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112. Itan, Y., Zhang, S.Y., Vogt, G., Abhyankar, A., Herman, M., Nitschke, P., Fried, D., Quintana-Murci, L., Abel, L., and Casanova, J.L. (2013). The human gene connectome as a map of short cuts for morbid allele discovery. Proc. Natl. Acad. Sci. USA 110, 5558-5563. https://doi.org/10.1073/pnas.1218167110.

113. Fourgeaud, J., Regnault, B., Ok, V., Da Rocha, N.D., Sitterlé, É., Mekouar, M., Faury, H., Milliancourt-Seels, C., Jagorel, F., Chrétien, D., et al. (2024). Performance of clinical metagenomics in France: a prospective observational study. Lancet Microbe 5, e52-e61. https://doi.org/10.1016/S2666-5247(23)00244-6.

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117. Pertea, M., Kim, D., Pertea, G.M., Leek, J.T., and Salzberg, S.L. (2016). Transcript-level expression analysis of RNA-seq experiments with HISAT, StringTie, and Ballgown. Nat. Protoc. 11, 1650-1667. https://doi.org/10.1038/nprot.2016.095.

118. Thorvaldsdóttir, H., Robinson, J.T., and Mesirov, J.P. (2013). Integrative Genomics Viewer (IGV): high-performance genomics data visualization and exploration. Brief. Bioinform. 14, 178-192. https://doi.org/10.1093/bib/bbs017.

119. Xu, G.J., Kula, T., Xu, Q., Li, M.Z., Vernon, S.D., Ndung'u, T., Ruxrungtham, K., Sanchez, J., Brander, C., Chung, R.T., et al. (2015). Viral immunology. Comprehensive serological profiling of human populations using a synthetic human virome. Science 348, aaa0698. https://doi.org/10.1126/science.aaa0698.

120. Waterboer, T., Sehr, P., Michael, K.M., Franceschi, S., Nieland, J.D., Joos, T.O., Templin, M.F., and Pawlita, M. (2005). Multiplex human papillomavirus serology based on in situ-purified glutathione S-transferase fusion proteins. Clin. Chem. 51, 1845-1853. https://doi.org/10.1373/clinchem.2005.052381.

121. Bunis, D.G., Bronevetsky, Y., Krow-Lucal, E., Bhakta, N.R., Kim, C.C., Nerella, S., Jones, N., Mendoza, V.F., Bryson, Y.J., Gern, J.E., et al. (2021). Single-cell mapping of progressive fetal-to-adult transition in human naive T cells. Cell Rep. 34, 108573. https://doi.org/10.1016/j.celrep.2020.108573.
<figure><img src="image_46.png" /></figure>

122. Martens, J.H.A., and Stunnenberg, H.G. (2013). BLUEPRINT: mapping human blood cell epigenomes. Haematologica 98, 1487-1489. https://doi.org/10.3324/haematol.2013.094243.

123. Hao, Y., Hao, S., Andersen-Nissen, E., Mauck, W.M., Zheng, S., Butler, A., Lee, M.J., Wilk, A.J., Darby, C., Zager, M., et al. (2021). Integrated analysis of multimodal single-cell data. Cell 184, 3573-3587.e29. https://doi.org/10.1016/j.cell.2021.04.048.

124. Sobrino, S., Magnani, A., Semeraro, M., Martignetti, L., Cortal, A., Denis, A., Couzin, C., Picard, C., Bustamante, J., Magrin, E., et al. (2023). Severe hematopoietic stem cell inflammation compromises chronic granulomatous disease gene therapy. Cell Rep. Med. 4, 100919. https://doi.org/10.1016/j.xcrm.2023.100919.

125. Windhager, J., Bodenmiller, B., and Eling, N. (2021). An End-to-End Workflow for Multiplexed Image Processing and Analysis. Preprint at bioRxiv. https://doi.org/10.1101/2021.11.12.468357.

126. Chevrier, S., Crowell, H.L., Zanotelli, V.R.T., Engler, S., Robinson, M.D., and Bodenmiller, B. (2018). Compensation of signal spillover in suspension and imaging mass cytometry. Cell Syst. 6, 612-620.e5. https://doi.org/10.1016/j.cels.2018.02.010.

127. Bankhead, P., Loughrey, M.B., Fernández, J.A., Dombrowski, Y., McArt, D.G., Dunne, P.D., McQuaid, S., Gray, R.T., Murray, L.J., Coleman, H.G., et al. (2017). QuPath: open source software for digital pathology image analysis. Sci. Rep. 7, 16878. https://doi.org/10.1038/s41598-017-17204-5.

128. Cladel, N.M., Budgeon, L.R., Balogh, K.K., Cooper, T.K., Hu, J., and Christensen, N.D. (2015). A novel pre-clinical murine model to study the life cycle and progression of cervical and anal papillomavirus infections. PLOS One 10, e0120128. https://doi.org/10.1371/journal.pone.0120128.

129. Cladel, N.M., Budgeon, L.R., Balogh, K.K., Cooper, T.K., Hu, J., and Christensen, N.D. (2016). Mouse papillomavirus MmuPV1 infects oral mucosa and preferentially targets the base of the tongue. Virology 488, 73-80. https://doi.org/10.1016/j.virol.2015.10.030.

130. Cladel, N.M., Jiang, P., Li, J.J., Peng, X., Cooper, T.K., Majerciak, V., Balogh, K.K., Meyer, T.J., Brendle, S.A., Budgeon, L.R., et al. (2019). Papillomavirus can be transmitted through the blood and produce infections in blood recipients: evidence from two animal models. Emerg. Microbes Infect. 8, 1108-1121. https://doi.org/10.1080/22221751.2019.1637072.

131. Cladel, N.M., Budgeon, L.R., Cooper, T.K., Balogh, K.K., Christensen, N.D., Myers, R., Majerciak, V., Gotte, D., Zheng, Z.-M., and Hu, J. (2017). Mouse papillomavirus infections spread to cutaneous sites with progression to malignancy. J. Gen. Virol. 98, 2520-2529. https://doi.org/10.1099/jgv.0.000926.
122. Martens, J.H.A., and Stunnenberg, H.G. (2013). BLUEPRINT: mapping human blood cell epigenomes. Haematologica 98, 1487-1489. https://doi.org/10.3324/haematol.2013.094243.

123. Hao, Y., Hao, S., Andersen-Nissen, E., Mauck, W.M., Zheng, S., Butler, A., Lee, M.J., Wilk, A.J., Darby, C., Zager, M., et al. (2021). Integrated analysis of multimodal single-cell data. Cell 184, 3573-3587.e29. https://doi.org/10.1016/j.cell.2021.04.048.

124. Sobrino, S., Magnani, A., Semeraro, M., Martignetti, L., Cortal, A., Denis, A., Couzin, C., Picard, C., Bustamante, J., Magrin, E., et al. (2023). Severe hematopoietic stem cell inflammation compromises chronic granulomatous disease gene therapy. Cell Rep. Med. 4, 100919. https://doi.org/10.1016/j.xcrm.2023.100919.

125. Windhager, J., Bodenmiller, B., and Eling, N. (2021). An End-to-End Workflow for Multiplexed Image Processing and Analysis. Preprint at bioRxiv. https://doi.org/10.1101/2021.11.12.468357.

126. Chevrier, S., Crowell, H.L., Zanotelli, V.R.T., Engler, S., Robinson, M.D., and Bodenmiller, B. (2018). Compensation of signal spillover in suspension and imaging mass cytometry. Cell Syst. 6, 612-620.e5. https://doi.org/10.1016/j.cels.2018.02.010.

127. Bankhead, P., Loughrey, M.B., Fernández, J.A., Dombrowski, Y., McArt, D.G., Dunne, P.D., McQuaid, S., Gray, R.T., Murray, L.J., Coleman, H.G., et al. (2017). QuPath: open source software for digital pathology image analysis. Sci. Rep. 7, 16878. https://doi.org/10.1038/s41598-017-17204-5.

128. Cladel, N.M., Budgeon, L.R., Balogh, K.K., Cooper, T.K., Hu, J., and Christensen, N.D. (2015). A novel pre-clinical murine model to study the life cycle and progression of cervical and anal papillomavirus infections. PLOS One 10, e0120128. https://doi.org/10.1371/journal.pone.0120128.

129. Cladel, N.M., Budgeon, L.R., Balogh, K.K., Cooper, T.K., Hu, J., and Christensen, N.D. (2016). Mouse papillomavirus MmuPV1 infects oral mucosa and preferentially targets the base of the tongue. Virology 488, 73-80. https://doi.org/10.1016/j.virol.2015.10.030.

130. Cladel, N.M., Jiang, P., Li, J.J., Peng, X., Cooper, T.K., Majerciak, V., Balogh, K.K., Meyer, T.J., Brendle, S.A., Budgeon, L.R., et al. (2019). Papillomavirus can be transmitted through the blood and produce infections in blood recipients: evidence from two animal models. Emerg. Microbes Infect. 8, 1108-1121. https://doi.org/10.1080/22221751.2019.1637072.

131. Cladel, N.M., Budgeon, L.R., Cooper, T.K., Balogh, K.K., Christensen, N.D., Myers, R., Majerciak, V., Gotte, D., Zheng, Z.-M., and Hu, J. (2017). Mouse papillomavirus infections spread to cutaneous sites with progression to malignancy. J. Gen. Virol. 98, 2520-2529. https://doi.org/10.1099/jgv.0.000926.
## STAR★METHODS

### KEY RESOURCES TABLE

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td colspan="3">Antibodies</td></tr><tr><td>anti-Flt 3-L antibody (F-6) HRP</td><td>Santa Cruz Biotechnology</td><td>Cat# sc-365266 HRP; RRID:AB_10848274</td></tr><tr><td>anti-human Flt-3 ligand/FLT3L antibody</td><td>R&amp;D Systems</td><td>Cat# MAB308; RRID:AB_2104978</td></tr><tr><td>rabbit anti-FLT3 (8F2) mAb</td><td>Cell Signaling</td><td>Cat# 3462; RRID:AB_2107052</td></tr><tr><td>rabbit anti-phospho-FLT3 (Tyr589/591) (30D4) mAb</td><td>Cell Signaling</td><td>Cat# 3464; RRID:AB_2107051</td></tr><tr><td>anti-V5 tag antibody</td><td>Thermo Fisher Scientific</td><td>Cat# R962-25; RRID:AB_2556566</td></tr><tr><td>anti-vinculin antibody HRP</td><td>Santa Cruz Biotechnology</td><td>Cat# sc-73614 HRP; RRID:AB_2941767</td></tr><tr><td>goat anti-mouse IgG (H + L)-HRP conjugate</td><td>Bio-Rad</td><td>Cat# 1706516; RRID:AB_2921252</td></tr><tr><td>goat anti-rabbit IgG (H + L)-HRP conjugate</td><td>Bio-Rad</td><td>Cat# 170-6515 (also 1706515); RRID:AB_11125142</td></tr><tr><td>recombinant anti-Flt3 ligand [EP1140Y]</td><td>Abcam</td><td>Cat# ab52648; RRID:AB_2104974</td></tr><tr><td>recombinant rabbit IgG, monoclonal [EPR25A]</td><td>Abcam</td><td>Cat# ab172730; RRID:AB_2687931</td></tr><tr><td>PE-conjugated goat anti-rabbit IgG (H+L) secondary antibody</td><td>Thermo Fisher Scientific</td><td>Cat# A10542; RRID:AB_1500781</td></tr><tr><td>163Dy CXCR3</td><td>Standard BioTools</td><td>Cat# 3163004B; RRID:AB_2810969</td></tr><tr><td>152Sm TCRgd</td><td>Standard BioTools</td><td>Cat# 3152008B; RRID:AB_2687643</td></tr><tr><td>142Nd CD19</td><td>Standard BioTools</td><td>Cat# 3142001 (also 3142001B); RRID:AB_2651155</td></tr><tr><td>144 ND CD38</td><td>Standard BioTools</td><td>Cat# 3144014B; RRID:AB_2687640</td></tr><tr><td>143 ND CD123</td><td>Standard BioTools</td><td>Cat# 3151001 (also 3151001B); RRID:AB_2661794</td></tr><tr><td>153Eu Va7.2</td><td>Standard BioTools</td><td>Cat# 3153024B; RRID:AB_2891190</td></tr><tr><td>154Sm CD3</td><td>Standard BioTools</td><td>Cat# 3154003B; RRID:AB_2811086</td></tr><tr><td>155Gd CD45RA</td><td>Standard BioTools</td><td>Cat# 3155011B; RRID:AB_2810246</td></tr><tr><td>158Gd CD27</td><td>Standard BioTools</td><td>Cat# 3158010B; RRID:AB_2858231</td></tr><tr><td>159Tb CD1c</td><td>Biolegend</td><td>Cat# 331502 (also 331501); RRID:AB_1088995</td></tr><tr><td>161Dy CLEC9A</td><td>Standard BioTools</td><td>Cat# 3161018B; RRID:AB_2810252</td></tr><tr><td>164Dy CD161</td><td>Standard BioTools</td><td>Cat# 3164009B; RRID:AB_2687651</td></tr><tr><td>168Er CD8</td><td>Standard BioTools</td><td>Cat# 3168002B; RRID:AB_2892771</td></tr><tr><td>170Er iNKT</td><td>Standard BioTools</td><td>Cat# 3170015B; RRID: N/A</td></tr><tr><td>175Lu CCR4</td><td>Standard BioTools</td><td>Cat# 3175035A; RRID:AB_2921320</td></tr><tr><td>174Yb CD4</td><td>Biolegend</td><td>Cat# 300501 (also 300502); RRID:AB_314069</td></tr><tr><td>162Dy CD21</td><td>Miltenyi Biotec</td><td>Cat# 130-124-315; RRID:AB_2811646</td></tr><tr><td>165Ho NKG2C</td><td>Miltenyi Biotec</td><td>Cat# 130-122-278; RRID:AB_2801859</td></tr><tr><td>148Nd CD20</td><td>Biolegend</td><td>Cat# 302302 (also 302301); RRID:AB_314250</td></tr><tr><td>173Yb HLA-DR</td><td>Standard BioTools</td><td>Cat# 3173005B; RRID:AB_2810248</td></tr><tr><td>156Gd CCR10</td><td>Miltenyi Biotec Inc.</td><td>Cat# 130-122-317; RRID:AB_2801898</td></tr><tr><td>089Y CD45</td><td>Standard BioTools</td><td>Cat# 3089003B; RRID:AB_2938863</td></tr><tr><td>116Cd CD66b</td><td>Biolegend</td><td>Cat# 396902; RRID:AB_2814367</td></tr><tr><td>141Pr CCR6</td><td>Standard BioTools</td><td>Cat# 3141003A; RRID:AB_2687639</td></tr><tr><td>158Gd CD127</td><td>Standard BioTools</td><td>Cat# 3143012B; RRID:AB_2810240</td></tr><tr><td>147Sm CD11c</td><td>Standard BioTools</td><td>Cat# 3147008 (also 3147008B); RRID:AB_2687850</td></tr><tr><td>149Sm CD25</td><td>Standard BioTools</td><td>Cat# 3149010B; RRID:AB_2756416</td></tr><tr><td>150Nd NKVFS1</td><td>Bio Rad</td><td>Cat# MCA2243GA; RRID:AB_323743</td></tr><tr><td>167Er CCR7</td><td>Standard BioTools</td><td>Cat# 3167009A; RRID:AB_2858236</td></tr><tr><td>169Tm NKG2A</td><td>Standard BioTools</td><td>Cat# 3169013B; RRID:AB_2756426</td></tr><tr><td>0171Yb CXCR5</td><td>Standard BioTools</td><td>Cat# 3171014B; RRID:AB_2858239</td></tr></tbody></table>

(Continued on next page)
## STAR★METHODS

### KEY RESOURCES TABLE

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td colspan="3">Antibodies</td></tr><tr><td>anti-Flt 3-L antibody (F-6) HRP</td><td>Santa Cruz Biotechnology</td><td>Cat# sc-365266 HRP; RRID:AB_10848274</td></tr><tr><td>anti-human Flt-3 ligand/FLT3L antibody</td><td>R&amp;D Systems</td><td>Cat# MAB308; RRID:AB_2104978</td></tr><tr><td>rabbit anti-FLT3 (8F2) mAb</td><td>Cell Signaling</td><td>Cat# 3462; RRID:AB_2107052</td></tr><tr><td>rabbit anti-phospho-FLT3 (Tyr589/591) (30D4) mAb</td><td>Cell Signaling</td><td>Cat# 3464; RRID:AB_2107051</td></tr><tr><td>anti-V5 tag antibody</td><td>Thermo Fisher Scientific</td><td>Cat# R962-25; RRID:AB_2556566</td></tr><tr><td>anti-vinculin antibody HRP</td><td>Santa Cruz Biotechnology</td><td>Cat# sc-73614 HRP; RRID:AB_2941767</td></tr><tr><td>goat anti-mouse IgG (H + L)-HRP conjugate</td><td>Bio-Rad</td><td>Cat# 1706516; RRID:AB_2921252</td></tr><tr><td>goat anti-rabbit IgG (H + L)-HRP conjugate</td><td>Bio-Rad</td><td>Cat# 170-6515 (also 1706515); RRID:AB_11125142</td></tr><tr><td>recombinant anti-Flt3 ligand [EP1140Y]</td><td>Abcam</td><td>Cat# ab52648; RRID:AB_2104974</td></tr><tr><td>recombinant rabbit IgG, monoclonal [EPR25A]</td><td>Abcam</td><td>Cat# ab172730; RRID:AB_2687931</td></tr><tr><td>PE-conjugated goat anti-rabbit IgG (H+L) secondary antibody</td><td>Thermo Fisher Scientific</td><td>Cat# A10542; RRID:AB_1500781</td></tr><tr><td>163Dy CXCR3</td><td>Standard BioTools</td><td>Cat# 3163004B; RRID:AB_2810969</td></tr><tr><td>152Sm TCRgd</td><td>Standard BioTools</td><td>Cat# 3152008B; RRID:AB_2687643</td></tr><tr><td>142Nd CD19</td><td>Standard BioTools</td><td>Cat# 3142001 (also 3142001B); RRID:AB_2651155</td></tr><tr><td>144 ND CD38</td><td>Standard BioTools</td><td>Cat# 3144014B; RRID:AB_2687640</td></tr><tr><td>143 ND CD123</td><td>Standard BioTools</td><td>Cat# 3151001 (also 3151001B); RRID:AB_2661794</td></tr><tr><td>153Eu Va7.2</td><td>Standard BioTools</td><td>Cat# 3153024B; RRID:AB_2891190</td></tr><tr><td>154Sm CD3</td><td>Standard BioTools</td><td>Cat# 3154003B; RRID:AB_2811086</td></tr><tr><td>155Gd CD45RA</td><td>Standard BioTools</td><td>Cat# 3155011B; RRID:AB_2810246</td></tr><tr><td>158Gd CD27</td><td>Standard BioTools</td><td>Cat# 3158010B; RRID:AB_2858231</td></tr><tr><td>159Tb CD1c</td><td>Biolegend</td><td>Cat# 331502 (also 331501); RRID:AB_1088995</td></tr><tr><td>161Dy CLEC9A</td><td>Standard BioTools</td><td>Cat# 3161018B; RRID:AB_2810252</td></tr><tr><td>164Dy CD161</td><td>Standard BioTools</td><td>Cat# 3164009B; RRID:AB_2687651</td></tr><tr><td>168Er CD8</td><td>Standard BioTools</td><td>Cat# 3168002B; RRID:AB_2892771</td></tr><tr><td>170Er iNKT</td><td>Standard BioTools</td><td>Cat# 3170015B; RRID: N/A</td></tr><tr><td>175Lu CCR4</td><td>Standard BioTools</td><td>Cat# 3175035A; RRID:AB_2921320</td></tr><tr><td>174Yb CD4</td><td>Biolegend</td><td>Cat# 300501 (also 300502); RRID:AB_314069</td></tr><tr><td>162Dy CD21</td><td>Miltenyi Biotec</td><td>Cat# 130-124-315; RRID:AB_2811646</td></tr><tr><td>165Ho NKG2C</td><td>Miltenyi Biotec</td><td>Cat# 130-122-278; RRID:AB_2801859</td></tr><tr><td>148Nd CD20</td><td>Biolegend</td><td>Cat# 302302 (also 302301); RRID:AB_314250</td></tr><tr><td>173Yb HLA-DR</td><td>Standard BioTools</td><td>Cat# 3173005B; RRID:AB_2810248</td></tr><tr><td>156Gd CCR10</td><td>Miltenyi Biotec Inc.</td><td>Cat# 130-122-317; RRID:AB_2801898</td></tr><tr><td>089Y CD45</td><td>Standard BioTools</td><td>Cat# 3089003B; RRID:AB_2938863</td></tr><tr><td>116Cd CD66b</td><td>Biolegend</td><td>Cat# 396902; RRID:AB_2814367</td></tr><tr><td>141Pr CCR6</td><td>Standard BioTools</td><td>Cat# 3141003A; RRID:AB_2687639</td></tr><tr><td>158Gd CD127</td><td>Standard BioTools</td><td>Cat# 3143012B; RRID:AB_2810240</td></tr><tr><td>147Sm CD11c</td><td>Standard BioTools</td><td>Cat# 3147008 (also 3147008B); RRID:AB_2687850</td></tr><tr><td>149Sm CD25</td><td>Standard BioTools</td><td>Cat# 3149010B; RRID:AB_2756416</td></tr><tr><td>150Nd NKVFS1</td><td>Bio Rad</td><td>Cat# MCA2243GA; RRID:AB_323743</td></tr><tr><td>167Er CCR7</td><td>Standard BioTools</td><td>Cat# 3167009A; RRID:AB_2858236</td></tr><tr><td>169Tm NKG2A</td><td>Standard BioTools</td><td>Cat# 3169013B; RRID:AB_2756426</td></tr><tr><td>0171Yb CXCR5</td><td>Standard BioTools</td><td>Cat# 3171014B; RRID:AB_2858239</td></tr></tbody></table>

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Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>166Er CD24</td><td>Standard BioTools</td><td>Cat# 3166007B; RRID:AB_2938867</td></tr><tr><td>145ND CD31</td><td>Standard BioTools</td><td>Cat# 3145004B; RRID:AB_2737262</td></tr><tr><td>160Gd CD14</td><td>Standard BioTools</td><td>Cat# 3160001B; RRID:AB_2687634</td></tr><tr><td>176Yb CD56</td><td>Standard BioTools</td><td>Cat# 3176008B; RRID:AB_2938870</td></tr><tr><td>172Yb CD57</td><td>Biolegend</td><td>Cat# 359602; RRID:AB_2562403</td></tr><tr><td>150Nd KIR3DL1L2</td><td>Miltenyi Biotec Inc.</td><td>Cat# 130-126-489; RRID:AB_2889458</td></tr><tr><td>146Nd IgD</td><td>Standard BioTools</td><td>Cat# 3146005B; RRID:AB_2811082</td></tr><tr><td>209Bi CD16</td><td>Standard BioTools</td><td>Cat# 3209002B; RRID:AB_2756431</td></tr><tr><td>CD45 PerCP-Cy5.5</td><td>BD Biosciences</td><td>Cat# 345809; RRID:AB_2868830</td></tr><tr><td>CD19 APC</td><td>BD Biosciences</td><td>Cat# 345791; RRID:AB_2868817</td></tr><tr><td>CD3 FITC</td><td>BD Biosciences</td><td>Cat# 345764; RRID:AB_2916364</td></tr><tr><td>CD16 PE</td><td>BD Biosciences</td><td>Cat# 332779; RRID:AB_2868628</td></tr><tr><td>CD56 PE</td><td>BD Biosciences</td><td>Cat# 345812; RRID:AB_2629216</td></tr><tr><td>CD4 APC</td><td>BD Biosciences</td><td>Cat# 345771; RRID:AB_2868799</td></tr><tr><td>CD8 PE</td><td>BD Biosciences</td><td>Cat# 345773; RRID:AB_2868801</td></tr><tr><td>CD3 PE</td><td>BD Biosciences</td><td>Cat# 345765; RRID:AB_2868796</td></tr><tr><td>CD4 FITC</td><td>BD Biosciences</td><td>Cat# 345768; RRID:AB_2868797</td></tr><tr><td>CD25 APC</td><td>BD Biosciences</td><td>Cat# 340907; RRID:AB_2819021</td></tr><tr><td>CD127 PE</td><td>Thermo Fisher Scientific</td><td>Cat# 12-1278-42; RRID:AB_10717663</td></tr><tr><td>CD45 V500</td><td>BD Biosciences</td><td>Cat# 655873; RRID:AB_2870390</td></tr><tr><td>Lin1 FITC</td><td>BD Biosciences</td><td>Cat# 340546; RRID:AB_400053</td></tr><tr><td>Anti-HLA-DR PerCP</td><td>BD Biosciences</td><td>Cat# 347402; RRID:AB_2868847</td></tr><tr><td>CD123 PE</td><td>BD Biosciences</td><td>Cat# 340545; RRID:AB_400052</td></tr><tr><td>CD11c APC</td><td>BD Biosciences</td><td>Cat# 333144; RRID:AB_2868645</td></tr><tr><td>CD38 V450</td><td>BD Biosciences</td><td>Cat# 646851; RRID:AB_1937282</td></tr><tr><td>CD19 PE</td><td>BD Biosciences</td><td>Cat# 345777; RRID:AB_2868805</td></tr><tr><td>CD20 PerCP-Cy5.5</td><td>BD Biosciences</td><td>Cat# 345794; RRID:AB_2868820</td></tr><tr><td>CD10 FITC</td><td>BD Biosciences</td><td>Cat# 332775; RRID:AB_2868624</td></tr><tr><td>CD27 APC</td><td>BD Biosciences</td><td>Cat# 337169; RRID:AB_647368</td></tr><tr><td>CD21 PE</td><td>BD Biosciences</td><td>Cat# 555422; RRID:AB_395816</td></tr><tr><td>CD3 APC-R700</td><td>BD Biosciences</td><td>Cat# 659119; RRID:AB_2870468</td></tr><tr><td>CD4 APC-H7</td><td>BD Biosciences</td><td>Cat# 641398; RRID:AB_1645732</td></tr><tr><td>CD127 PE-Cy7</td><td>BD Biosciences</td><td>Cat# 560822; RRID:AB_2033938</td></tr><tr><td>CD45RA FITC</td><td>BD Biosciences</td><td>Cat# 335039; RRID:AB_2868682</td></tr><tr><td>CD185 (CXCR5) PerCP-Cy5.5</td><td>BD Biosciences</td><td>Cat# 562781; RRID:AB_2313576</td></tr><tr><td>CD183 (CXCR3) PE</td><td>BD Biosciences</td><td>Cat# 557185; RRID:AB_396596</td></tr><tr><td>CD196 (CCR6) BV421</td><td>BD Biosciences</td><td>Cat# 562515; RRID:AB_11154229</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD3</td><td>BD Biosciences</td><td>Cat# 555332; RRID:AB_395739</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD19</td><td>BD Biosciences</td><td>Cat# 555412; RRID:AB_395812</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD20</td><td>BD Biosciences</td><td>Cat# 560962; RRID:AB_2033953</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD14</td><td>BD Biosciences</td><td>Cat# 555397; RRID:AB_395798</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD16</td><td>BD Biosciences</td><td>Cat# 555406; RRID:AB_395806</td></tr><tr><td>BD™ CD56 FITC</td><td>BD Biosciences</td><td>Cat# 345811; RRID:AB_2868832</td></tr><tr><td>APC Mouse Anti-Human CD11C</td><td>BD Biosciences</td><td>Cat# 559877; RRID:AB_398680</td></tr><tr><td>BD Pharmingen™ PE Mouse Anti-Human HLA-DR</td><td>BD Biosciences</td><td>Cat# 555812; RRID:AB_396146</td></tr><tr><td>BD Pharmingen™ Pacific Blue™ Mouse Anti-Human CD14</td><td>BD Biosciences</td><td>Cat# 558121; RRID:AB_397041</td></tr><tr><td>BD Horizon™ BV650 Mouse Anti-Human CD16</td><td>BD Biosciences</td><td>Cat# 563692; RRID:AB_2869511</td></tr></tbody></table>

(Continued on next page)
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<table><thead><tr><td>REAGENT or RESOURCE</td><td>SOURCE</td><td>IDENTIFIER</td></tr></thead><tbody><tr><td>166Er CD24</td><td>Standard BioTools</td><td>Cat# 3166007B; RRID:AB_2938867</td></tr><tr><td>145ND CD31</td><td>Standard BioTools</td><td>Cat# 3145004B; RRID:AB_2737262</td></tr><tr><td>160Gd CD14</td><td>Standard BioTools</td><td>Cat# 3160001B; RRID:AB_2687634</td></tr><tr><td>176Yb CD56</td><td>Standard BioTools</td><td>Cat# 3176008B; RRID:AB_2938870</td></tr><tr><td>172Yb CD57</td><td>Biolegend</td><td>Cat# 359602; RRID:AB_2562403</td></tr><tr><td>150Nd KIR3DL1L2</td><td>Miltenyi Biotec Inc.</td><td>Cat# 130-126-489; RRID:AB_2889458</td></tr><tr><td>146Nd IgD</td><td>Standard BioTools</td><td>Cat# 3146005B; RRID:AB_2811082</td></tr><tr><td>209Bi CD16</td><td>Standard BioTools</td><td>Cat# 3209002B; RRID:AB_2756431</td></tr><tr><td>CD45 PerCP-Cy5.5</td><td>BD Biosciences</td><td>Cat# 345809; RRID:AB_2868830</td></tr><tr><td>CD19 APC</td><td>BD Biosciences</td><td>Cat# 345791; RRID:AB_2868817</td></tr><tr><td>CD3 FITC</td><td>BD Biosciences</td><td>Cat# 345764; RRID:AB_2916364</td></tr><tr><td>CD16 PE</td><td>BD Biosciences</td><td>Cat# 332779; RRID:AB_2868628</td></tr><tr><td>CD56 PE</td><td>BD Biosciences</td><td>Cat# 345812; RRID:AB_2629216</td></tr><tr><td>CD4 APC</td><td>BD Biosciences</td><td>Cat# 345771; RRID:AB_2868799</td></tr><tr><td>CD8 PE</td><td>BD Biosciences</td><td>Cat# 345773; RRID:AB_2868801</td></tr><tr><td>CD3 PE</td><td>BD Biosciences</td><td>Cat# 345765; RRID:AB_2868796</td></tr><tr><td>CD4 FITC</td><td>BD Biosciences</td><td>Cat# 345768; RRID:AB_2868797</td></tr><tr><td>CD25 APC</td><td>BD Biosciences</td><td>Cat# 340907; RRID:AB_2819021</td></tr><tr><td>CD127 PE</td><td>Thermo Fisher Scientific</td><td>Cat# 12-1278-42; RRID:AB_10717663</td></tr><tr><td>CD45 V500</td><td>BD Biosciences</td><td>Cat# 655873; RRID:AB_2870390</td></tr><tr><td>Lin1 FITC</td><td>BD Biosciences</td><td>Cat# 340546; RRID:AB_400053</td></tr><tr><td>Anti-HLA-DR PerCP</td><td>BD Biosciences</td><td>Cat# 347402; RRID:AB_2868847</td></tr><tr><td>CD123 PE</td><td>BD Biosciences</td><td>Cat# 340545; RRID:AB_400052</td></tr><tr><td>CD11c APC</td><td>BD Biosciences</td><td>Cat# 333144; RRID:AB_2868645</td></tr><tr><td>CD38 V450</td><td>BD Biosciences</td><td>Cat# 646851; RRID:AB_1937282</td></tr><tr><td>CD19 PE</td><td>BD Biosciences</td><td>Cat# 345777; RRID:AB_2868805</td></tr><tr><td>CD20 PerCP-Cy5.5</td><td>BD Biosciences</td><td>Cat# 345794; RRID:AB_2868820</td></tr><tr><td>CD10 FITC</td><td>BD Biosciences</td><td>Cat# 332775; RRID:AB_2868624</td></tr><tr><td>CD27 APC</td><td>BD Biosciences</td><td>Cat# 337169; RRID:AB_647368</td></tr><tr><td>CD21 PE</td><td>BD Biosciences</td><td>Cat# 555422; RRID:AB_395816</td></tr><tr><td>CD3 APC-R700</td><td>BD Biosciences</td><td>Cat# 659119; RRID:AB_2870468</td></tr><tr><td>CD4 APC-H7</td><td>BD Biosciences</td><td>Cat# 641398; RRID:AB_1645732</td></tr><tr><td>CD127 PE-Cy7</td><td>BD Biosciences</td><td>Cat# 560822; RRID:AB_2033938</td></tr><tr><td>CD45RA FITC</td><td>BD Biosciences</td><td>Cat# 335039; RRID:AB_2868682</td></tr><tr><td>CD185 (CXCR5) PerCP-Cy5.5</td><td>BD Biosciences</td><td>Cat# 562781; RRID:AB_2313576</td></tr><tr><td>CD183 (CXCR3) PE</td><td>BD Biosciences</td><td>Cat# 557185; RRID:AB_396596</td></tr><tr><td>CD196 (CCR6) BV421</td><td>BD Biosciences</td><td>Cat# 562515; RRID:AB_11154229</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD3</td><td>BD Biosciences</td><td>Cat# 555332; RRID:AB_395739</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD19</td><td>BD Biosciences</td><td>Cat# 555412; RRID:AB_395812</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD20</td><td>BD Biosciences</td><td>Cat# 560962; RRID:AB_2033953</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD14</td><td>BD Biosciences</td><td>Cat# 555397; RRID:AB_395798</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD16</td><td>BD Biosciences</td><td>Cat# 555406; RRID:AB_395806</td></tr><tr><td>BD™ CD56 FITC</td><td>BD Biosciences</td><td>Cat# 345811; RRID:AB_2868832</td></tr><tr><td>APC Mouse Anti-Human CD11C</td><td>BD Biosciences</td><td>Cat# 559877; RRID:AB_398680</td></tr><tr><td>BD Pharmingen™ PE Mouse Anti-Human HLA-DR</td><td>BD Biosciences</td><td>Cat# 555812; RRID:AB_396146</td></tr><tr><td>BD Pharmingen™ Pacific Blue™ Mouse Anti-Human CD14</td><td>BD Biosciences</td><td>Cat# 558121; RRID:AB_397041</td></tr><tr><td>BD Horizon™ BV650 Mouse Anti-Human CD16</td><td>BD Biosciences</td><td>Cat# 563692; RRID:AB_2869511</td></tr></tbody></table>

(Continued on next page)
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<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>Brilliant Violet 605™ anti-human CD45 Antibody</td><td>BioLegend</td><td>Cat# 368523 (also 368524); RRID:AB_2715825</td></tr><tr><td>Pacific Blue™ anti-human HLA-DR Antibody</td><td>BioLegend</td><td>Cat# 307633 (also 307623, 307624); RRID:AB_1595444</td></tr><tr><td>APC/Cyanine7 anti-human CD1c Antibody</td><td>BioLegend</td><td>Cat# 331519 (also 331520); RRID:AB_10643413</td></tr><tr><td>BD Horizon™ BV711 Mouse Anti-Human CD141</td><td>BD Biosciences</td><td>Cat# 563155; RRID:AB_2738033</td></tr><tr><td>PE anti-human CD370 (CLEC9A/DNGR1) Antibody</td><td>BioLegend</td><td>Cat# 353803 (also 353804); RRID:AB_10959651</td></tr><tr><td>PE/Cyanine7 anti-human CD123 Antibody</td><td>BioLegend</td><td>Cat# 306009 (also 306010); RRID:AB_493577</td></tr><tr><td>APC anti-human CD34 Antibody</td><td>BioLegend</td><td>Cat# 343608 (also 343607); RRID:AB_2228972</td></tr><tr><td>BD Pharmingen™ APC-H7 Mouse Anti-Human CD45RA</td><td>BD Biosciences</td><td>Cat# 560674; RRID:AB_1727497</td></tr><tr><td>Pacific Blue™ anti-human CD123 Antibody</td><td>BioLegend</td><td>Cat# 306044 (also 306043); RRID:AB_2750165</td></tr><tr><td>PE anti-human CD10 Antibody</td><td>BioLegend</td><td>Cat# 312204 (also 312203); RRID:AB_314915</td></tr><tr><td>CD38 Antibody, anti-human, PE-Vio® 770</td><td>Miltenyi Biotec</td><td>Cat# 130-113-428; RRID:AB_2733230</td></tr><tr><td>Brilliant Violet 711™ anti-human CD45RA Antibody</td><td>BioLegend</td><td>Cat# 304138 (also 304137); RRID:AB_11218999</td></tr><tr><td>PE/Cyanine5 anti-human CD90 (Thy1) Antibody</td><td>BioLegend</td><td>Cat# 328112 (also 328111); RRID:AB_893432</td></tr><tr><td>Brilliant Violet 605™ anti-human CD38 Antibody</td><td>BioLegend</td><td>Cat# 303532 (also 303531); RRID:AB_2562915</td></tr><tr><td>APC/Cyanine7 anti-human CD34 Antibody</td><td>BioLegend</td><td>Cat# 343514 (also 343513); RRID:AB_1877168</td></tr><tr><td>PE/Dazzle™ 594 anti-human CD133 Antibody</td><td>BioLegend</td><td>Cat# 372812 (also 372811); RRID:AB_2650878</td></tr><tr><td>PE/Cyanine7 anti-human CD123 Antibody</td><td>BioLegend</td><td>Cat# 306010 (also 306009); RRID:AB_493576</td></tr><tr><td>Brilliant Violet 510™ anti-human CD10 Antibody</td><td>BioLegend</td><td>Cat# 312220 (also 312219); RRID:AB_2563835</td></tr><tr><td>BD OptiBuild™ BV421 Mouse Anti-Human CD110</td><td>BD Biosciences</td><td>Cat# 743576; RRID:AB_2741598</td></tr><tr><td>PE conjugated Lin custom panel</td><td>Miltenyi Biotec</td><td>RRID: N/A</td></tr><tr><td>Brilliant Violet 570™ anti-human CD45</td><td>Sony</td><td>Cat# 2120170; RRID: N/A</td></tr><tr><td>Brilliant Violet 510™ anti-human CD20 Antibody</td><td>BioLegend</td><td>Cat# 302339 (also 302340); RRID:AB_2561721</td></tr><tr><td>PE/Cy7 anti-human CD34</td><td>Sony</td><td>Cat# 2317580; RRID: N/A</td></tr><tr><td>APC/Cy7 anti-human CD19</td><td>Sony</td><td>Cat# 2111090; RRID: N/A</td></tr><tr><td>BD™ CD10 APC</td><td>BD Biosciences</td><td>Cat# 332777; RRID:AB_2868626</td></tr><tr><td>BD Horizon™ BV421 Mouse Anti-Human IgM</td><td>BD Biosciences</td><td>Cat# 562618; RRID:AB_2737681</td></tr><tr><td>BD OptiBuild™ BV711 Mouse Anti-Human IgD</td><td>BD Biosciences</td><td>Cat# 740794; RRID:AB_2740457</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD24</td><td>BD Biosciences</td><td>Cat# 555427; RRID:AB_395821</td></tr><tr><td>BD Pharmingen™ PE Mouse Anti-Human CD38</td><td>BD Biosciences</td><td>Cat# 555460; RRID:AB_395853</td></tr><tr><td>Monoclonal Mouse Anti-Human Glycophorin C (Concentrate)</td><td>Agilent</td><td>Cat# M082001; RRID: N/A</td></tr><tr><td>Polyclonal Rabbit Anti-Human Myeloperoxidase</td><td>Agilent</td><td>Cat# A039829; RRID: N/A</td></tr><tr><td>Polyclonal Rabbit Anti-Human Von Willebrand Factor</td><td>Agilent</td><td>Cat# A0082 (also A008202-5, A008202-2); RRID:AB_2315602</td></tr><tr><td>PE anti-human CD45 Antibody</td><td>BioLegend</td><td>Cat# 304008 (also 304007, 304039, 304058); RRID:AB_314396</td></tr><tr><td>APC anti-human HLA-A,B,C Antibody</td><td>BioLegend</td><td>Cat# 311409 (also 311410); RRID:AB_314878</td></tr><tr><td>CD3 Monoclonal Antibody (UCHT1), Alexa Fluor™ 532, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 58-0038-42; RRID:AB_11218675</td></tr><tr><td>TCR gamma/delta Monoclonal Antibody (B1.1), FITC, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 11-9959-41; RRID:AB_10669048</td></tr><tr><td>APC/Fire™ 750 anti-human TCR Vδ2 Antibody</td><td>BioLegend</td><td>Cat# 331419; RRID:AB_2687325</td></tr><tr><td>Brilliant Violet 605™ anti-human CD56 (NCAM) Antibody</td><td>BioLegend</td><td>Cat# 362537; RRID:AB_2565856</td></tr><tr><td>BD Horizon™ BV750 Mouse Anti-Human CD4</td><td>BD Biosciences</td><td>Cat# 566356; RRID:AB_2744426</td></tr><tr><td>Pacific Blue™ anti-human CD8 Antibody</td><td>BioLegend</td><td>Cat# 344717; RRID:AB_10551438</td></tr><tr><td>APC anti-human TCR Vα7.2 Antibody</td><td>BioLegend</td><td>Cat# 351708; RRID:AB_10933430</td></tr></tbody></table>

(Continued on next page)
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<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>Brilliant Violet 605™ anti-human CD45 Antibody</td><td>BioLegend</td><td>Cat# 368523 (also 368524); RRID:AB_2715825</td></tr><tr><td>Pacific Blue™ anti-human HLA-DR Antibody</td><td>BioLegend</td><td>Cat# 307633 (also 307623, 307624); RRID:AB_1595444</td></tr><tr><td>APC/Cyanine7 anti-human CD1c Antibody</td><td>BioLegend</td><td>Cat# 331519 (also 331520); RRID:AB_10643413</td></tr><tr><td>BD Horizon™ BV711 Mouse Anti-Human CD141</td><td>BD Biosciences</td><td>Cat# 563155; RRID:AB_2738033</td></tr><tr><td>PE anti-human CD370 (CLEC9A/DNGR1) Antibody</td><td>BioLegend</td><td>Cat# 353803 (also 353804); RRID:AB_10959651</td></tr><tr><td>PE/Cyanine7 anti-human CD123 Antibody</td><td>BioLegend</td><td>Cat# 306009 (also 306010); RRID:AB_493577</td></tr><tr><td>APC anti-human CD34 Antibody</td><td>BioLegend</td><td>Cat# 343608 (also 343607); RRID:AB_2228972</td></tr><tr><td>BD Pharmingen™ APC-H7 Mouse Anti-Human CD45RA</td><td>BD Biosciences</td><td>Cat# 560674; RRID:AB_1727497</td></tr><tr><td>Pacific Blue™ anti-human CD123 Antibody</td><td>BioLegend</td><td>Cat# 306044 (also 306043); RRID:AB_2750165</td></tr><tr><td>PE anti-human CD10 Antibody</td><td>BioLegend</td><td>Cat# 312204 (also 312203); RRID:AB_314915</td></tr><tr><td>CD38 Antibody, anti-human, PE-Vio® 770</td><td>Miltenyi Biotec</td><td>Cat# 130-113-428; RRID:AB_2733230</td></tr><tr><td>Brilliant Violet 711™ anti-human CD45RA Antibody</td><td>BioLegend</td><td>Cat# 304138 (also 304137); RRID:AB_11218999</td></tr><tr><td>PE/Cyanine5 anti-human CD90 (Thy1) Antibody</td><td>BioLegend</td><td>Cat# 328112 (also 328111); RRID:AB_893432</td></tr><tr><td>Brilliant Violet 605™ anti-human CD38 Antibody</td><td>BioLegend</td><td>Cat# 303532 (also 303531); RRID:AB_2562915</td></tr><tr><td>APC/Cyanine7 anti-human CD34 Antibody</td><td>BioLegend</td><td>Cat# 343514 (also 343513); RRID:AB_1877168</td></tr><tr><td>PE/Dazzle™ 594 anti-human CD133 Antibody</td><td>BioLegend</td><td>Cat# 372812 (also 372811); RRID:AB_2650878</td></tr><tr><td>PE/Cyanine7 anti-human CD123 Antibody</td><td>BioLegend</td><td>Cat# 306010 (also 306009); RRID:AB_493576</td></tr><tr><td>Brilliant Violet 510™ anti-human CD10 Antibody</td><td>BioLegend</td><td>Cat# 312220 (also 312219); RRID:AB_2563835</td></tr><tr><td>BD OptiBuild™ BV421 Mouse Anti-Human CD110</td><td>BD Biosciences</td><td>Cat# 743576; RRID:AB_2741598</td></tr><tr><td>PE conjugated Lin custom panel</td><td>Miltenyi Biotec</td><td>RRID: N/A</td></tr><tr><td>Brilliant Violet 570™ anti-human CD45</td><td>Sony</td><td>Cat# 2120170; RRID: N/A</td></tr><tr><td>Brilliant Violet 510™ anti-human CD20 Antibody</td><td>BioLegend</td><td>Cat# 302339 (also 302340); RRID:AB_2561721</td></tr><tr><td>PE/Cy7 anti-human CD34</td><td>Sony</td><td>Cat# 2317580; RRID: N/A</td></tr><tr><td>APC/Cy7 anti-human CD19</td><td>Sony</td><td>Cat# 2111090; RRID: N/A</td></tr><tr><td>BD™ CD10 APC</td><td>BD Biosciences</td><td>Cat# 332777; RRID:AB_2868626</td></tr><tr><td>BD Horizon™ BV421 Mouse Anti-Human IgM</td><td>BD Biosciences</td><td>Cat# 562618; RRID:AB_2737681</td></tr><tr><td>BD OptiBuild™ BV711 Mouse Anti-Human IgD</td><td>BD Biosciences</td><td>Cat# 740794; RRID:AB_2740457</td></tr><tr><td>BD Pharmingen™ FITC Mouse Anti-Human CD24</td><td>BD Biosciences</td><td>Cat# 555427; RRID:AB_395821</td></tr><tr><td>BD Pharmingen™ PE Mouse Anti-Human CD38</td><td>BD Biosciences</td><td>Cat# 555460; RRID:AB_395853</td></tr><tr><td>Monoclonal Mouse Anti-Human Glycophorin C (Concentrate)</td><td>Agilent</td><td>Cat# M082001; RRID: N/A</td></tr><tr><td>Polyclonal Rabbit Anti-Human Myeloperoxidase</td><td>Agilent</td><td>Cat# A039829; RRID: N/A</td></tr><tr><td>Polyclonal Rabbit Anti-Human Von Willebrand Factor</td><td>Agilent</td><td>Cat# A0082 (also A008202-5, A008202-2); RRID:AB_2315602</td></tr><tr><td>PE anti-human CD45 Antibody</td><td>BioLegend</td><td>Cat# 304008 (also 304007, 304039, 304058); RRID:AB_314396</td></tr><tr><td>APC anti-human HLA-A,B,C Antibody</td><td>BioLegend</td><td>Cat# 311409 (also 311410); RRID:AB_314878</td></tr><tr><td>CD3 Monoclonal Antibody (UCHT1), Alexa Fluor™ 532, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 58-0038-42; RRID:AB_11218675</td></tr><tr><td>TCR gamma/delta Monoclonal Antibody (B1.1), FITC, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 11-9959-41; RRID:AB_10669048</td></tr><tr><td>APC/Fire™ 750 anti-human TCR Vδ2 Antibody</td><td>BioLegend</td><td>Cat# 331419; RRID:AB_2687325</td></tr><tr><td>Brilliant Violet 605™ anti-human CD56 (NCAM) Antibody</td><td>BioLegend</td><td>Cat# 362537; RRID:AB_2565856</td></tr><tr><td>BD Horizon™ BV750 Mouse Anti-Human CD4</td><td>BD Biosciences</td><td>Cat# 566356; RRID:AB_2744426</td></tr><tr><td>Pacific Blue™ anti-human CD8 Antibody</td><td>BioLegend</td><td>Cat# 344717; RRID:AB_10551438</td></tr><tr><td>APC anti-human TCR Vα7.2 Antibody</td><td>BioLegend</td><td>Cat# 351708; RRID:AB_10933430</td></tr></tbody></table>

(Continued on next page)
Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>PE/Cyanine7 anti-human TCR Vα24-Jα18 (iNKT cell) Antibody</td><td>BioLegend</td><td>Cat# 342912; RRID:AB_2562229</td></tr><tr><td>Brilliant Violet 785™ anti-human CD20 Antibody</td><td>BioLegend</td><td>Cat# 302356; RRID:AB_2566316</td></tr><tr><td>CD279 (PD-1) Monoclonal Antibody (MIH4), PE, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 12-9969-42; RRID:AB_10736473</td></tr><tr><td>Brilliant Violet 711™ anti-human IFN-γ Antibody</td><td>BioLegend</td><td>Cat# 502540; RRID:AB_2563506</td></tr><tr><td>Brilliant Violet 510™ anti-human TNF-α Antibody</td><td>BioLegend</td><td>Cat# 502950; RRID:AB_2565860</td></tr><tr><td>PE/Dazzle™ 594 anti-human IL-10 Antibody</td><td>BioLegend</td><td>Cat# 506812; RRID:AB_2632782</td></tr><tr><td>BD Horizon™ BV605 Mouse Anti-Human CD45RA</td><td>BD Biosciences</td><td>Cat# 562886; RRID:AB_2737865</td></tr><tr><td>BD Pharmingen™ Alexa Fluor® 700 Mouse anti-Human CD197 (CCR7)</td><td>BD Biosciences</td><td>Cat# 561143; RRID:AB_10562031</td></tr><tr><td>BD Horizon™ BV421 Mouse Anti-Human CD3</td><td>BD Biosciences</td><td>Cat# 562426; RRID:AB_11152082</td></tr><tr><td>BD Horizon™ BUV395 Mouse Anti-Human CD8</td><td>BD Biosciences</td><td>Cat# 563795; RRID:AB_2722501</td></tr><tr><td>APC anti-human CD4 Antibody</td><td>BioLegend</td><td>Cat# 300514; RRID:AB_314082</td></tr><tr><td>PE/Cyanine7 anti-human TCR α/β Antibody</td><td>BioLegend</td><td>Cat# 306720; RRID:AB_10640829</td></tr><tr><td>TCR gamma/delta Monoclonal Antibody (B1.1), PE, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 12-9959-42; RRID:AB_1603300</td></tr><tr><td>BD Horizon™ BUV737 Mouse Anti-Human IFN-γ</td><td>BD Biosciences</td><td>Cat# 564620; RRID:AB_2869591</td></tr><tr><td>PerCP anti-human TNF-α Antibody</td><td>BioLegend</td><td>Cat# 502924; RRID:AB_2561287</td></tr><tr><td>BD Pharmingen™ PE Mouse Anti-Human IL-9</td><td>BD Biosciences</td><td>Cat# 560807; RRID:AB_2033985</td></tr><tr><td>BD Horizon™ BV421 Rat Anti-Human IL-13</td><td>BD Biosciences</td><td>Cat# 563580; RRID:AB_2738290</td></tr><tr><td>Alexa Fluor® 488 anti-human IL-4 Antibody</td><td>BioLegend</td><td>Cat# 500710; RRID:AB_1877131</td></tr><tr><td>Brilliant Violet 510™ anti-human IL-17A Antibody</td><td>BioLegend</td><td>Cat# 512330; RRID:AB_2562744</td></tr><tr><td>BD Horizon™ BV650 Mouse Anti-Human IL-17F</td><td>BD Biosciences</td><td>Cat# 564264; RRID:AB_2869555</td></tr><tr><td>BD Horizon™ BV750 Rat Anti-Human IL-2</td><td>BD Biosciences</td><td>Cat# 566361; RRID:AB_2739710</td></tr><tr><td>IL-21 Monoclonal Antibody (eBio3A3-N2 (3A3-N2)), eFluor™ 660, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 50-7219-42; RRID:AB_10598202</td></tr><tr><td>IL-22 Monoclonal Antibody (22URTI), PerCP-eFluor™ 710, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 46-7229-42; RRID:AB_10596639</td></tr><tr><td>CD19</td><td>Standard BioTools</td><td>Cat# 3142014D; RRID:AB_2924313</td></tr><tr><td>Vimentin</td><td>Standard BioTools</td><td>Cat# 3143027D; RRID: N/A</td></tr><tr><td>CD14</td><td>Standard BioTools</td><td>Cat# 3144025D; RRID:AB_2924314</td></tr><tr><td>NKp46/NCR1</td><td>R&amp;D Systems</td><td>Cat# MAB1850-100; RRID:AB_2149153</td></tr><tr><td>pan-cytokeratin</td><td>Standard BioTools</td><td>Cat# 3148022D; RRID:AB_2927682</td></tr><tr><td>CD15</td><td>Standard BioTools</td><td>Cat# 3149026D; RRID:AB_2928037</td></tr><tr><td>CD134 (OX40)</td><td>Standard BioTools</td><td>Cat# 3151024D; RRID:AB_2811014</td></tr><tr><td>CD45</td><td>Standard BioTools</td><td>Cat# 3152018D; RRID:AB_2909538</td></tr><tr><td>TCR-γδ</td><td>Santa-cruz Biotechnology</td><td>Cat# sc-100289; RRID:AB_1130061</td></tr><tr><td>CD11c</td><td>Abcam</td><td>Cat# ab216655; RRID:AB_2864379</td></tr><tr><td>FOXP3</td><td>Standard BioTools</td><td>Cat# 3155016D; RRID:AB_2910136</td></tr><tr><td>CD4</td><td>Standard BioTools</td><td>Cat# 3156033D; RRID:AB_2811051</td></tr><tr><td>E-cadherin</td><td>Standard BioTools</td><td>Cat# 3158029D; RRID:AB_2893074</td></tr><tr><td>CD68</td><td>Standard BioTools</td><td>Cat# 3159035D; RRID:AB_2810859</td></tr><tr><td>Langerin</td><td>Eurobio scientific</td><td>Cat# DDX0362; RRID:AB_1148742</td></tr><tr><td>CD141</td><td>Cell signaling</td><td>Cat# 34149SF; RRID: N/A</td></tr><tr><td>CD8a</td><td>Standard BioTools</td><td>Cat# 3162035D; RRID:AB_2909535</td></tr><tr><td>CD370/CLEC9A</td><td>Abcam</td><td>Cat# ab245121; RRID: N/A</td></tr><tr><td>CD103</td><td>Abcam</td><td>Cat# ab271889; RRID: N/A</td></tr><tr><td>CD45RA</td><td>Standard BioTools</td><td>Cat# 3166031D; RRID:AB_2924387</td></tr></tbody></table>

(Continued on next page)
Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>PE/Cyanine7 anti-human TCR Vα24-Jα18 (iNKT cell) Antibody</td><td>BioLegend</td><td>Cat# 342912; RRID:AB_2562229</td></tr><tr><td>Brilliant Violet 785™ anti-human CD20 Antibody</td><td>BioLegend</td><td>Cat# 302356; RRID:AB_2566316</td></tr><tr><td>CD279 (PD-1) Monoclonal Antibody (MIH4), PE, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 12-9969-42; RRID:AB_10736473</td></tr><tr><td>Brilliant Violet 711™ anti-human IFN-γ Antibody</td><td>BioLegend</td><td>Cat# 502540; RRID:AB_2563506</td></tr><tr><td>Brilliant Violet 510™ anti-human TNF-α Antibody</td><td>BioLegend</td><td>Cat# 502950; RRID:AB_2565860</td></tr><tr><td>PE/Dazzle™ 594 anti-human IL-10 Antibody</td><td>BioLegend</td><td>Cat# 506812; RRID:AB_2632782</td></tr><tr><td>BD Horizon™ BV605 Mouse Anti-Human CD45RA</td><td>BD Biosciences</td><td>Cat# 562886; RRID:AB_2737865</td></tr><tr><td>BD Pharmingen™ Alexa Fluor® 700 Mouse anti-Human CD197 (CCR7)</td><td>BD Biosciences</td><td>Cat# 561143; RRID:AB_10562031</td></tr><tr><td>BD Horizon™ BV421 Mouse Anti-Human CD3</td><td>BD Biosciences</td><td>Cat# 562426; RRID:AB_11152082</td></tr><tr><td>BD Horizon™ BUV395 Mouse Anti-Human CD8</td><td>BD Biosciences</td><td>Cat# 563795; RRID:AB_2722501</td></tr><tr><td>APC anti-human CD4 Antibody</td><td>BioLegend</td><td>Cat# 300514; RRID:AB_314082</td></tr><tr><td>PE/Cyanine7 anti-human TCR α/β Antibody</td><td>BioLegend</td><td>Cat# 306720; RRID:AB_10640829</td></tr><tr><td>TCR gamma/delta Monoclonal Antibody (B1.1), PE, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 12-9959-42; RRID:AB_1603300</td></tr><tr><td>BD Horizon™ BUV737 Mouse Anti-Human IFN-γ</td><td>BD Biosciences</td><td>Cat# 564620; RRID:AB_2869591</td></tr><tr><td>PerCP anti-human TNF-α Antibody</td><td>BioLegend</td><td>Cat# 502924; RRID:AB_2561287</td></tr><tr><td>BD Pharmingen™ PE Mouse Anti-Human IL-9</td><td>BD Biosciences</td><td>Cat# 560807; RRID:AB_2033985</td></tr><tr><td>BD Horizon™ BV421 Rat Anti-Human IL-13</td><td>BD Biosciences</td><td>Cat# 563580; RRID:AB_2738290</td></tr><tr><td>Alexa Fluor® 488 anti-human IL-4 Antibody</td><td>BioLegend</td><td>Cat# 500710; RRID:AB_1877131</td></tr><tr><td>Brilliant Violet 510™ anti-human IL-17A Antibody</td><td>BioLegend</td><td>Cat# 512330; RRID:AB_2562744</td></tr><tr><td>BD Horizon™ BV650 Mouse Anti-Human IL-17F</td><td>BD Biosciences</td><td>Cat# 564264; RRID:AB_2869555</td></tr><tr><td>BD Horizon™ BV750 Rat Anti-Human IL-2</td><td>BD Biosciences</td><td>Cat# 566361; RRID:AB_2739710</td></tr><tr><td>IL-21 Monoclonal Antibody (eBio3A3-N2 (3A3-N2)), eFluor™ 660, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 50-7219-42; RRID:AB_10598202</td></tr><tr><td>IL-22 Monoclonal Antibody (22URTI), PerCP-eFluor™ 710, eBioscience™</td><td>Thermo Fisher Scientific</td><td>Cat# 46-7229-42; RRID:AB_10596639</td></tr><tr><td>CD19</td><td>Standard BioTools</td><td>Cat# 3142014D; RRID:AB_2924313</td></tr><tr><td>Vimentin</td><td>Standard BioTools</td><td>Cat# 3143027D; RRID: N/A</td></tr><tr><td>CD14</td><td>Standard BioTools</td><td>Cat# 3144025D; RRID:AB_2924314</td></tr><tr><td>NKp46/NCR1</td><td>R&amp;D Systems</td><td>Cat# MAB1850-100; RRID:AB_2149153</td></tr><tr><td>pan-cytokeratin</td><td>Standard BioTools</td><td>Cat# 3148022D; RRID:AB_2927682</td></tr><tr><td>CD15</td><td>Standard BioTools</td><td>Cat# 3149026D; RRID:AB_2928037</td></tr><tr><td>CD134 (OX40)</td><td>Standard BioTools</td><td>Cat# 3151024D; RRID:AB_2811014</td></tr><tr><td>CD45</td><td>Standard BioTools</td><td>Cat# 3152018D; RRID:AB_2909538</td></tr><tr><td>TCR-γδ</td><td>Santa-cruz Biotechnology</td><td>Cat# sc-100289; RRID:AB_1130061</td></tr><tr><td>CD11c</td><td>Abcam</td><td>Cat# ab216655; RRID:AB_2864379</td></tr><tr><td>FOXP3</td><td>Standard BioTools</td><td>Cat# 3155016D; RRID:AB_2910136</td></tr><tr><td>CD4</td><td>Standard BioTools</td><td>Cat# 3156033D; RRID:AB_2811051</td></tr><tr><td>E-cadherin</td><td>Standard BioTools</td><td>Cat# 3158029D; RRID:AB_2893074</td></tr><tr><td>CD68</td><td>Standard BioTools</td><td>Cat# 3159035D; RRID:AB_2810859</td></tr><tr><td>Langerin</td><td>Eurobio scientific</td><td>Cat# DDX0362; RRID:AB_1148742</td></tr><tr><td>CD141</td><td>Cell signaling</td><td>Cat# 34149SF; RRID: N/A</td></tr><tr><td>CD8a</td><td>Standard BioTools</td><td>Cat# 3162035D; RRID:AB_2909535</td></tr><tr><td>CD370/CLEC9A</td><td>Abcam</td><td>Cat# ab245121; RRID: N/A</td></tr><tr><td>CD103</td><td>Abcam</td><td>Cat# ab271889; RRID: N/A</td></tr><tr><td>CD45RA</td><td>Standard BioTools</td><td>Cat# 3166031D; RRID:AB_2924387</td></tr></tbody></table>

(Continued on next page)
Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>GZM-B</td><td>Standard BioTools</td><td>Cat# 3167021D, RRID:AB_2811057</td></tr><tr><td>Ki67</td><td>Standard BioTools</td><td>Cat# 3168022D, RRID:AB_2811061</td></tr><tr><td>L1</td><td>Merck</td><td>Cat# MAB837, RRID:AB_95315</td></tr><tr><td>CD3</td><td>Standard BioTools</td><td>Cat# 3170019D, RRID:AB_2811048</td></tr><tr><td>CD49a</td><td>R&amp;D systems</td><td>Cat# AF5676, RRID:AB_2129238</td></tr><tr><td>HLA-DR</td><td>Standard BioTools</td><td>Cat# 3174025D, RRID:AB_2924389</td></tr><tr><td>CD25</td><td>Standard BioTools</td><td>Cat# 3175036D, RRID:AB_2909537</td></tr><tr><td>CD303</td><td>Eurobio scientific</td><td>Cat# DDX0043, RRID:AB_1149764</td></tr><tr><td>Anti-Human IgA (alpha) FITC</td><td>Inova Diagnostics</td><td>Cat# 504045, RRID: N/A</td></tr><tr><td>Antibody against mouse papillomavirus E4</td><td>in house</td><td>Rabbit serum, RRID: N/A</td></tr><tr><td>Antibody against mouse papillomavirus L1</td><td>in house</td><td>MPV.B9, RRID: N/A</td></tr><tr><td>Goat Anti-Mouse IgG, Human ads-AP</td><td>Southern Biotech</td><td>Cat# 1030-04, RRID:AB_2794293</td></tr><tr><td>Goat Anti-Mouse IgG3, Human ads-AP</td><td>Southern Biotech</td><td>Cat# 1100-04, RRID: N/A</td></tr><tr><td>PNPP (p-Nitrophenyl Phosphate, Disodium Sal)</td><td>Sigma</td><td>Cat# 487666, RRID: N/A</td></tr></tbody></table>

Bacterial and virus strains

<table><tr><td>Mycobacterium bovis BCG</td><td>Vogt and Nathan<sup>93</sup>, PMID: 21911939</td><td>N/A</td></tr><tr><td>NEB® 10-beta Competent E. coli (High Efficiency)</td><td>New England Biolabs</td><td>Cat# C3019H</td></tr><tr><td>NEB® Stable Competent E. coli (High Efficiency)</td><td>New England Biolabs</td><td>Cat# C3040H</td></tr><tr><td>Mouse papillomavirus</td><td>HSD:NU mouse tail lesions, Hu et al.<sup>94</sup></td><td>MmuPV1</td></tr></table>

Biological samples

<table><tr><td>Plasma from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>Peripheral blood mononuclear cells from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>Skin Biopsies from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>Bone marrow mononuclear cells from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>Bone biopsies from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>Fresh and FFPE tail and muzzle tissues from Rag1ko, B6, and Flt3lko</td><td>Infected tissues from the current study</td><td>N/A</td></tr></table>

Chemicals, peptides, and recombinant proteins

<table><tr><td>ImmunoCult™ Human CD3/CD28/CD2 T Cell Activator</td><td>STEMCELL Technologies</td><td>Cat# 10970</td></tr><tr><td>Gibco™ Human IL-2 Recombinant Protein</td><td>Thermo Fisher Scientific</td><td>Cat# PHC0023</td></tr><tr><td>Recombinant Human Flt-3 Ligand/FLT3L (HEK293) Protein, CF</td><td>R&amp;D Systems</td><td>Cat# 308-FKHB-010</td></tr><tr><td>X-tremeGENE™ 9 DNA Transfection Reagent</td><td>Roche</td><td>Cat# 6365809001</td></tr><tr><td>LIVE/DEAD™ Fixable Aqua Dead Cell Stain Kit</td><td>Thermo Fisher Scientific</td><td>Cat# L34957</td></tr><tr><td>Dispase</td><td>STEMCELL Technologies</td><td>Cat# 07923</td></tr><tr><td>Collagenase D</td><td>Roche</td><td>Cat# 1088866001</td></tr><tr><td>Gibco™ Trypsin-EDTA (0.05%)</td><td>Thermo Fisher Scientific</td><td>Cat# 11580626</td></tr><tr><td>GolgiPlug</td><td>BD Biosciences</td><td>Cat# 555029</td></tr><tr><td>Zombie NIR™ Fixable Viability Kit</td><td>BioLegend</td><td>Cat# 423105</td></tr><tr><td>FcBlock</td><td>Miltenyi Biotec</td><td>Cat# 130-059-901</td></tr><tr><td>Recombinant human Interleukin-12</td><td>R&amp;D Systems</td><td>Cat# 219-IL-025</td></tr><tr><td>Recombinant human Interleukin-23</td><td>R&amp;D Systems</td><td>Cat# 1290-IL-010</td></tr><tr><td>Phorbol 12-myristate 13-acetate</td><td>Merck</td><td>Cat# P1585</td></tr><tr><td>Phorbol 12-myristate 13-acetate</td><td>Merck</td><td>Cat# P8139</td></tr></table>

(Continued on next page)
Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>GZM-B</td><td>Standard BioTools</td><td>Cat# 3167021D, RRID:AB_2811057</td></tr><tr><td>Ki67</td><td>Standard BioTools</td><td>Cat# 3168022D, RRID:AB_2811061</td></tr><tr><td>L1</td><td>Merck</td><td>Cat# MAB837, RRID:AB_95315</td></tr><tr><td>CD3</td><td>Standard BioTools</td><td>Cat# 3170019D, RRID:AB_2811048</td></tr><tr><td>CD49a</td><td>R&amp;D systems</td><td>Cat# AF5676, RRID:AB_2129238</td></tr><tr><td>HLA-DR</td><td>Standard BioTools</td><td>Cat# 3174025D, RRID:AB_2924389</td></tr><tr><td>CD25</td><td>Standard BioTools</td><td>Cat# 3175036D, RRID:AB_2909537</td></tr><tr><td>CD303</td><td>Eurobio scientific</td><td>Cat# DDX0043, RRID:AB_1149764</td></tr><tr><td>Anti-Human IgA (alpha) FITC</td><td>Inova Diagnostics</td><td>Cat# 504045, RRID: N/A</td></tr><tr><td>Antibody against mouse papillomavirus E4</td><td>in house</td><td>Rabbit serum, RRID: N/A</td></tr><tr><td>Antibody against mouse papillomavirus L1</td><td>in house</td><td>MPV.B9, RRID: N/A</td></tr><tr><td>Goat Anti-Mouse IgG, Human ads-AP</td><td>Southern Biotech</td><td>Cat# 1030-04, RRID:AB_2794293</td></tr><tr><td>Goat Anti-Mouse IgG3, Human ads-AP</td><td>Southern Biotech</td><td>Cat# 1100-04, RRID: N/A</td></tr><tr><td>PNPP (p-Nitrophenyl Phosphate, Disodium Sal)</td><td>Sigma</td><td>Cat# 487666, RRID: N/A</td></tr></tbody></table>

Bacterial and virus strains

<table><tr><td>Mycobacterium bovis BCG</td><td>Vogt and Nathan<sup>93</sup>, PMID: 21911939</td><td>N/A</td></tr><tr><td>NEB® 10-beta Competent E. coli (High Efficiency)</td><td>New England Biolabs</td><td>Cat# C3019H</td></tr><tr><td>NEB® Stable Competent E. coli (High Efficiency)</td><td>New England Biolabs</td><td>Cat# C3040H</td></tr><tr><td>Mouse papillomavirus</td><td>HSD:NU mouse tail lesions, Hu et al.<sup>94</sup></td><td>MmuPV1</td></tr></table>

Biological samples

<table><tr><td>Plasma from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>Peripheral blood mononuclear cells from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>Skin Biopsies from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>Bone marrow mononuclear cells from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>Bone biopsies from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>Fresh and FFPE tail and muzzle tissues from Rag1ko, B6, and Flt3lko</td><td>Infected tissues from the current study</td><td>N/A</td></tr></table>

Chemicals, peptides, and recombinant proteins

<table><tr><td>ImmunoCult™ Human CD3/CD28/CD2 T Cell Activator</td><td>STEMCELL Technologies</td><td>Cat# 10970</td></tr><tr><td>Gibco™ Human IL-2 Recombinant Protein</td><td>Thermo Fisher Scientific</td><td>Cat# PHC0023</td></tr><tr><td>Recombinant Human Flt-3 Ligand/FLT3L (HEK293) Protein, CF</td><td>R&amp;D Systems</td><td>Cat# 308-FKHB-010</td></tr><tr><td>X-tremeGENE™ 9 DNA Transfection Reagent</td><td>Roche</td><td>Cat# 6365809001</td></tr><tr><td>LIVE/DEAD™ Fixable Aqua Dead Cell Stain Kit</td><td>Thermo Fisher Scientific</td><td>Cat# L34957</td></tr><tr><td>Dispase</td><td>STEMCELL Technologies</td><td>Cat# 07923</td></tr><tr><td>Collagenase D</td><td>Roche</td><td>Cat# 1088866001</td></tr><tr><td>Gibco™ Trypsin-EDTA (0.05%)</td><td>Thermo Fisher Scientific</td><td>Cat# 11580626</td></tr><tr><td>GolgiPlug</td><td>BD Biosciences</td><td>Cat# 555029</td></tr><tr><td>Zombie NIR™ Fixable Viability Kit</td><td>BioLegend</td><td>Cat# 423105</td></tr><tr><td>FcBlock</td><td>Miltenyi Biotec</td><td>Cat# 130-059-901</td></tr><tr><td>Recombinant human Interleukin-12</td><td>R&amp;D Systems</td><td>Cat# 219-IL-025</td></tr><tr><td>Recombinant human Interleukin-23</td><td>R&amp;D Systems</td><td>Cat# 1290-IL-010</td></tr><tr><td>Phorbol 12-myristate 13-acetate</td><td>Merck</td><td>Cat# P1585</td></tr><tr><td>Phorbol 12-myristate 13-acetate</td><td>Merck</td><td>Cat# P8139</td></tr></table>

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Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>Interleukin-2 Protein, Recombinant human</td><td>Merck</td><td>Cat# IL002</td></tr><tr><td>Recombinant Human TGF-β1</td><td>Peprotech</td><td>Cat# 100-21C</td></tr><tr><td>Recombinant Human IL-1β</td><td>Peprotech</td><td>Cat# 200-01B</td></tr><tr><td>Recombinant Human IL-6</td><td>Peprotech</td><td>Cat# 200-06</td></tr><tr><td>Recombinant Human IL-21</td><td>Peprotech</td><td>Cat# 200-21</td></tr><tr><td>Recombinant Human IL-23</td><td>Peprotech</td><td>Cat# 200-23</td></tr><tr><td>Ionomycin calcium salt from Streptomyces conglobatus</td><td>Merck</td><td>Cat# I0634</td></tr><tr><td>Zombie UV™ Fixable Viability Kit</td><td>Biolegend</td><td>Cat# 423107</td></tr><tr><td>Brefeldin A</td><td>Merck</td><td>Cat# B7651</td></tr><tr><td>KLH-conjugated MmuPV1 E4 peptide (PKTTPPRRELFPPPTPLTQPP)</td><td>China Peptide</td><td>N/A</td></tr><tr><td>DpnI</td><td>New England Biolabs</td><td>Cat# R0176L</td></tr><tr><td>Protamine sulfate</td><td>Merck</td><td>Cat# P3369-10G</td></tr><tr><td>Lymphoprep</td><td>STEMCELL Technologies</td><td>Cat# 07801</td></tr><tr><td>BD Pharmingen™ 7-AAD</td><td>BD Biosciences</td><td>Cat# 559925</td></tr><tr><td>Cell-ID™ Intercalator-Ir</td><td>Standard BioTools</td><td>Cat# 201192A</td></tr><tr><td>PKTTPPRRELFPPPTPLTQPP</td><td>Brendle et al.<sup>95</sup></td><td>Mouse papillomavirus E4 peptide</td></tr><tr><td>Mouse papillomavirus L1 virus like particles</td><td>Brendle et al.<sup>96,97</sup></td><td>MmuPV1L1VLPs</td></tr></tbody></table>

Critical commercial assays

<table><tbody><tr><td>Zero Blunt™ TOPO™ PCR Cloning Kit</td><td>Thermo Fisher Scientific</td><td>Cat# 450245</td></tr><tr><td>pcDNA™3.1 Directional TOPO™ Expression Kit</td><td>Thermo Fisher Scientific</td><td>Cat# K490001</td></tr><tr><td>High-Capacity RNA-to-cDNA™ Kit</td><td>Thermo Fisher Scientific</td><td>Cat# 4387406</td></tr><tr><td>SuperScript™ II Reverse Transcriptase</td><td>Thermo Fisher Scientific</td><td>Cat# 18064014</td></tr><tr><td>LEGENDplex™ Human Inflammation Panel 1</td><td>Biolegend</td><td>Cat# 740809</td></tr><tr><td>Human Flt-3 Ligand/FLT3L DuoSet ELISA</td><td>R&amp;D Systems</td><td>Cat# DY308</td></tr><tr><td>CD34 MicroBead Kit UltraPure, human</td><td>Miltenyi Biotec</td><td>Cat# 130-100-453</td></tr><tr><td>CD34 MicroBead Kit, human</td><td>Miltenyi Biotec</td><td>Cat# 130-046-702</td></tr><tr><td>CD19 MicroBeads, human</td><td>Miltenyi Biotec</td><td>Cat# 130-050-301</td></tr><tr><td>CD271 MicroBeads</td><td>Miltenyi Biotec</td><td>Cat# 130-099-023</td></tr><tr><td>T Cell Activation/Expansion Kit, human</td><td>Miltenyi Biotec</td><td>Cat# 130-091-441</td></tr><tr><td>Chromium Next GEM Single Cell 3' Reagent Kit v3.1</td><td>10X Genomics</td><td>Cat# CG000315</td></tr><tr><td>Chromium Next GEM Single Cell 5' Reagent Kit v2</td><td>10X Genomics</td><td>Cat# CG000331</td></tr><tr><td>SureSelectXT Human All Exon V6</td><td>Agilent</td><td>Cat# 5190-8864</td></tr><tr><td>Big Dye Terminator v3.1 cycle sequencing kit</td><td>Applied Biosystems</td><td>Cat# 4337455</td></tr><tr><td>M.O.M.® (Mouse on Mouse) ImmPRESS® HRP (Peroxidase) Polymer Kit</td><td>Vector Laboratories</td><td>Cat# MP-2400</td></tr><tr><td>ImmPRESS anti-rabbit IgG polymer system</td><td>Vector Laboratories</td><td>Cat# MP-7801</td></tr><tr><td>ImmPACT® NovaRED® Substrate</td><td>Vector Laboratories</td><td>Cat# SK-4805</td></tr><tr><td>RNAScope® 2.5 VS Probe- V-MusPV-E4</td><td>Advanced Cell Diagnostics, Inc.</td><td>Cat# 473289</td></tr><tr><td>RNAScope 2.5 HD Assay - BROWN</td><td>Advanced Cell Diagnostics, Inc.</td><td>Cat# 322360</td></tr><tr><td>Amersham Megaprime DNA Labeling System</td><td>Amersham</td><td>Cat# RPN1604</td></tr><tr><td>The Brilliant III qPCR kit</td><td>Agilent</td><td>Cat# 600880</td></tr><tr><td>The RevertAid First Strand cDNA synthesis kit</td><td>Thermo Fisher Scientific</td><td>Cat# K1622</td></tr></tbody></table>

Deposited data

<table><tbody><tr><td>scRNAseq on fresh PBMCs</td><td>This manuscript</td><td>NCBI BioProject: PRJNA999252</td></tr><tr><td>scRNAseq on fresh sorted HLA-DR<sup>+</sup>Lin<sup>+</sup> (CD3, CD19, CD20, and CD56) cells</td><td>This manuscript</td><td>NCBI BioProject: PRJNA999252</td></tr><tr><td>scRNAseq on digested skin biopsies</td><td>This manuscript</td><td>NCBI BioProject: PRJNA999252</td></tr></tbody></table>

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Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>Interleukin-2 Protein, Recombinant human</td><td>Merck</td><td>Cat# IL002</td></tr><tr><td>Recombinant Human TGF-β1</td><td>Peprotech</td><td>Cat# 100-21C</td></tr><tr><td>Recombinant Human IL-1β</td><td>Peprotech</td><td>Cat# 200-01B</td></tr><tr><td>Recombinant Human IL-6</td><td>Peprotech</td><td>Cat# 200-06</td></tr><tr><td>Recombinant Human IL-21</td><td>Peprotech</td><td>Cat# 200-21</td></tr><tr><td>Recombinant Human IL-23</td><td>Peprotech</td><td>Cat# 200-23</td></tr><tr><td>Ionomycin calcium salt from Streptomyces conglobatus</td><td>Merck</td><td>Cat# I0634</td></tr><tr><td>Zombie UV™ Fixable Viability Kit</td><td>Biolegend</td><td>Cat# 423107</td></tr><tr><td>Brefeldin A</td><td>Merck</td><td>Cat# B7651</td></tr><tr><td>KLH-conjugated MmuPV1 E4 peptide (PKTTPPRRELFPPTPLTQPP)</td><td>China Peptide</td><td>N/A</td></tr><tr><td>DpnI</td><td>New England Biolabs</td><td>Cat# R0176L</td></tr><tr><td>Protamine sulfate</td><td>Merck</td><td>Cat# P3369-10G</td></tr><tr><td>Lymphoprep</td><td>STEMCELL Technologies</td><td>Cat# 07801</td></tr><tr><td>BD Pharmingen™ 7-AAD</td><td>BD Biosciences</td><td>Cat# 559925</td></tr><tr><td>Cell-ID™ Intercalator-Ir</td><td>Standard BioTools</td><td>Cat# 201192A</td></tr><tr><td>PKTTPPRRELFPPTPLTQPP</td><td>Brendle et al.<sup>95</sup></td><td>Mouse papillomavirus E4 peptide</td></tr><tr><td>Mouse papillomavirus L1 virus like particles</td><td>Brendle et al.<sup>96,97</sup></td><td>MmuPV1L1VLPs</td></tr></tbody></table>

Critical commercial assays

<table><tbody><tr><td>Zero Blunt™ TOPO™ PCR Cloning Kit</td><td>Thermo Fisher Scientific</td><td>Cat# 450245</td></tr><tr><td>pcDNA™3.1 Directional TOPO™ Expression Kit</td><td>Thermo Fisher Scientific</td><td>Cat# K490001</td></tr><tr><td>High-Capacity RNA-to-cDNA™ Kit</td><td>Thermo Fisher Scientific</td><td>Cat# 4387406</td></tr><tr><td>SuperScript™ II Reverse Transcriptase</td><td>Thermo Fisher Scientific</td><td>Cat# 18064014</td></tr><tr><td>LEGENDplex™ Human Inflammation Panel 1</td><td>Biolegend</td><td>Cat# 740809</td></tr><tr><td>Human Flt-3 Ligand/FLT3L DuoSet ELISA</td><td>R&amp;D Systems</td><td>Cat# DY308</td></tr><tr><td>CD34 MicroBead Kit UltraPure, human</td><td>Miltenyi Biotec</td><td>Cat# 130-100-453</td></tr><tr><td>CD34 MicroBead Kit, human</td><td>Miltenyi Biotec</td><td>Cat# 130-046-702</td></tr><tr><td>CD19 MicroBeads, human</td><td>Miltenyi Biotec</td><td>Cat# 130-050-301</td></tr><tr><td>CD271 MicroBeads</td><td>Miltenyi Biotec</td><td>Cat# 130-099-023</td></tr><tr><td>T Cell Activation/Expansion Kit, human</td><td>Miltenyi Biotec</td><td>Cat# 130-091-441</td></tr><tr><td>Chromium Next GEM Single Cell 3' Reagent Kit v3.1</td><td>10X Genomics</td><td>Cat# CG000315</td></tr><tr><td>Chromium Next GEM Single Cell 5' Reagent Kit v2</td><td>10X Genomics</td><td>Cat# CG000331</td></tr><tr><td>SureSelectXT Human All Exon V6</td><td>Agilent</td><td>Cat# 5190-8864</td></tr><tr><td>Big Dye Terminator v3.1 cycle sequencing kit</td><td>Applied Biosystems</td><td>Cat# 4337455</td></tr><tr><td>M.O.M.® (Mouse on Mouse) ImmPRESS® HRP (Peroxidase) Polymer Kit</td><td>Vector Laboratories</td><td>Cat# MP-2400</td></tr><tr><td>ImmPRESS anti-rabbit IgG polymer system</td><td>Vector Laboratories</td><td>Cat# MP-7801</td></tr><tr><td>ImmPACT® NovaRED® Substrate</td><td>Vector Laboratories</td><td>Cat# SK-4805</td></tr><tr><td>RNAScope® 2.5 VS Probe- V-MusPV-E4</td><td>Advanced Cell Diagnostics, Inc.</td><td>Cat# 473289</td></tr><tr><td>RNAScope 2.5 HD Assay - BROWN</td><td>Advanced Cell Diagnostics, Inc.</td><td>Cat# 322360</td></tr><tr><td>Amersham Megaprime DNA Labeling System</td><td>Amersham</td><td>Cat# RPN1604</td></tr><tr><td>The Brilliant III qPCR kit</td><td>Agilent</td><td>Cat# 600880</td></tr><tr><td>The RevertAid First Strand cDNA synthesis kit</td><td>Thermo Fisher Scientific</td><td>Cat# K1622</td></tr></tbody></table>

Deposited data

<table><tbody><tr><td>scRNAseq on fresh PBMCs</td><td>This manuscript</td><td>NCBI BioProject: PRJNA999252</td></tr><tr><td>scRNAseq on fresh sorted HLA-DR<sup>+</sup>Lin<sup>+</sup> (CD3, CD19, CD20, and CD56) cells</td><td>This manuscript</td><td>NCBI BioProject: PRJNA999252</td></tr><tr><td>scRNAseq on digested skin biopsies</td><td>This manuscript</td><td>NCBI BioProject: PRJNA999252</td></tr></tbody></table>

(Continued on next page)
Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>scRNAseq on fresh CD34<sup>+</sup> enriched BMNCs</td><td>This manuscript</td><td>EMBL-EBI Biostudies: S-BSST1333</td></tr><tr><td>High-throughput sequencing (HTS) of T cell receptor β (TRB) and B cell receptor (IGH) dataset (control and patients)</td><td>Adaptive Biotechnologies, This manuscript</td><td>https://clients.adaptivebiotech.com/pub/momenilandi-2024-cell (DOI: 10.21417/MM2024C)</td></tr></tbody></table>

Experimental models: Cell lines

<table><tr><td>HEK293T cells</td><td>ATCC</td><td>CRL-11268</td></tr><tr><td>K562-FLT3</td><td>This manuscript</td><td>N/A</td></tr><tr><td>T cell blasts from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>EBV-B cells from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr></table>

Experimental models: Organisms/strains

<table><tr><td>C57BL/6J</td><td>The Jackson Laboratory</td><td>https://www.jax.org/strain/000664</td></tr><tr><td>C57BL/6-Flt3ltm1Imx/TacMmjax</td><td>The Jackson Laboratory</td><td>https://www.jax.org/strain/025411 -</td></tr><tr><td>B6.129S7-Rag1tm1Mom/J</td><td>The Jackson Laboratory, Beziat et al.<sup>98</sup></td><td>https://www.jax.org/strain/002216</td></tr></table>

Oligonucleotides

<table><tr><td>cDNA-FLT3L-Ex1 For 5' tttcggctctggctgtcac 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-FLT3L-Ex9 Rev15' ctgtgtccaggctatgcatc 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-For5' caccatgacagtgctggccgcagc 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev15' tcagtgctccacaagcagcaggtcctg 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev25' gtgctccacaagcagcaggtcctg 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.83)15' tcagtatcctccccaggatgaggcctt 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.83)25' gtatcctccccaggatgaggcctt 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(full9c.83)15' tcagtatcctccccctgtaaaatgggatgatag 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(full9c.83)25' gtatcctccccctgtaaaatgggatgataga 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.121)15' ctatgcatcctctggctggtgactcccctc 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.121)25' tgcatcctctggctggtgactcccctc 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.260)15' tcatggggacaagggctttgtacagag 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.260)25' tggggacaagggctttgtacagag 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3L-del7-For5' gtgccccccgt ccccagtcctccaggacctg 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>Homrec-FLT3L-For5' CCAGTACC CTTCACCATgacagtgctggccgcagcct 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>Homrec-FLT3L-Rev5' ctggggacgg ggggcaccgggctgacactgcagctcca 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>pcDNA-open-Rev5' GGTGAAGGG TACTGGATCCGAGCT 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>pTrip-dNGFR-For5' AGGCACCAC CGACAACCTCATCCCT 3'</td><td>Eurofins</td><td>N/A</td></tr></table>

(Continued on next page)
Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>scRNAseq on fresh CD34<sup>+</sup> enriched BMNCs</td><td>This manuscript</td><td>EMBL-EBI Biostudies: S-BSST1333</td></tr><tr><td>High-throughput sequencing (HTS) of T cell receptor β (TRB) and B cell receptor (IGH) dataset (control and patients)</td><td>Adaptive Biotechnologies, This manuscript</td><td>https://clients.adaptivebiotech.com/pub/momenilandi-2024-cell (DOI: 10.21417/MM2024C)</td></tr></tbody></table>

Experimental models: Cell lines

<table><tr><td>HEK293T cells</td><td>ATCC</td><td>CRL-11268</td></tr><tr><td>K562-FLT3</td><td>This manuscript</td><td>N/A</td></tr><tr><td>T cell blasts from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr><tr><td>EBV-B cells from indicated individuals</td><td>This manuscript</td><td>N/A</td></tr></table>

Experimental models: Organisms/strains

<table><tr><td>C57BL/6J</td><td>The Jackson Laboratory</td><td>https://www.jax.org/strain/000664</td></tr><tr><td>C57BL/6-Flt3ltm1Imx/TacMmjax</td><td>The Jackson Laboratory</td><td>https://www.jax.org/strain/025411 -</td></tr><tr><td>B6.129S7-Rag1tm1Mom/J</td><td>The Jackson Laboratory, Beziat et al.<sup>98</sup></td><td>https://www.jax.org/strain/002216</td></tr></table>

Oligonucleotides

<table><tr><td>cDNA-FLT3L-Ex1 For 5' tttcggtcttggctgtcac 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-FLT3L-Ex9 Rev15' ctgtgtccaggctatgcatc 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-For5' caccatgacagtgctggccgcagc 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev15' tcagtgctccacaagcagcaggtcctg 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev25' gtgctccacaagcagcaggtcctg 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.83)15' tcagtatcctccccaggatgaggcctt 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.83)25' gtatcctccccaggatgaggcctt 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(full9c.83)15' tcagtatcctccccctgtaaaatgggatgatag 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(full9c.83)25' gtatcctccccctgtaaaatgggatgataga 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.121)15' ctatgcatcctctggctggtgactcccctc 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.121)25' tgcatcctctggctggtgactcccctc 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.260)15' tcatggggacaagggctttgtacagag 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>cDNA-topodirect-Rev(c.260)25' tggggacaagggctttgtacagag 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3L-del7-For5' gtgccccccgt ccccagtcctccaggacctg 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>Homrec-FLT3L-For5' CCAGTACC CTTCACCATgacagtgctggccgcagcct 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>Homrec-FLT3L-Rev5' ctggggacgg ggggcaccgggctgacactgcagctcca 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>pcDNA-open-Rev5' GGTGAAGGG TACTGGATCCGAGCT 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>pTrip-dNGFR-For5' AGGCACCAC CGACAACCTCATCCCT 3'</td><td>Eurofins</td><td>N/A</td></tr></table>

(Continued on next page)
Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>pTrip-Rev5' CTCCTCTTGTGCTTCTAGCCAG 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>pTrip-FLT3L-For5' GAGAACCTGGACCT atgacagtgtggtcgcca 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>pTrip-FLT3L-Rev15' TTTTCTAGGTCTCGA tcagtgcctccacaagcagca 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3LG-S22P-For5' gctgctgagcccgggactcag 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3LG-S22P-Rev5' agcagcaggaggagatagg 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3LG-S118Afs-For5' ccccccccagctgtcttcgctt 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3LG-S118Afs-Rev5' ctgaaaggcacatttggt gacaaa 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3LG-gDNA-For5' ATTTCACCACTCCAG CCTGG 3'</td><td>Thermo Fisher Scientific</td><td>N/A</td></tr><tr><td>FLT3LG-gDNA-Rev5' CCTTTAC CGGGCTGACACTG 3'</td><td>Thermo Fisher Scientific</td><td>N/A</td></tr><tr><td>Flt3lko mutant reverse5' ATT TGT CAC GTC CTG CAC GAC G 3'</td><td>Integrated DNA Technologies</td><td>N/A</td></tr><tr><td>Common forward5' TGG CAG CTG AAG TGA CTG AC 3'</td><td>Integrated DNA Technologies</td><td>N/A</td></tr><tr><td>Flt3l wild type reverse5' AAG CCA AAG CTG GAT GAC AG 3'</td><td>Integrated DNA Technologies</td><td>N/A</td></tr><tr><td>Forward primers for the mouse papillomavirus5'-TAGCTTTGTCTGCCCCGCACT-3'</td><td>Integrated DNA Technologies, Brendle et al.<sup>96</sup></td><td>N/A</td></tr><tr><td>Reverse primers for the mouse papillomavirus5'-GTCAGTGGTGTTCGGTGGGAA-3'</td><td>Integrated DNA Technologies, Brendle et al.<sup>96</sup></td><td>N/A</td></tr><tr><td>Probe for the mouse papillomavirus5'FAM-CG GCCCGAAGACAACACCGCCACG-3'TAMRA</td><td>Integrated DNA Technologies, Brendle et al.<sup>96</sup></td><td>N/A</td></tr><tr><td>FLT3LG</td><td>Thermo Fisher Scientific</td><td>Hs00181740_m1</td></tr><tr><td>FLT3LG</td><td>Thermo Fisher Scientific</td><td>Hs00953092_g1</td></tr><tr><td>GUSB</td><td>Thermo Fisher Scientific</td><td>4326320E</td></tr></tbody></table>

Recombinant DNA

<table><tr><td>pcDNA3.1/V5-His</td><td>Thermo Fisher Scientific</td><td>V79020</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T1/T2)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T3)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T4)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T5)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T6)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T7)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-mFLT3LG (T1/2) -Ser118Alafs23*</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-mFLT3LG (T1/2)-Ser22Pro</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-sFLT3LG (T7)-Ser118Alafs23*</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T7)-Ser22Pro</td><td>This manuscript</td><td>N/A</td></tr><tr><td>psPAX2</td><td>Addgene</td><td>12260</td></tr><tr><td>pCMV-VSV-G</td><td>Addgene</td><td>8454</td></tr><tr><td>pHXB2-Env</td><td>NIH-AIDS Reagent Program</td><td>1069</td></tr><tr><td>pTrip-SFFV-ΔNGFR-2A</td><td>In house, Philippot et al.<sup>99</sup></td><td>N/A</td></tr><tr><td>pTrip-SFFV-mFLT3LG (T1)-ΔNGFR-2A</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pTrip-SFFV-sFLT3LG (T7)-ΔNGFR-2A</td><td>This manuscript</td><td>N/A</td></tr><tr><td>The mouse papillomavirus</td><td>HQ625439.1 (GenBank) cloned in PUC19</td><td>MmuPV1</td></tr><tr><td>Mouse papillomavirus L1</td><td>Brendle et al.<sup>96</sup></td><td>MmuPV1 L1</td></tr></table>

(Continued on next page)
Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td>pTrip-Rev5' CTCCTCTTGTGCTTCTAGCCAG 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>pTrip-FLT3L-For5' GAGAACCTGGACCT atgacagtgtggtcgcca 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>pTrip-FLT3L-Rev15' TTTTCTAGGTCTCGA tcagtgcctccacaagcagca 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3LG-S22P-For5' gctgctgagcccgggactcag 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3LG-S22P-Rev5' agcagcaggaggagatagg 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3LG-S118Afs-For5' ccccccccagctgtcttcgctt 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3LG-S118Afs-Rev5' ctgaaaggcacatttggt gacaaa 3'</td><td>Eurofins</td><td>N/A</td></tr><tr><td>FLT3LG-gDNA-For5' ATTTCACCACTCCAG CCTGG 3'</td><td>Thermo Fisher Scientific</td><td>N/A</td></tr><tr><td>FLT3LG-gDNA-Rev5' CCTTTAC CGGGCTGACACTG 3'</td><td>Thermo Fisher Scientific</td><td>N/A</td></tr><tr><td>Flt3lko mutant reverse5' ATT TGT CAC GTC CTG CAC GAC G 3'</td><td>Integrated DNA Technologies</td><td>N/A</td></tr><tr><td>Common forward5' TGG CAG CTG AAG TGA CTG AC 3'</td><td>Integrated DNA Technologies</td><td>N/A</td></tr><tr><td>Flt3l wild type reverse5' AAG CCA AAG CTG GAT GAC AG 3'</td><td>Integrated DNA Technologies</td><td>N/A</td></tr><tr><td>Forward primers for the mouse papillomavirus5'-TAGCTTTGTCTGCCCCGCACT-3'</td><td>Integrated DNA Technologies, Brendle et al.<sup>96</sup></td><td>N/A</td></tr><tr><td>Reverse primers for the mouse papillomavirus5'-GTCAGTGGTGTTCGGTGGGAA-3'</td><td>Integrated DNA Technologies, Brendle et al.<sup>96</sup></td><td>N/A</td></tr><tr><td>Probe for the mouse papillomavirus5'FAM-CG GCCCGAAGACAACACCGCCACG-3'TAMRA</td><td>Integrated DNA Technologies, Brendle et al.<sup>96</sup></td><td>N/A</td></tr><tr><td>FLT3LG</td><td>Thermo Fisher Scientific</td><td>Hs00181740_m1</td></tr><tr><td>FLT3LG</td><td>Thermo Fisher Scientific</td><td>Hs00953092_g1</td></tr><tr><td>GUSB</td><td>Thermo Fisher Scientific</td><td>4326320E</td></tr></tbody></table>

Recombinant DNA

<table><tr><td>pcDNA3.1/V5-His</td><td>Thermo Fisher Scientific</td><td>V79020</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T1/T2)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T3)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T4)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T5)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T6)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T7)</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-mFLT3LG (T1/2) -Ser118Alafs23*</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-mFLT3LG (T1/2)-Ser22Pro</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-sFLT3LG (T7)-Ser118Alafs23*</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pCDNA3.1/V5-His-FLT3LG (T7)-Ser22Pro</td><td>This manuscript</td><td>N/A</td></tr><tr><td>psPAX2</td><td>Addgene</td><td>12260</td></tr><tr><td>pCMV-VSV-G</td><td>Addgene</td><td>8454</td></tr><tr><td>pHXB2-Env</td><td>NIH-AIDS Reagent Program</td><td>1069</td></tr><tr><td>pTrip-SFFV-ΔNGFR-2A</td><td>In house, Philippot et al.<sup>99</sup></td><td>N/A</td></tr><tr><td>pTrip-SFFV-mFLT3LG (T1)-ΔNGFR-2A</td><td>This manuscript</td><td>N/A</td></tr><tr><td>pTrip-SFFV-sFLT3LG (T7)-ΔNGFR-2A</td><td>This manuscript</td><td>N/A</td></tr><tr><td>The mouse papillomavirus</td><td>HQ625439.1 (GenBank) cloned in PUC19</td><td>MmuPV1</td></tr><tr><td>Mouse papillomavirus L1</td><td>Brendle et al.<sup>96</sup></td><td>MmuPV1 L1</td></tr></table>

(Continued on next page)
Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td colspan="3">Software and algorithms</td></tr><tr><td>Eperico Image scope</td><td>Leicabiosystems</td><td>https://www.leicabiosystems.com/</td></tr><tr><td>R</td><td>The R Project for Statistical Computing</td><td>https://www.r-project.org/</td></tr><tr><td>Harmony</td><td>Korsunsky et al.<sup>100</sup></td><td>https://github.com/immunogenomics/harmony</td></tr><tr><td>Cell Ranger</td><td>10X Genomics</td><td>https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger</td></tr><tr><td>STAR (2.6.1d)</td><td>Dobin et al.<sup>101</sup></td><td>https://github.com/alexdobin/STAR</td></tr><tr><td>Seurat R package</td><td>Stuart et al.<sup>102</sup></td><td>https://cran.r-project.org/web/packages/Seurat/index.html</td></tr><tr><td>Burrows-Wheeler aligner</td><td>Li and Durbin<sup>103</sup></td><td>V0.7.12</td></tr><tr><td>GATK</td><td>McKenna et al.<sup>104</sup></td><td>https://www.broadinstitute.org/gatk</td></tr><tr><td>SAMtools</td><td>Li et al.<sup>105</sup></td><td>http://samtools.sourceforge.net/</td></tr><tr><td>muscat</td><td>Crowell et al.<sup>106</sup></td><td>https://bioconductor.org/packages/release/bioc/html/muscat.html</td></tr><tr><td>DESeq2</td><td>Love et al.<sup>107</sup></td><td>https://bioconductor.org/packages/release/bioc/html/DESeq2.html</td></tr><tr><td>Picard</td><td>http://broadinstitute.github.io/picard/</td><td>N/A</td></tr><tr><td>MERLIN 1.1.2 software</td><td>Abecasis et al.<sup>108</sup></td><td>http://csg.sph.umich.edu/abecasis/merlin/download/</td></tr><tr><td>Alamut Visual Plus</td><td>https://www.sophiagenetics.com/platform/alamut-visual-plus/</td><td>N/A</td></tr><tr><td>BioRender</td><td>https://biorender.com/</td><td>N/A</td></tr></tbody></table>

RESOURCE AVAILABILITY

Lead contact

Further information and requests for resources and reagents should be directed to and will be fulfilled by the lead contact, Vivien Beziat (vivien.beziat@inserm.fr).

Materials availability

Under a Material/Data Transfer Agreement (MTA) with INSERM or the Rockefeller University, all raw and processed data as well as biological materials are available upon request from the lead contact.

Data and code availability

The single-cell RNA-sequencing data have been deposited at NCBI BioProject and Biostudies EMBL-EBI, and are publicly available as of the publication date. The T cell and B cell receptor datasets have been deposited at Adaptive Biotechnologies. Accession numbers and access links are listed in the key resources table. There is no original code reported in this publication. Upon request, the lead contact will provide and share any more information or row data files needed to reanalyze the data presented in this work.

EXPERIMENTAL MODEL AND STUDY PARTICIPANT DETAILS

The age, sex, ethnicity, ancestry, and race of the studied patients is reported in the main text of the paper and in the method details below. Information on gender and socioeconomic status of the patients and healthy controls was not collected. Written informed consent (IC) was obtained from participants in accordance with local regulations in France, Belgium, the United States of America (USA), and Iran with approval from the appropriate institutional review board (IRB). Experiments were performed in France, Belgium, USA, Iran, and Qatar, in accordance with local regulations and with the approval of the IRB of INSERM and the Rockefeller University for France and the USA, respectively. Patients were included in the C18-41 Genetic Predisposition to Severe Infections study
Continued

<table><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td colspan="3">Software and algorithms</td></tr><tr><td>Eperico Image scope</td><td>Leicabiosystems</td><td>https://www.leicabiosystems.com/</td></tr><tr><td>R</td><td>The R Project for Statistical Computing</td><td>https://www.r-project.org/</td></tr><tr><td>Harmony</td><td>Korsunsky et al.<sup>100</sup></td><td>https://github.com/immunogenomics/harmony</td></tr><tr><td>Cell Ranger</td><td>10X Genomics</td><td>https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger</td></tr><tr><td>STAR (2.6.1d)</td><td>Dobin et al.<sup>101</sup></td><td>https://github.com/alexdobin/STAR</td></tr><tr><td>Seurat R package</td><td>Stuart et al.<sup>102</sup></td><td>https://cran.r-project.org/web/packages/Seurat/index.html</td></tr><tr><td>Burrows-Wheeler aligner</td><td>Li and Durbin<sup>103</sup></td><td>V0.7.12</td></tr><tr><td>GATK</td><td>McKenna et al.<sup>104</sup></td><td>https://www.broadinstitute.org/gatk</td></tr><tr><td>SAMtools</td><td>Li et al.<sup>105</sup></td><td>http://samtools.sourceforge.net/</td></tr><tr><td>muscat</td><td>Crowell et al.<sup>106</sup></td><td>https://bioconductor.org/packages/release/bioc/html/muscat.html</td></tr><tr><td>DESeq2</td><td>Love et al.<sup>107</sup></td><td>https://bioconductor.org/packages/release/bioc/html/DESeq2.html</td></tr><tr><td>Picard</td><td>http://broadinstitute.github.io/picard/</td><td>N/A</td></tr><tr><td>MERLIN 1.1.2 software</td><td>Abecasis et al.<sup>108</sup></td><td>http://csg.sph.umich.edu/abecasis/merlin/download/</td></tr><tr><td>Alamut Visual Plus</td><td>https://www.sophiagenetics.com/platform/alamut-visual-plus/</td><td>N/A</td></tr><tr><td>BioRender</td><td>https://biorender.com/</td><td>N/A</td></tr></tbody></table>

RESOURCE AVAILABILITY

Lead contact

Further information and requests for resources and reagents should be directed to and will be fulfilled by the lead contact, Vivien Beziat (vivien.beziat@inserm.fr).

Materials availability

Under a Material/Data Transfer Agreement (MTA) with INSERM or the Rockefeller University, all raw and processed data as well as biological materials are available upon request from the lead contact.

Data and code availability

The single-cell RNA-sequencing data have been deposited at NCBI BioProject and Biostudies EMBL-EBI, and are publicly available as of the publication date. The T cell and B cell receptor datasets have been deposited at Adaptive Biotechnologies. Accession numbers and access links are listed in the key resources table. There is no original code reported in this publication. Upon request, the lead contact will provide and share any more information or row data files needed to reanalyze the data presented in this work.

EXPERIMENTAL MODEL AND STUDY PARTICIPANT DETAILS

The age, sex, ethnicity, ancestry, and race of the studied patients is reported in the main text of the paper and in the method details below. Information on gender and socioeconomic status of the patients and healthy controls was not collected. Written informed consent (IC) was obtained from participants in accordance with local regulations in France, Belgium, the United States of America (USA), and Iran with approval from the appropriate institutional review board (IRB). Experiments were performed in France, Belgium, USA, Iran, and Qatar, in accordance with local regulations and with the approval of the IRB of INSERM and the Rockefeller University for France and the USA, respectively. Patients were included in the C18-41 Genetic Predisposition to Severe Infections study
approved by the Sud Est II ethics committee (approval 2022-A00257-36), and the ethics committee of Leuven University Hospital (S62030), and was performed in accordance with the requirements of these bodies. Bone marrow (BM) mononuclear cells were harvested from controls and patients after written, informed consent had been obtained. Control BM cells corresponded to the unused residue of allogeneic hematopoietic stem cells harvested from a healthy adult donor. The patients' BM cells were obtained from the residual BM after myelogram analysis as part of the diagnostic procedure. The constitution of a collection of BM samples was approved by the regional investigational review board and the French Ministry of Research (references: DC 2011-1338 and DC 2011-1421). The collection was used in accordance with French legislation and the ethical tenets of the Declaration of Helsinki. Virscan studies (on human subjects) were performed with informed written consent, and the procedures were approved by the Institutional Research Ethics Boards of Sidra Medicine and the Qatar Biobank. All work on mice was approved by the Institutional Animal Care and Use Committee of the Pennsylvania State University College of Medicine (PSUCOM), and all procedures were performed in strict accordance with guidelines and regulations.

## METHOD DETAILS

### Human patients

IC was obtained during the medical examination of the patients in France, Belgium, and residence country of the patients; Iran, in accordance with local regulations and with institutional review board (IRB) approval. A detailed clinical case report is provided below.

### Case reports

All three patients were siblings born to first-degree consanguineous Iranian parents (Figure 1A; Table S1).

P1 (female) was born in 1989. She suffered from failure to thrive (FTT) from the age of one year. From the age of five years, she presented recurrent infections, such as otitis media leading to hearing impairment, pulmonary infections, and severe disseminated warts (Figure 1B). She also suffered from chronic diarrhea from the age of five years, for which no clear etiology was established at the time. She displayed leukopenia and anemia from the age of seven years. At the age of 10 years, she developed polyarthritis. Tests for antinuclear antibodies and anti-neutrophil cytoplasmic antibodies were negative. P1 died from an undocumented invasive infection after a car accident at the age of 29 years.

P2 (male) was born in 1991. He was healthy until the age of one year, when he presented FTT and chronic diarrhea. Celiac disease was suspected, but the introduction of a gluten-free diet (GFD) led to no improvement. P2 also suffered from early-onset infectious complications: recurrent upper and lower respiratory tract infections (URTI & LRTI) with bilateral hearing impairment due to bilateral otosclerosis at the age of nine years, disseminated common warts from the age of four years (Figures 1B and S1A) and recurrent episodes of oral HSV infection and shingles. He had no adverse reaction to the BCG and MMR live attenuated vaccines. At recent evaluations at the ages of 28 and 31 years, P2 was cachectic and complained of diarrhea and fatigue. Physical examination revealed digital clubbing of the hands and feet, disseminated warts, reduced vesicular lung sounds with expiratory wheezing, and normal cardiac, abdominal and neurological examinations. Chest X ray was normal, and lung computed tomography (CT) revealed bilateral bronchial wall thickening, with no bronchiectasis or lymphadenopathy. P2 repeatedly complained of recurrent diarrhea and weight loss. Abdominal ultrasound revealed dilated intrahepatic biliary ducts; upper GI tract endoscopy revealed normal histological features of the duodenal mucosa; fecal calprotectin and elastase levels were normal; tests for anti-tissue transglutaminase (ATT), anti-endomysial (EMA) and anti-gliadin IgG were negative; IgA-EMA and IgA anti-gliadin tests were negative; IgA-ATT results were weakly positive (37.5 U/mL; normal < 15) in a context of polyclonal IgA hypergammaglobulinemia (38.2 g/L); high loads of microsporidia were observed in the stools (*Encephalitozoon/Enterocytozoon*); HPEV-1 replication rates were high in the stools whereas this virus was only weakly detectable in blood. P2 was diagnosed with chronic cholangitis, possibly due to chronic microsporidiosis (with biliary infection) and plausible chronic HPEV-1 infection. See Table S1 for the detailed laboratory findings.

P3 (female) was born in 1995. Like P2, she presented FTT and diarrhea from the age of one year. Celiac disease was suspected but the introduction of a gluten-free diet (GFD) yielded no improvement, as in P2. P3 also suffered from early-onset infectious complications: recurrent URTI with chronic otorrhea and hearing loss, and recurrent LRTI; HPV infection with disseminated HPV2⁺ common warts from the age of four years (Figure 1B), disseminated HPV5⁺ flat warts (Figures 1B and S1A); recurrent episodes of oral HSV infection; and episodes of *Staphylococcus aureus* cellulitis. P3 displayed no adverse reaction to the BCG and MMR live attenuated vaccines. P3 also presented seronegative idiopathic polyarthritis and Raynaud's phenomenon from the age of four years, with intermittent swelling affecting the knees, fingers, and shoulders. At recent evaluations at the ages of 24 and 28 years, P3 was cachectic and also complained of diarrhea, chronic cough with yellowish sputum and fatigue. Physical examination found reduced lung vesicular sounds, verrucous plaques and nodules on the hands, feet, lower arms and soles, subungual verruca and HPV7⁺ anogenital warts, with normal cardiac, abdominal and neurological examinations. Chest X ray was normal and lung computed tomography (CT) showed zones of air trapping suggestive of small airway disease (probably constrictive bronchiolitis due to recurrent infections), without bronchiectasis or lymphadenopathy. P3's chronic diarrhea was investigated further: abdominal ultrasound revealed a dilated bile duct and intrahepatic biliary ducts; upper GI tract endoscopy showed mild neutrophilic duodenitis and no signs suggestive of celiac disease; fecal calprotectin and elastase levels were normal; negative results were obtained in tests for IgG and IgA-ATT, IgG and IgA-EMA, IgA and IgA anti-gliadin; fecal loads of microsporidia were high (*Encephalitozoon/Enterocytozoon*), and high rates
approved by the Sud Est II ethics committee (approval 2022-A00257-36), and the ethics committee of Leuven University Hospital (S62030), and was performed in accordance with the requirements of these bodies. Bone marrow (BM) mononuclear cells were harvested from controls and patients after written, informed consent had been obtained. Control BM cells corresponded to the unused residue of allogeneic hematopoietic stem cells harvested from a healthy adult donor. The patients' BM cells were obtained from the residual BM after myelogram analysis as part of the diagnostic procedure. The constitution of a collection of BM samples was approved by the regional investigational review board and the French Ministry of Research (references: DC 2011-1338 and DC 2011-1421). The collection was used in accordance with French legislation and the ethical tenets of the Declaration of Helsinki. Virscan studies (on human subjects) were performed with informed written consent, and the procedures were approved by the Institutional Research Ethics Boards of Sidra Medicine and the Qatar Biobank. All work on mice was approved by the Institutional Animal Care and Use Committee of the Pennsylvania State University College of Medicine (PSUCOM), and all procedures were performed in strict accordance with guidelines and regulations.

## METHOD DETAILS

### Human patients

IC was obtained during the medical examination of the patients in France, Belgium, and residence country of the patients; Iran, in accordance with local regulations and with institutional review board (IRB) approval. A detailed clinical case report is provided below.

### Case reports

All three patients were siblings born to first-degree consanguineous Iranian parents (Figure 1A; Table S1).

P1 (female) was born in 1989. She suffered from failure to thrive (FTT) from the age of one year. From the age of five years, she presented recurrent infections, such as otitis media leading to hearing impairment, pulmonary infections, and severe disseminated warts (Figure 1B). She also suffered from chronic diarrhea from the age of five years, for which no clear etiology was established at the time. She displayed leukopenia and anemia from the age of seven years. At the age of 10 years, she developed polyarthritis. Tests for antinuclear antibodies and anti-neutrophil cytoplasmic antibodies were negative. P1 died from an undocumented invasive infection after a car accident at the age of 29 years.

P2 (male) was born in 1991. He was healthy until the age of one year, when he presented FTT and chronic diarrhea. Celiac disease was suspected, but the introduction of a gluten-free diet (GFD) led to no improvement. P2 also suffered from early-onset infectious complications: recurrent upper and lower respiratory tract infections (URTI & LRTI) with bilateral hearing impairment due to bilateral otosclerosis at the age of nine years, disseminated common warts from the age of four years (Figures 1B and S1A) and recurrent episodes of oral HSV infection and shingles. He had no adverse reaction to the BCG and MMR live attenuated vaccines. At recent evaluations at the ages of 28 and 31 years, P2 was cachectic and complained of diarrhea and fatigue. Physical examination revealed digital clubbing of the hands and feet, disseminated warts, reduced vesicular lung sounds with expiratory wheezing, and normal cardiac, abdominal and neurological examinations. Chest X ray was normal, and lung computed tomography (CT) revealed bilateral bronchial wall thickening, with no bronchiectasis or lymphadenopathy. P2 repeatedly complained of recurrent diarrhea and weight loss. Abdominal ultrasound revealed dilated intrahepatic biliary ducts; upper GI tract endoscopy revealed normal histological features of the duodenal mucosa; fecal calprotectin and elastase levels were normal; tests for anti-tissue transglutaminase (ATT), anti-endomysial (EMA) and anti-gliadin IgG were negative; IgA-EMA and IgA anti-gliadin tests were negative; IgA-ATT results were weakly positive (37.5 U/mL; normal < 15) in a context of polyclonal IgA hypergammaglobulinemia (38.2 g/L); high loads of microsporidia were observed in the stools (*Encephalitozoon/Enterocytozoon*); HPEV-1 replication rates were high in the stools whereas this virus was only weakly detectable in blood. P2 was diagnosed with chronic cholangitis, possibly due to chronic microsporidiosis (with biliary infection) and plausible chronic HPEV-1 infection. See Table S1 for the detailed laboratory findings.

P3 (female) was born in 1995. Like P2, she presented FTT and diarrhea from the age of one year. Celiac disease was suspected but the introduction of a gluten-free diet (GFD) yielded no improvement, as in P2. P3 also suffered from early-onset infectious complications: recurrent URTI with chronic otorrhea and hearing loss, and recurrent LRTI; HPV infection with disseminated HPV2⁺ common warts from the age of four years (Figure 1B), disseminated HPV5⁺ flat warts (Figures 1B and S1A); recurrent episodes of oral HSV infection; and episodes of *Staphylococcus aureus* cellulitis. P3 displayed no adverse reaction to the BCG and MMR live attenuated vaccines. P3 also presented seronegative idiopathic polyarthritis and Raynaud's phenomenon from the age of four years, with intermittent swelling affecting the knees, fingers, and shoulders. At recent evaluations at the ages of 24 and 28 years, P3 was cachectic and also complained of diarrhea, chronic cough with yellowish sputum and fatigue. Physical examination found reduced lung vesicular sounds, verrucous plaques and nodules on the hands, feet, lower arms and soles, subungual verruca and HPV7⁺ anogenital warts, with normal cardiac, abdominal and neurological examinations. Chest X ray was normal and lung computed tomography (CT) showed zones of air trapping suggestive of small airway disease (probably constrictive bronchiolitis due to recurrent infections), without bronchiectasis or lymphadenopathy. P3's chronic diarrhea was investigated further: abdominal ultrasound revealed a dilated bile duct and intrahepatic biliary ducts; upper GI tract endoscopy showed mild neutrophilic duodenitis and no signs suggestive of celiac disease; fecal calprotectin and elastase levels were normal; negative results were obtained in tests for IgG and IgA-ATT, IgG and IgA-EMA, IgA and IgA anti-gliadin; fecal loads of microsporidia were high (*Encephalitozoon/Enterocytozoon*), and high rates
of HPEV-1 replication were detected in stools. We concluded that P3 suffered from chronic cholangitis, possibly due to chronic mi-crosporidiosis (with biliary infection) and plausible chronic HPEV-1 infection. See Table S1 for detailed laboratory findings.

## Whole-exome sequencing (WES) and linkage analysis (LA)

Genomic DNA was isolated from peripheral blood collected from patients, parents, and siblings. WES was performed for P1, P2, P3, their parents and healthy siblings. The DNA preparation was enriched in exons with the SureSelect XT Library prep Kit (Agilent Technologies, CA, USA). Sequencing was performed on an Illumina HiSeq 4000 (San Diego, CA, USA) sequencer with paired-end sequencing, a read length of 101 bp and 100× coverage for the three patients or with a NovaSeq 6000 (Illumina, CA, USA) sequencer with a read length of 150 bp and 100× coverage for the other members of the family, by Macrogen (Macrogen, Inc., Seoul, South Korea). Burrows-Wheeler aligner (BWA) was used to align the reads with the reference genome sequence (GRCh37/hg19).¹⁰³ GATK's Haplotype Caller from the Genome Analysis Toolkit (GATK version 3.6)¹⁰⁴ SAMtools,¹⁰⁵ and Picard tools were used for variant calling. All variants were annotated with our in-house annotation software.²⁷,¹⁰⁹,¹¹⁰ The homozygosity rate was calculated as previously described.¹¹¹ We used the WES data for linkage analysis. Assuming a disorder with AR inheritance and complete penetrance, multi-point LOD scores were calculated with MERLIN software.¹⁰⁸

We focused principally on protein coding sequence-altering variants (nonsense, splice-site variants, coding indels, and missense variants) with minor allele frequencies (MAFs) in the general population reported in the gnomAD database (v2.1.1), and our in-home exome database including >20000 exomes. Variants with high combined annotation-dependent deletion (CADD) (v1.6) scores were filtered out. The mutation significance cutoff (MSC) was calculated as previously described.¹¹²

## Sanger sequencing

Genomic DNA samples were extracted from peripheral blood from patients, their parents, and healthy siblings. The region encompassing the FLT3LG mutation was amplified by PCR. The PCR products were sequenced by standard Sanger sequencing with the BigDye Terminator Cycle Sequencing Kit (Applied Biosystems). Sequencing reaction products were purified by centrifugation through Sephadex G-50 Superfine resin. Sequencing was performed on an ABI 3500 DNA sequencer (Applied Biosystems, Life Technologies, CA, USA). The primers used for sequencing were those used for PCR. Sequence data were aligned with the genomic sequence of FLT3LG (NCBI) with Benchling (https://benchling.com).

## Metagenomic next-generation sequencing (mNGS)

Urine samples and stools from P2 and P3 were analyzed by metagenomics next-generation sequencing (mNGS) to search for known or unexpected microorganisms. Samples were prepared and sequenced at high sequencing depth in a context of routine mNGS analyses of the patient's samples, as previously described.¹¹³ Briefly, total nucleic acids were extracted with (stools) or without (urine) prior nuclease treatment. They were then subjected to reverse transcription with random primers and pre-amplification, before production of the final NGS libraries. Sequencing was performed on an Illumina NextSeq500 instrument, generating the following numbers of raw reads: 71.1 M (urine P2), 64.0 M (urine P3), 55.0 M (stools P2), 65.7 M (stools P3). Microseek was used for the detection of microorganisms.¹¹⁴

## Cell culture and isolation

### Primary cells

Peripheral blood mononuclear cells (PBMCs) were isolated by Ficoll-Hypaque centrifugation (Merck) from cytopheresis products from patients or healthy donors. Cells were either used fresh or were cryopreserved and stored in liquid nitrogen until use. Primary T-cell blasts were generated by culturing PBMCs in ImmunoCult™-XF T Cell Expansion Medium (STEMCELL Technologies) supplemented with 10 ng/mL rIL-2 (Thermo Fisher Scientific) (referred to hereafter as T-cell medium or TCM) with replacement of the medium every 48 to 72 hours. Cells were activated with soluble antibody complex reagents (ImmunoCult™ Human CD3/CD28/CD2 T-Cell Activator; STEMCELL Technologies) at an initial dilution of 1:80, with reactivation at two-week intervals to ensure continued growth in culture.

### Cell lines

The HEK293T cell line was purchased from the American Type Culture Collection (ATCC) and cultured in DMEM supplemented with 10% FBS (Gibco). The K562 cell line (kindly donated by Prof. Jerome Tamburini, Institut Cochin) was cultured in RPMI-1640 medium with GlutaMAX (Gibco) supplemented with 10% FBS. EBV-B cell lines were generated in-house by infecting PBMCs with EBV. EBV-B cells were cultured in RPMI-1640 medium with GlutaMAX (Gibco) supplemented with 10% FBS.

### RNA-seq analyses

We determined the splicing events occurring in different immune cell types by re-analyzing published RNA-seq datasets for human cells (GSE107011).¹¹⁵ We downloaded the raw sequence files from the Gene Expression Omnibus (GEO) with the SRA toolkit (fastq-dump), assessed their quality with FastQC (Babraham Bioinformatics), and removed low-quality reads and bases with Trimmomatic v.0.33.¹¹⁶ The biological replicates for each cell type were aligned with the hg38 reference human genome assembly with HISAT2 v2.2.1¹¹⁷ and were combined to obtain greater coverage at exon-splicing junctions. The resulting SAM files were converted to
of HPEV-1 replication were detected in stools. We concluded that P3 suffered from chronic cholangitis, possibly due to chronic mi-crosporidiosis (with biliary infection) and plausible chronic HPEV-1 infection. See Table S1 for detailed laboratory findings.

## Whole-exome sequencing (WES) and linkage analysis (LA)

Genomic DNA was isolated from peripheral blood collected from patients, parents, and siblings. WES was performed for P1, P2, P3, their parents and healthy siblings. The DNA preparation was enriched in exons with the SureSelect XT Library prep Kit (Agilent Technologies, CA, USA). Sequencing was performed on an Illumina HiSeq 4000 (San Diego, CA, USA) sequencer with paired-end sequencing, a read length of 101 bp and 100× coverage for the three patients or with a NovaSeq 6000 (Illumina, CA, USA) sequencer with a read length of 150 bp and 100× coverage for the other members of the family, by Macrogen (Macrogen, Inc., Seoul, South Korea). Burrows-Wheeler aligner (BWA) was used to align the reads with the reference genome sequence (GRCh37/hg19).¹⁰³ GATK's Haplotype Caller from the Genome Analysis Toolkit (GATK version 3.6)¹⁰⁴ SAMtools,¹⁰⁵ and Picard tools were used for variant calling. All variants were annotated with our in-house annotation software.²⁷,¹⁰⁹,¹¹⁰ The homozygosity rate was calculated as previously described.¹¹¹ We used the WES data for linkage analysis. Assuming a disorder with AR inheritance and complete penetrance, multi-point LOD scores were calculated with MERLIN software.¹⁰⁸

We focused principally on protein coding sequence-altering variants (nonsense, splice-site variants, coding indels, and missense variants) with minor allele frequencies (MAFs) in the general population reported in the gnomAD database (v2.1.1), and our in-home exome database including >20000 exomes. Variants with high combined annotation-dependent deletion (CADD) (v1.6) scores were filtered out. The mutation significance cutoff (MSC) was calculated as previously described.¹¹²

## Sanger sequencing

Genomic DNA samples were extracted from peripheral blood from patients, their parents, and healthy siblings. The region encompassing the FLT3LG mutation was amplified by PCR. The PCR products were sequenced by standard Sanger sequencing with the BigDye Terminator Cycle Sequencing Kit (Applied Biosystems). Sequencing reaction products were purified by centrifugation through Sephadex G-50 Superfine resin. Sequencing was performed on an ABI 3500 DNA sequencer (Applied Biosystems, Life Technologies, CA, USA). The primers used for sequencing were those used for PCR. Sequence data were aligned with the genomic sequence of FLT3LG (NCBI) with Benchling (https://benchling.com).

## Metagenomic next-generation sequencing (mNGS)

Urine samples and stools from P2 and P3 were analyzed by metagenomics next-generation sequencing (mNGS) to search for known or unexpected microorganisms. Samples were prepared and sequenced at high sequencing depth in a context of routine mNGS analyses of the patient's samples, as previously described.¹¹³ Briefly, total nucleic acids were extracted with (stools) or without (urine) prior nuclease treatment. They were then subjected to reverse transcription with random primers and pre-amplification, before production of the final NGS libraries. Sequencing was performed on an Illumina NextSeq500 instrument, generating the following numbers of raw reads: 71.1 M (urine P2), 64.0 M (urine P3), 55.0 M (stools P2), 65.7 M (stools P3). Microseek was used for the detection of microorganisms.¹¹⁴

## Cell culture and isolation

### Primary cells

Peripheral blood mononuclear cells (PBMCs) were isolated by Ficoll-Hypaque centrifugation (Merck) from cytopheresis products from patients or healthy donors. Cells were either used fresh or were cryopreserved and stored in liquid nitrogen until use. Primary T-cell blasts were generated by culturing PBMCs in ImmunoCult™-XF T Cell Expansion Medium (STEMCELL Technologies) supplemented with 10 ng/mL rIL-2 (Thermo Fisher Scientific) (referred to hereafter as T-cell medium or TCM) with replacement of the medium every 48 to 72 hours. Cells were activated with soluble antibody complex reagents (ImmunoCult™ Human CD3/CD28/CD2 T-Cell Activator; STEMCELL Technologies) at an initial dilution of 1:80, with reactivation at two-week intervals to ensure continued growth in culture.

### Cell lines

The HEK293T cell line was purchased from the American Type Culture Collection (ATCC) and cultured in DMEM supplemented with 10% FBS (Gibco). The K562 cell line (kindly donated by Prof. Jerome Tamburini, Institut Cochin) was cultured in RPMI-1640 medium with GlutaMAX (Gibco) supplemented with 10% FBS. EBV-B cell lines were generated in-house by infecting PBMCs with EBV. EBV-B cells were cultured in RPMI-1640 medium with GlutaMAX (Gibco) supplemented with 10% FBS.

## RNA-seq analyses

We determined the splicing events occurring in different immune cell types by re-analyzing published RNA-seq datasets for human cells (GSE107011).¹¹⁵ We downloaded the raw sequence files from the Gene Expression Omnibus (GEO) with the SRA toolkit (fastq-dump), assessed their quality with FastQC (Babraham Bioinformatics), and removed low-quality reads and bases with Trimmomatic v.0.33.¹¹⁶ The biological replicates for each cell type were aligned with the hg38 reference human genome assembly with HISAT2 v2.2.1¹¹⁷ and were combined to obtain greater coverage at exon-splicing junctions. The resulting SAM files were converted to
BAM format, sorted, and indexed with samtools v1.12.¹⁰⁵ The BAM files were loaded into Integrated Genome Viewer (IGV) to visualize the reads aligned with the FLT3LG gene and the spliced reads were counted with the Sashimi plot function. ¹¹⁸

## mRNA extraction, cDNA synthesis, and assessment of gene expression by RT-PCR/RT-qPCR

Total RNA was isolated from the indicated cells with the RNeasy Extraction Kit (QIAGEN) or the Quick-RNA Microprep Kit (Zymo Research Corporation). RNA samples were reverse-transcribed with SuperScript II reverse transcriptase (Thermo Fisher Scientific) and oligo dT primers (Thermo Fisher Scientific) or the High-Capacity RNA-to-cDNA™ Kit (Thermo Fisher Scientific), in accordance with the manufacturer's instructions. RT-qPCR was performed with the TaqMan Gene Expression Master Mix (Applied Biosystems) and specific FAM-MGB probes for FLT3LG (Hs00953092_g1 and Hs00181740_m1) (Thermo Fisher Scientific). A human GUSB-VIC probe (4326320E, Thermo Fisher Scientific) was used to normalize the data. RT-qPCR amplification was performed on the Applied Biosystems VIIA 7 system. Relative expression is expressed as 2⁻ΔCT values after normalization against GUSB (endogenous control) expression (ΔCt).

## Characterization of FLT3LG mRNA splice variants by Topo cloning

Bulk cDNA samples were generated as previous described from the T-cell blasts of P3, the mother, and two unrelated healthy donors. The cDNA from the FLT3LG transcripts was amplified with CloneAmp HiFi PCR Premix (Takara) and the following primers binding to exons 1 and 9, respectively, covering the full coding sequence: forward primer: 5' TTTCGGTCTCTGGCTGTCAC-3', reverse primer: 5'-CTGTGTCCAGGCTATGCATC-3'. PCR products were inserted into the pCR™-Blunt II-TOPO® vector with the Zero blunt TOPO PCR cloning kit (Thermo Fisher Scientific) and were used to transform NEB 10-beta competent E. coli cells (New England Biolabs). The bacteria then were spread on LB-agar plates supplemented with kanamycin and incubated overnight at 37°C. Individual colonies were picked, amplified in DreamTaq Green PCR master mix (Thermo Fisher Scientific), and sequenced with the M13 forward and reverse primers from the TOPO cloning kit. The PCR products were subjected to Sanger sequencing as described above and the sequence obtained was aligned with the FLT3LG cDNA (GenBank: NM_001459.4) map with the Benchling application for characterization of the alternative splice variants.

## Cloning and mutagenesis

For the seven principal transcripts identified, as described above, the full-length WT FLT3L coding sequence (obtained from a cDNA from a healthy donor) was inserted into the pcDNA3.1D plasmid containing a V5 epitope/His by PCR with specific primers for each locus, with the directional TOPO expression kit (Thermo Fisher Scientific). The Tag sequence was located directly after the last coding codon. The patients' frameshift variant (c.343delC) and the only missense variant reported in the homozygous state in the gnomAD database (https://gnomad.broadinstitute.org/) were constructed by site-directed mutagenesis with CloneAmp Hifi premix (Takara). DpnI (#R0176L, New England Biolabs) was used to digest the resulting PCR products (2 h at 37°C). The product was amplified in NEB 10-beta competent E. coli cells (New England Biolabs) and purified with a maxiprep kit (Qiagen) according to the manufacturer's recommendations. Lentiviral plasmids carrying both the functional FLT3LG transcripts (T1 & T7) were inserted into the empty pTRIP-SFFV-deltaNGFR vector (modified from pTRIP-SFFV-mtagBFP-2A; addgene, plasmid #102585) by PCR with modified primers. The empty pTRIP-SFFV-ΔNGFR vector was digested with XhoI and BamHI for 1 h at 37°C, and the cDNA of interest, previously amplified by PCR (from pcDNA plasmid), was inserted by homologous recombination with the In-Fusion® HD Cloning Kit, according to the manufacturer's instructions (#638911, Takara). Each coding sequence was confirmed by Sanger sequencing.

## Transient transfection

HEK293T cells were used to seed six-well plates at a density of 1 × 10⁵ cells per well and were incubated overnight. The cells were then transiently transfected with the various constructs in the presence of Opti-MEM (Thermo Fisher Scientific) and X-tremeGENE 9 DNA Transfection Reagent (#6365787001, Roche), in accordance with the manufacturers' instructions, and incubated for 24-72 h before use in various experiments.

## Cell lysates and analysis of the FLT3L protein by western blotting

Cells were washed with cold PBS and lysed in 100 µL RIPA buffer (50 mM Tris pH 7.4, 150 mM NaCl, 2 mM EDTA, 1% NP-40) supplemented with a mixture of protease and phosphatase inhibitors added to the buffer immediately before use: aprotinin (Sigma, 10 mg/mL), PMSF (Sigma, 1 mM), leupeptin (Sigma, 10 mg/mL), phosSTOP (Roche), and cOmplete Mini Protease Inhibitor Cocktail (Roche). Lysates were incubated for 30 minutes at 4°C and mixed by vortexing every 10 min. The cells were centrifuged for 20 minutes at 16000 x g and 4°C, and the supernatant was collected for immunoblotting. Protein yield was determined with the Bradford protein assay (Bio-Rad), and equal amounts of total protein were separated by SDS-PAGE (10% polyacrylamide gel). A recombinant human FLT3L protein (R&D, 308-FKHB-010) was used as positive control. Proteins were transferred onto a polyvinylidene difluoride (PVDF) membrane in a wet transfer system (Bio-Rad). The membrane was blocked by incubation with a blocking buffer (5% BSA or milk in TBS) for 1 h at room temperature. Immunoblotting was performed with the following primary antibodies (overnight incubation at 4°C): anti-Flt 3-L antibody (F-6) HRP (sc-365266 HRP, Santa Cruz), anti-human Flt-3 ligand/FLT3L antibody (MAB308-100, R&D Systems), rabbit anti-FLT3 (8F2) mAb (3462, Cell Signaling), rabbit anti-phospho-FLT3 (Tyr589/591) (30D4) mAb (3464, Cell Signaling), anti-V5 tag antibody (R962-25, Thermo Fisher Scientific) and anti-vinculin antibody (sc-73614 HRP, Santa Cruz). All the primary antibodies
BAM format, sorted, and indexed with samtools v1.12.¹⁰⁵ The BAM files were loaded into Integrated Genome Viewer (IGV) to visualize the reads aligned with the *FLT3LG* gene and the spliced reads were counted with the Sashimi plot function. ¹¹⁸

## mRNA extraction, cDNA synthesis, and assessment of gene expression by RT-PCR/RT-qPCR

Total RNA was isolated from the indicated cells with the RNeasy Extraction Kit (QIAGEN) or the Quick-RNA Microprep Kit (Zymo Research Corporation). RNA samples were reverse-transcribed with SuperScript II reverse transcriptase (Thermo Fisher Scientific) and oligo dT primers (Thermo Fisher Scientific) or the High-Capacity RNA-to-cDNA™ Kit (Thermo Fisher Scientific), in accordance with the manufacturer's instructions. RT-qPCR was performed with the TaqMan Gene Expression Master Mix (Applied Biosystems) and specific FAM-MGB probes for FLT3LG (Hs00953092_g1 and Hs00181740_m1) (Thermo Fisher Scientific). A human GUSB-VIC probe (4326320E, Thermo Fisher Scientific) was used to normalize the data. RT-qPCR amplification was performed on the Applied Biosystems VIIA 7 system. Relative expression is expressed as 2^ΔCt^ values after normalization against GUSB (endogenous control) expression (ΔCt).

## Characterization of FLT3LG mRNA splice variants by Topo cloning

Bulk cDNA samples were generated as previous described from the T-cell blasts of P3, the mother, and two unrelated healthy donors. The cDNA from the FLT3LG transcripts was amplified with CloneAmp HiFi PCR Premix (Takara) and the following primers binding to exons 1 and 9, respectively, covering the full coding sequence: forward primer: 5' TTTCGGTCTCTGGCTGTCAC-3', reverse primer: 5'-CTGTGTCCAGGCTATGCATC-3'. PCR products were inserted into the pCR™-Blunt II-TOPO® vector with the Zero blunt TOPO PCR cloning kit (Thermo Fisher Scientific) and were used to transform NEB 10-beta competent E. coli cells (New England Biolabs). The bacteria then were spread on LB-agar plates supplemented with kanamycin and incubated overnight at 37°C. Individual colonies were picked, amplified in DreamTaq Green PCR master mix (Thermo Fisher Scientific), and sequenced with the M13 forward and reverse primers from the TOPO cloning kit. The PCR products were subjected to Sanger sequencing as described above and the sequence obtained was aligned with the FLT3LG cDNA (GenBank: NM_001459.4) map with the Benchling application for characterization of the alternative splice variants.

## Cloning and mutagenesis

For the seven principal transcripts identified, as described above, the full-length WT FLT3L coding sequence (obtained from a cDNA from a healthy donor) was inserted into the pcDNA3.1D plasmid containing a V5 epitope/His by PCR with specific primers for each locus, with the directional TOPO expression kit (Thermo Fisher Scientific). The Tag sequence was located directly after the last coding codon. The patients' frameshift variant (c.343delC) and the only missense variant reported in the homozygous state in the gnomAD database (https://gnomad.broadinstitute.org/) were constructed by site-directed mutagenesis with CloneAmp Hifi premix (Takara). DpnI (#R0176L, New England Biolabs) was used to digest the resulting PCR products (2 h at 37°C). The product was amplified in NEB 10-beta competent E. coli cells (New England Biolabs) and purified with a maxiprep kit (Qiagen) according to the manufacturer's recommendations. Lentiviral plasmids carrying both the functional FLT3LG transcripts (T1 & T7) were inserted into the empty pTRIP-SFFV-deltaNGFR vector (modified from pTRIP-SFFV-mtagBFP-2A; addgene, plasmid #102585) by PCR with modified primers. The empty pTRIP-SFFV-ΔNGFR vector was digested with XhoI and BamHI for 1 h at 37°C, and the cDNA of interest, previously amplified by PCR (from pcDNA plasmid), was inserted by homologous recombination with the In-Fusion® HD Cloning Kit, according to the manufacturer's instructions (#638911, Takara). Each coding sequence was confirmed by Sanger sequencing.

## Transient transfection

HEK293T cells were used to seed six-well plates at a density of 1 × 10⁵ cells per well and were incubated overnight. The cells were then transiently transfected with the various constructs in the presence of Opti-MEM (Thermo Fisher Scientific) and X-tremeGENE 9 DNA Transfection Reagent (#6365787001, Roche), in accordance with the manufacturers' instructions, and incubated for 24-72 h before use in various experiments.

## Cell lysates and analysis of the FLT3L protein by western blotting

Cells were washed with cold PBS and lysed in 100 µL RIPA buffer (50 mM Tris pH 7.4, 150 mM NaCl, 2 mM EDTA, 1% NP-40) supplemented with a mixture of protease and phosphatase inhibitors added to the buffer immediately before use: aprotinin (Sigma, 10 mg/mL), PMSF (Sigma, 1 mM), leupeptin (Sigma, 10 mg/mL), phosSTOP (Roche), and cOmplete Mini Protease Inhibitor Cocktail (Roche). Lysates were incubated for 30 minutes at 4°C and mixed by vortexing every 10 min. The cells were centrifuged for 20 minutes at 16000 x g and 4°C, and the supernatant was collected for immunoblotting. Protein yield was determined with the Bradford protein assay (Bio-Rad), and equal amounts of total protein were separated by SDS-PAGE (10% polyacrylamide gel). A recombinant human FLT3L protein (R&D, 308-FKHB-010) was used as positive control. Proteins were transferred onto a polyvinylidene difluoride (PVDF) membrane in a wet transfer system (Bio-Rad). The membrane was blocked by incubation with a blocking buffer (5% BSA or milk in TBS) for 1 h at room temperature. Immunoblotting was performed with the following primary antibodies (overnight incubation at 4°C): anti-Flt 3-L antibody (F-6) HRP (sc-365266 HRP, Santa Cruz), anti-human Flt-3 ligand/FLT3L antibody (MAB308-100, R&D Systems), rabbit anti-FLT3 (8F2) mAb (3462, Cell Signaling), rabbit anti-phospho-FLT3 (Tyr589/591) (30D4) mAb (3464, Cell Signaling), anti-V5 tag antibody (R962-25, Thermo Fisher Scientific) and anti-vinculin antibody (sc-73614 HRP, Santa Cruz). All the primary antibodies
were diluted in blocking buffer. For primary Abs not conjugated to HRP, membranes were washed in wash buffer (0.1% Tween 20 in PBS) and incubated with the following secondary antibodies: goat anti-mouse IgG (H + L)-HRP conjugate (1706516, Bio-Rad) or goat anti-rabbit IgG (H + L)-HRP conjugate (1706515, Bio-Rad). Staining was detected with the Clarity Western ECL substrate (Biorad, #1705061) or SuperSignal West Femto (Thermo Fisher Scientific, #34096) with ChemiDoc MP (Biorad). The images were analyzed with Image Lab 6.1 software (Biorad).

## Lentivirus transduction

HEK293T cells were used to seed a six-well plate at a density of $5 \times 10^5$ cells per well, in 2 mL per well DMEM supplemented with 10% FBS, two days before transduction. The cells were then transfected with pCMV-VSV-G (0.2 µg), pHXB2-env (0.2 µg; NIH-AIDS Reagent Program; #1069), psPAX2 (1 µg; a gift from D. Trono; Addgene plasmid #12260) and pTRIP-SFFV-deltaNGFR EV vector or pTRIP-SFFV-deltaNGFR containing the mFL or sFL coding sequences (1.6 µg) in Opti-MEM (Gibco; 300 µl) plus 9 µL X-tremeGENE 9 (Roche), according to the manufacturers' protocol. After 6 h of transfection, the medium was replaced with 3 mL DMEM supplemented with 10% FBS, and the cells were incubated for a further 24 h to produce lentiviral particles. On the day of HEK293T cell transfection, T-cell blasts from patients and controls were thawed, placed in TCM and reactivated with ImmunoCult™ Human CD3/CD28/CD2 T-Cell Activator (STEMCELL Technologies) in a 96-well plate, at a density of $2 \times 10^6$ cells/mL in a total volume of 100 µL/well. The next day (day 0), the supernatant of the HEK293T cells was collected and passed through a filter with 0.2 mm pores. Protamine sulfate (Sigma; $10 \mu g mL^{-1}$; 8 mg/mL) was added to the viral supernatant, and 100 µL of the supernatant was then added to the activated T cells, which were subjected to spinoculation for 2 hours at 1200 x g and room temperature. The spinoculated cells were then incubated for 48 hours at 37°C under an atmosphere containing 5% CO₂. After two days, the cells were harvested and transferred to small flasks containing TCM supplemented with 1% penicillin/streptomycin.

## Soluble FLT3L determination

The soluble form of FLT3L was determined in plasma or supernatant collected from cell lines expressing the protein (HEK293T cells and T-cell blasts) by sandwich ELISA (Human Flt-3 Ligand/FLT3L DuoSet ELISA, DY308, R&D Systems) in duplicate and in more than two independent experiments, in accordance with the manufacturer's protocol. The data obtained were subjected to statistical analysis by the regression analysis approach, in accordance with the instructions supplied with the kit.

## Analysis of FLT3L expression by flow cytometry

Transiently transfected HEK293T cells were harvested from the wells and washed once with PBS and cold FACS buffer (2% FBS in PBS). The cells were then stained simultaneously with a mixture of Live/Dead Fixable Aqua Dead Cell marker (reconstituted in DMSO; used at a 1:100 dilution in FACS buffer) and a primary Ab against FLT3L (recombinant anti-Flt3 ligand [EP1140Y] (ab52648), Abcam) or an isotype control (recombinant rabbit IgG, monoclonal [EPR25A] - (ab172730), Abcam) antibody for 60 min at 4°C in the dark. The cells were washed three times with FACS buffer and stained with a PE-conjugated goat anti-rabbit IgG (H+L) secondary antibody (A10542, Thermo Fisher Scientific) at a 1:500 dilution, for 30-60 min at 4°C in the dark. Cells were washed twice with FACS buffer and acquired with a Gallios (Beckman Coulter) machine. Data were then analyzed with FlowJo software v10. FLT3L expression on the cell surface of T-cell blasts (non-transduced and transduced) was analyzed with the same staining and analysis procedure.

## Deep immunophenotyping of whole blood

Deep immunophenotyping was performed by both conventional flow cytometry and mass cytometry (CyTOF) methods. For CyTOF, we used 200 µL fresh whole blood for the patients and controls. We used a custom-designed panel with antibodies indicated in STAR Methods, according to Fluidigm's instructions. Cells were frozen at -80°C after overnight dead-cell staining and acquisition was performed on a Helios mass cytometer (Fluidigm). All the samples were processed within 24 hours of sampling. Data analysis was performed with OMIQ software.

For classical immunophenotyping of the patients, fresh whole-blood samples were stained by incubation for 30 minutes at room temperature with antibodies against cell-surface markers, indicated in STAR Methods, and analyzed on a BD FACSLyric™ flow cytometer.

For a more detailed analysis of DC subsets, more than 10 M ($1 \times 10^7$) freshly isolated PBMCs were subjected to cell-surface staining with the following Abs: anti-CD3 FITC (#555332, BD), anti-CD19 FITC (#555412, BD), anti-CD14 FITC (#555397, BD), anti-CD16 FITC (#555406, BD), anti-CD56 FITC (#345811, BD), anti-CD11c APC (#559877, BD), anti-CD45 BV605 (#368523, Biolegend), anti-HLA-DR Pacific Blue (#307633, Biolegend), anti-CD1c APC-Cy7 (#331520, Biolegend), anti-CD141 BV711 (#563155, BD), anti-CD370 (CLEC9A) PE (#353804, Biolegend), anti-CD123 PE-Cy7 (#306010, Biolegend). Staining was performed with the Ab mixture and the Aqua Live/Dead Cell Stain Kit (Thermo Fisher Scientific) for 30 minutes at 4°C. Samples were analyzed on a Fortessa X20 (BD) and data were analyzed with FlowJo 10.8.1 software.

For the analysis of progenitors in blood, we used a published gating strategy.⁴¹ PBMC samples (stored in liquid nitrogen) were thawed and immediately stained with the following Abs: anti-CD3-FITC (#555332, BD), anti-CD19 FITC (#555412, BD), anti-CD14 FITC (#555397, BD), anti-CD16 FITC (#555406, BD), anti-CD56 FITC (#345811, BD), anti-CD34 APC (#343607, Biolegend), anti-CD45RA APC-H7 (#560674, BD), anti-CD123 Pacific Blue (#306044, Biolegend), anti-CD45 BV605 (#368523, Biolegend), anti-CD10 PE (#312204, Biolegend), and anti-CD38 PE-Vio770 (#130-113-428, Miltenyi Biotec).
were diluted in blocking buffer. For primary Abs not conjugated to HRP, membranes were washed in wash buffer (0.1% Tween 20 in PBS) and incubated with the following secondary antibodies: goat anti-mouse IgG (H + L)-HRP conjugate (1706516, Bio-Rad) or goat anti-rabbit IgG (H + L)-HRP conjugate (1706515, Bio-Rad). Staining was detected with the Clarity Western ECL substrate (Biorad, #1705061) or SuperSignal West Femto (Thermo Fisher Scientific, #34096) with ChemiDoc MP (Biorad). The images were analyzed with Image Lab 6.1 software (Biorad).

## Lentivirus transduction

HEK293T cells were used to seed a six-well plate at a density of $5 \times 10^5$ cells per well, in 2 mL per well DMEM supplemented with 10% FBS, two days before transduction. The cells were then transfected with pCMV-VSV-G (0.2 µg), pHXB2-env (0.2 µg; NIH-AIDS Reagent Program; #1069), psPAX2 (1 µg; a gift from D. Trono; Addgene plasmid #12260) and pTRIP-SFFV-deltaNGFR EV vector or pTRIP-SFFV-deltaNGFR containing the mFL or sFL coding sequences (1.6 µg) in Opti-MEM (Gibco; 300 µl) plus 9 µL X-tremeGENE 9 (Roche), according to the manufacturers' protocol. After 6 h of transfection, the medium was replaced with 3 mL DMEM supplemented with 10% FBS, and the cells were incubated for a further 24 h to produce lentiviral particles. On the day of HEK293T cell transfection, T-cell blasts from patients and controls were thawed, placed in TCM and reactivated with ImmunoCult™ Human CD3/CD28/CD2 T-Cell Activator (STEMCELL Technologies) in a 96-well plate, at a density of $2 \times 10^6$ cells/mL in a total volume of 100 µL/well. The next day (day 0), the supernatant of the HEK293T cells was collected and passed through a filter with 0.2 mm pores. Protamine sulfate (Sigma; $10 \mu g mL^{-1}$; 8 mg/mL) was added to the viral supernatant, and 100 µL of the supernatant was then added to the activated T cells, which were subjected to spinoculation for 2 hours at 1200 x g and room temperature. The spinoculated cells were then incubated for 48 hours at 37°C under an atmosphere containing 5% CO₂. After two days, the cells were harvested and transferred to small flasks containing TCM supplemented with 1% penicillin/streptomycin.

## Soluble FLT3L determination

The soluble form of FLT3L was determined in plasma or supernatant collected from cell lines expressing the protein (HEK293T cells and T-cell blasts) by sandwich ELISA (Human Flt-3 Ligand/FLT3L DuoSet ELISA, DY308, R&D Systems) in duplicate and in more than two independent experiments, in accordance with the manufacturer's protocol. The data obtained were subjected to statistical analysis by the regression analysis approach, in accordance with the instructions supplied with the kit.

## Analysis of FLT3L expression by flow cytometry

Transiently transfected HEK293T cells were harvested from the wells and washed once with PBS and cold FACS buffer (2% FBS in PBS). The cells were then stained simultaneously with a mixture of Live/Dead Fixable Aqua Dead Cell marker (reconstituted in DMSO; used at a 1:100 dilution in FACS buffer) and a primary Ab against FLT3L (recombinant anti-Flt3 ligand [EP1140Y] (ab52648), Abcam) or an isotype control (recombinant rabbit IgG, monoclonal [EPR25A] - (ab172730), Abcam) antibody for 60 min at 4°C in the dark. The cells were washed three times with FACS buffer and stained with a PE-conjugated goat anti-rabbit IgG (H+L) secondary antibody (A10542, Thermo Fisher Scientific) at a 1:500 dilution, for 30-60 min at 4°C in the dark. Cells were washed twice with FACS buffer and acquired with a Gallios (Beckman Coulter) machine. Data were then analyzed with FlowJo software v10. FLT3L expression on the cell surface of T-cell blasts (non-transduced and transduced) was analyzed with the same staining and analysis procedure.

## Deep immunophenotyping of whole blood

Deep immunophenotyping was performed by both conventional flow cytometry and mass cytometry (CyTOF) methods. For CyTOF, we used 200 µL fresh whole blood for the patients and controls. We used a custom-designed panel with antibodies indicated in STAR Methods, according to Fluidigm's instructions. Cells were frozen at -80°C after overnight dead-cell staining and acquisition was performed on a Helios mass cytometer (Fluidigm). All the samples were processed within 24 hours of sampling. Data analysis was performed with OMIQ software.

For classical immunophenotyping of the patients, fresh whole-blood samples were stained by incubation for 30 minutes at room temperature with antibodies against cell-surface markers, indicated in STAR Methods, and analyzed on a BD FACSLyric™ flow cytometer.

For a more detailed analysis of DC subsets, more than 10 M ($1 \times 10^7$) freshly isolated PBMCs were subjected to cell-surface staining with the following Abs: anti-CD3 FITC (#555332, BD), anti-CD19 FITC (#555412, BD), anti-CD14 FITC (#555397, BD), anti-CD16 FITC (#555406, BD), anti-CD56 FITC (#345811, BD), anti-CD11c APC (#559877, BD), anti-CD45 BV605 (#368523, Biolegend), anti-HLA-DR Pacific Blue (#307633, Biolegend), anti-CD1c APC-Cy7 (#331520, Biolegend), anti-CD141 BV711 (#563155, BD), anti-CD370 (CLEC9A) PE (#353804, Biolegend), anti-CD123 PE-Cy7 (#306010, Biolegend). Staining was performed with the Ab mixture and the Aqua Live/Dead Cell Stain Kit (Thermo Fisher Scientific) for 30 minutes at 4°C. Samples were analyzed on a Fortessa X20 (BD) and data were analyzed with FlowJo 10.8.1 software.

For the analysis of progenitors in blood, we used a published gating strategy.⁴¹ PBMC samples (stored in liquid nitrogen) were thawed and immediately stained with the following Abs: anti-CD3-FITC (#555332, BD), anti-CD19 FITC (#555412, BD), anti-CD14 FITC (#555397, BD), anti-CD16 FITC (#555406, BD), anti-CD56 FITC (#345811, BD), anti-CD34 APC (#343607, Biolegend), anti-CD45RA APC-H7 (#560674, BD), anti-CD123 Pacific Blue (#306044, Biolegend), anti-CD45 BV605 (#368523, Biolegend), anti-CD10 PE (#312204, Biolegend), and anti-CD38 PE-Vio770 (#130-113-428, Miltenyi Biotec).
Staining was performed with the Ab mixture and the Aqua Live/Dead Cell Stain Kit (Thermo Fisher Scientific) for 30 minutes at 4°C. Samples were analyzed on a Fortessa X20 (BD) machine and data were analyzed with FlowJo 10.8.1 software.

## Bone marrow experiments

Bone marrow mononuclear cells (BMNCs) were isolated by density separation on Lymphoprep (Axis-Shield, Oslo, Norway). CD34⁺ hematopoietic progenitor cells were isolated with an indirect CD34 microbead kit (CD34 MicroBead Kit UltraPure, human, 130-100-453 or CD34 MicroBead Kit, human, 130-046-702) and a magnetic separator (Miltenyi Biotec, Bergisch Gladbach, Germany). The purity of the CD34⁺ cells was checked with a MACSQuant analyzer (Miltenyi Biotec). For the *in vitro* differentiation assays, the ability of the CD34⁺ cells to produce CFUs and BFUs was evaluated in a clonal assay on methylcellulose (MethoCult H04535 and H04435, Stemcell Technologies, Vancouver, Canada) after 12 days of culture, according to the standard protocol. For HSPC phenotyping, the patients and healthy donor cells were characterized with a multi-labeled antibody panel, as listed below. HSPCs were analyzed on a spectral (Sony) flow cytometer, and the data were then analyzed with FlowJo software (v10.8.1). The following Ab panel was used for the HSPC analysis: anti-CD45RA BV711 (#304138, Biolegend), anti-CD90 PE-Cy5 (#328112, Biolegend), anti-CD38 BV605 (#303532, Biolegend), anti-CD34 APC-Cy7 #343514, Biolegend), anti-CD133 PE-Dazzle594 (#372812, Biolegend), anti-CD123 PE-Cy7 (#306010, Biolegend), anti-CD10 BV510 (#312220, Biolegend), anti-CD110 BV421 (#743955, BD) and a Lin custom panel including anti-CD2, anti-CD3, anti-CD4, anti-CD8, anti-CD13, anti-CD14, anti-CD15, anti-CD16, anti-CD19, anti-CD20, anti-CD56, anti-CD235a antibodies, all conjugated with PE and obtained from Miltenyi Biotec.

For the analysis of the B cell precursors by flow cytometry, 200000 isolated BMNCs were used from the patients and one HD for the staining with the following panel: anti-CD45 BV570 (#2120170, Sony), anti-CD20 BV510 (#302339, Biolegend), anti CD34 PE-Cy7 (#2317580, Sony), anti CD19 APC-Cy7 (#2111090, Sony), anti-CD10 APC (#332777, BD), anti-IgM BV421 (#562618, BD), anti-IgD BV711 (#740794, BD), anti-CD24-FITC (#555427, BD), anti-CD38 PE (#555460, BD), and 7AAD (#130-111-568, Miltenyi). Cells were analyzed on an Agilent NovoCyte Flow Cytometers, and the data were then analyzed with FlowJo software (v10.8.1).

For analysis of the leukocyte subsets of the BM, CyTOF was performed on 100 µL fresh whole BM from P2, P3, and the controls, as described above (with the same panel as for blood, described in STAR Methods).

## Bone biopsy evaluation

Immunostaining was performed on a whole-bone biopsy section in a paraffin block with a standardized Bond Max Leica machine (Leica Microsystems), a Polymer Refine detection kit (Leica), and the following antibodies: anti-glycophorin C (M082001, Agilent), anti-myeloperoxidase (A039829, Agilent) and anti-factor VIII (A008202, Agilent) antibodies.

## VirScan-phage immunoprecipitation-sequencing (PhIP-Seq)

Antibody profiling by phage immunoprecipitation-sequencing (PhIP-Seq)¹¹⁹ and the corresponding data analysis were performed as previously described.⁹⁸ Briefly, the experiment was performed on plasma samples from P2, P3, and 20 unrelated controls (representative of the adult general population). We calculated species-specific significance cutoff values to estimate the minimum number of enriched, non-homologous peptides required for a sample to be considered seropositive, as previously described with an in-house dataset and a generalized linear model.¹¹⁹ The adjusted virus scores were then used to generate heatmap plots. Pooled human plasma used for IVIg (Privigen CSL Behring AG), and human IgG-depleted serum (Molecular Innovations, Inc.) were used as additional controls.

## Luminex anti-HPV serological tests

Plasma samples from P2, P3, and 20 unrelated healthy controls were sent to the German Cancer Research Center (DKFZ, Heidelberg, Germany) on dry ice for serological analysis. Antibodies against the L1 antigens of HPV types 1, 2, 4, 5, 6, 8, 9, 10, 11, 12, 15, 16, 17, 18, 21, 22, 23, 24, 27b, 31, 33, 36, 38, 41, 45, 48, 50, 52, 58, 60, 75, 80, 88, 92, 93, 96, 101 and 103 were analyzed simultaneously in Luminex-based multiplex serological tests, as previously described.⁹⁸,¹²⁰ In brief, HPV L1 antigens were expressed as recombinant glutathione S-transferase (GST) fusion proteins, loaded onto polystyrene beads and simultaneously presented to primary serum antibodies. The immunocomplexes formed were detected with a biotinylated secondary antibody and streptavidin-R-phycoerythrin as a reporter dye. Serum samples were tested at a dilution of 1:100 and antigen-specific seropositivity was determined on the basis of predefined cutoff values. The values obtained were then used to generate heatmap plots.

## High-throughput sequencing (HTS) of the human TCR repertoire

DNA was extracted from whole blood from P2, P3, and four unrelated controls with the DNeasy blood & tissue kit (#69504; Qiagen). The rearranged genomic TRB loci were amplified by multiplex PCR on the DNA preparations obtained and sequenced by Adaptive Biotechnologies Seattle, WA. Adaptive Biotechnologies use assay-based and computational techniques to minimize PCR amplification bias. The assay is quantitative, and the frequency of a given TCR sequence is representative of the frequency of the clone type concerned in the original sample. The PCR products were sequenced on an Illumina HiSeq platform. Custom algorithms were used to filter the raw sequences for errors and to align the sequences with reference genome sequences. The data were then analyzed with ImmunoSeqTM online tools and custom R scripts.
Staining was performed with the Ab mixture and the Aqua Live/Dead Cell Stain Kit (Thermo Fisher Scientific) for 30 minutes at 4°C. Samples were analyzed on a Fortessa X20 (BD) machine and data were analyzed with FlowJo 10.8.1 software.

## Bone marrow experiments

Bone marrow mononuclear cells (BMNCs) were isolated by density separation on Lymphoprep (Axis-Shield, Oslo, Norway). CD34⁺ hematopoietic progenitor cells were isolated with an indirect CD34 microbead kit (CD34 MicroBead Kit UltraPure, human, 130-100-453 or CD34 MicroBead Kit, human, 130-046-702) and a magnetic separator (Miltenyi Biotec, Bergisch Gladbach, Germany). The purity of the CD34⁺ cells was checked with a MACSQuant analyzer (Miltenyi Biotec). For the *in vitro* differentiation assays, the ability of the CD34⁺ cells to produce CFUs and BFUs was evaluated in a clonal assay on methylcellulose (MethoCult H04535 and H04435, Stemcell Technologies, Vancouver, Canada) after 12 days of culture, according to the standard protocol. For HSPC phenotyping, the patients and healthy donor cells were characterized with a multi-labeled antibody panel, as listed below. HSPCs were analyzed on a spectral (Sony) flow cytometer, and the data were then analyzed with FlowJo software (v10.8.1). The following Ab panel was used for the HSPC analysis: anti-CD45RA BV711 (#304138, Biolegend), anti-CD90 PE-Cy5 (#328112, Biolegend), anti-CD38 BV605 (#303532, Biolegend), anti-CD34 APC-Cy7 #343514, Biolegend), anti-CD133 PE-Dazzle594 (#372812, Biolegend), anti-CD123 PE-Cy7 (#306010, Biolegend), anti-CD10 BV510 (#312220, Biolegend), anti-CD110 BV421 (#743955, BD) and a Lin custom panel including anti-CD2, anti-CD3, anti-CD4, anti-CD8, anti-CD13, anti-CD14, anti-CD15, anti-CD16, anti-CD19, anti-CD20, anti-CD56, anti-CD235a antibodies, all conjugated with PE and obtained from Miltenyi Biotec.

For the analysis of the B cell precursors by flow cytometry, 200000 isolated BMNCs were used from the patients and one HD for the staining with the following panel: anti-CD45 BV570 (#2120170, Sony), anti-CD20 BV510 (#302339, Biolegend), anti CD34 PE-Cy7 (#2317580, Sony), anti CD19 APC-Cy7 (#2111090, Sony), anti-CD10 APC (#332777, BD), anti-IgM BV421 (#562618, BD), anti-IgD BV711 (#740794, BD), anti-CD24-FITC (#555427, BD), anti-CD38 PE (#555460, BD), and 7AAD (#130-111-568, Miltenyi). Cells were analyzed on an Agilent NovoCyte Flow Cytometers, and the data were then analyzed with FlowJo software (v10.8.1).

For analysis of the leukocyte subsets of the BM, CyTOF was performed on 100 µL fresh whole BM from P2, P3, and the controls, as described above (with the same panel as for blood, described in STAR Methods).

## Bone biopsy evaluation

Immunostaining was performed on a whole-bone biopsy section in a paraffin block with a standardized Bond Max Leica machine (Leica Microsystems), a Polymer Refine detection kit (Leica), and the following antibodies: anti-glycophorin C (M082001, Agilent), anti-myeloperoxidase (A039829, Agilent) and anti-factor VIII (A008202, Agilent) antibodies.

## VirScan-phage immunoprecipitation-sequencing (PhIP-Seq)

Antibody profiling by phage immunoprecipitation-sequencing (PhIP-Seq)¹¹⁹ and the corresponding data analysis were performed as previously described.⁹⁸ Briefly, the experiment was performed on plasma samples from P2, P3, and 20 unrelated controls (representative of the adult general population). We calculated species-specific significance cutoff values to estimate the minimum number of enriched, non-homologous peptides required for a sample to be considered seropositive, as previously described with an in-house dataset and a generalized linear model.¹¹⁹ The adjusted virus scores were then used to generate heatmap plots. Pooled human plasma used for IVIg (Privigen CSL Behring AG), and human IgG-depleted serum (Molecular Innovations, Inc.) were used as additional controls.

## Luminex anti-HPV serological tests

Plasma samples from P2, P3, and 20 unrelated healthy controls were sent to the German Cancer Research Center (DKFZ, Heidelberg, Germany) on dry ice for serological analysis. Antibodies against the L1 antigens of HPV types 1, 2, 4, 5, 6, 8, 9, 10, 11, 12, 15, 16, 17, 18, 21, 22, 23, 24, 27b, 31, 33, 36, 38, 41, 45, 48, 50, 52, 58, 60, 75, 80, 88, 92, 93, 96, 101 and 103 were analyzed simultaneously in Luminex-based multiplex serological tests, as previously described.⁹⁸,¹²⁰ In brief, HPV L1 antigens were expressed as recombinant glutathione S-transferase (GST) fusion proteins, loaded onto polystyrene beads and simultaneously presented to primary serum antibodies. The immunocomplexes formed were detected with a biotinylated secondary antibody and streptavidin-R-phycoerythrin as a reporter dye. Serum samples were tested at a dilution of 1:100 and antigen-specific seropositivity was determined on the basis of predefined cutoff values. The values obtained were then used to generate heatmap plots.

## High-throughput sequencing (HTS) of the human TCR repertoire

DNA was extracted from whole blood from P2, P3, and four unrelated controls with the DNeasy blood & tissue kit (#69504; Qiagen). The rearranged genomic TRB loci were amplified by multiplex PCR on the DNA preparations obtained and sequenced by Adaptive Biotechnologies Seattle, WA. Adaptive Biotechnologies use assay-based and computational techniques to minimize PCR amplification bias. The assay is quantitative, and the frequency of a given TCR sequence is representative of the frequency of the clone type concerned in the original sample. The PCR products were sequenced on an Illumina HiSeq platform. Custom algorithms were used to filter the raw sequences for errors and to align the sequences with reference genome sequences. The data were then analyzed with ImmunoSeqTM online tools and custom R scripts.
## High-throughput sequencing (HTS) of the human BCR repertoire

We assessed B-cell receptor (BCR) diversity in these patients with low B-cell counts by performing NGS of the immunoglobulin heavy chain (IGH) repertoire in sorted circulating B cells. CD19⁺ B cells from P2, P3, and unrelated age-matched healthy control PBMCs were sorted magnetically with anti-CD19 Ab-conjugated microbeads (CD19 MicroBeads, human, 130-050-301, Miltenyi Biotec, Ber- gisch Gladbach, Germany) according to the manufacturer's protocol. Isolated B cells were washed with 1X PBS, and DNA was ex- tracted with the DNeasy blood & tissue kit (#69504; Qiagen). The rearranged genomic IGH loci were amplified by multiplex PCR on the prepared DNA and sequenced by Adaptive Biotechnologies (Seattle, WA). Adaptive Biotechnologies uses assay-based and compu- tational techniques to minimize PCR amplification bias. The assay was quantitative, and the frequency of a given IGH sequence is representative of the frequency of the clone type concerned in the original sample. The PCR products were sequenced on an Illumina HiSeq platform. Custom algorithms were used to filter the raw sequences for errors and to align the sequences with reference genome sequences. The data were then analyzed with ImmunoSeqTM online tools, and finally, all analyses were conducted in R v.4 (http://www.R-project.org/) using several custom R scripts.

## Skin digestion and skin cell sorting

Skin punch biopsies (4 mm) were performed on healthy skin and warts from the patients (P2 and P3) and on healthy skin from three healthy donors. The biopsy specimens were collected in complete RPMI medium (10% FBS and 1% penicillin streptomycin). They were washed with 1X PBS to eliminate the contaminating blood surrounding the tissue and were then incubated in 500 µL 1X dispase (#07923, STEMCELL Technologies) medium in a 24-well plate for 2 h at 37°C, with shaking every 30 minutes. Segments of epidermis were then separated from the dermis. The dermis was cut into chunks with an area of about 1 mm², returned to the same dispase medium and digested overnight (12 h at 37°C) with 500 µL 2X collagenase (#1088866001, Roche). The epidermis was stored in travel medium at 4°C overnight. It was then washed in 1X PBS to eliminate the serum and transferred to a new 24-well plate for digestion for 45 min at 37°C with 500 µL trypsin-EDTA. The epidermal digestion reaction was stopped by adding FBS and the dermal digestion reaction was stopped by adding 6 mM EDTA in PBS (6% FBS). The cells in the digested samples were dissociated by pipetting up and down and the suspension was filtered through a 35 µm-mesh filter. Samples were centrifuged (350 x g, 10 min, 4°C) and imme- diately stained in PBS (supplemented with 0.04% BSA) with anti-CD45 PE (#304008, Biolegend) and anti-HLA-ABC APC (#311409, Biolegend) antibodies and Aqua Live-Dead stain, on ice for 30 min. The cells were then sorted, in microtubes, with a BD FACS Aria II sorter (BD Biosciences), into live CD45⁺ and CD45⁻ fractions. Cells from each fraction were then pooled in a specific ratio (20,000 epidermal CD45⁻ cells, all sorted epidermal CD45⁺ cells, 10,000 dermal CD45⁻ cells, and all sorted dermal CD45⁺ cells) and trans- ferred immediately to the 10X Genomics single-cell RNA sequencing platform.

## Single-cell RNA-sequencing

Single-cell RNA sequencing (scRNA-seq) was performed with different strategies, as explained below. First, to study the bone marrow, we enriched the fresh bone marrow aspirate of two healthy individuals and P2 in CD34⁺ cells as described above in the bone marrow section. Second, for studies of blood leukocyte subsets in the basal state, we used freshly isolated PBMCs from P2, P3, and two healthy donors for library preparation. Third, for enrichment in myeloid cells, we sorted the HLA-DR⁺ monocytes and DCs from the fresh PBMCs of P2, P3, and the two healthy controls (same controls as for the previous experiment on the blood). Cells were stained with a mixture of Abs against cell surface markers and Aqua Live/Dead Cell Stain (same staining as above), and were then sorted in microtubes with a BD FACS Aria II sorter (BD Biosciences). Cells were sorted by gating on HLA-DR⁺Lin⁻ (CD3, CD19, CD20, and CD56) cells. The freshly sorted cells were then immediately subjected to RNA sequencing. The Abs used for cell sorting were anti CD3-FITC (#555332, BD), anti-CD19 FITC (#555412, BD), anti-CD20 FITC (#560962, BD), anti-CD56 FITC (#345811, BD), anti-CD11c APC (#559877, BD), anti-HLA-DR PE (#555812, BD), anti-CD1c APC-Cy7 (#331520, Biolegend), anti-CD14 BV711 (#563155, BD), anti-CD123 PE-Cy7 (#306010, Biolegend), anti-CD14 Pacific Blue (#558121, BD), and anti-CD16 BV650 (#563692, BD) antibodies.

Fourth, we used fresh cells obtained from digested biopsy specimens from the healthy skin and lesions of P2 and P3 (as described in the skin digestion section) for scRNA-seq. Library preparation and sequencing were performed in the same manner for all ap- proaches. Briefly, the scRNA-seq libraries were generated with either the Chromium Next GEM Single-Cell 3' Reagent Kit v3.1 or the Chromium Next GEM Automated Single-Cell 5' Kit v2 with Feature Barcoding (10X Genomics) according to the manufacturer's protocol. The purified libraries were sequenced on a NovaSeq 6000 (Illumina) machine. We aimed to achieve a median depth of 50000 reads per cell for gene expression.

## Single-cell RNA-sequencing analysis

The data generated during this study were analyzed in an integrative approach. For the PBMC analysis, we added the data previously obtained for four other controls available in the in-house pipeline of the laboratory (data for four adult controls from different batches of experiments added to the data for the two healthy controls in the experiment), and publicly available control PBMC datasets downloaded from the 10X Genomics web portal (https://support.10xgenomics.com/single-cell-gene-expression/datasets). For the skin analysis, patients were compared with data for four healthy in-house controls acquired at different time points.

Cells were subjected to preprocessing with CellRanger. Approximately 10,000 cells were sequenced per sample. Data were filtered manually with common quality-control metrics and integrated with Harmony.¹⁰⁰ We performed two sequential graph-based
## High-throughput sequencing (HTS) of the human BCR repertoire

We assessed B-cell receptor (BCR) diversity in these patients with low B-cell counts by performing NGS of the immunoglobulin heavy chain (IGH) repertoire in sorted circulating B cells. CD19⁺ B cells from P2, P3, and unrelated age-matched healthy control PBMCs were sorted magnetically with anti-CD19 Ab-conjugated microbeads (CD19 MicroBeads, human, 130-050-301, Miltenyi Biotec, Ber- gisch Gladbach, Germany) according to the manufacturer's protocol. Isolated B cells were washed with 1X PBS, and DNA was ex- tracted with the DNeasy blood & tissue kit (#69504; Qiagen). The rearranged genomic IGH loci were amplified by multiplex PCR on the prepared DNA and sequenced by Adaptive Biotechnologies (Seattle, WA). Adaptive Biotechnologies uses assay-based and compu- tational techniques to minimize PCR amplification bias. The assay was quantitative, and the frequency of a given IGH sequence is representative of the frequency of the clone type concerned in the original sample. The PCR products were sequenced on an Illumina HiSeq platform. Custom algorithms were used to filter the raw sequences for errors and to align the sequences with reference genome sequences. The data were then analyzed with ImmunoSeqTM online tools, and finally, all analyses were conducted in R v.4 (http://www.R-project.org/) using several custom R scripts.

## Skin digestion and skin cell sorting

Skin punch biopsies (4 mm) were performed on healthy skin and warts from the patients (P2 and P3) and on healthy skin from three healthy donors. The biopsy specimens were collected in complete RPMI medium (10% FBS and 1% penicillin streptomycin). They were washed with 1X PBS to eliminate the contaminating blood surrounding the tissue and were then incubated in 500 µL 1X dispase (#07923, STEMCELL Technologies) medium in a 24-well plate for 2 h at 37°C, with shaking every 30 minutes. Segments of epidermis were then separated from the dermis. The dermis was cut into chunks with an area of about 1 mm², returned to the same dispase medium and digested overnight (12 h at 37°C) with 500 µL 2X collagenase (#1088866001, Roche). The epidermis was stored in travel medium at 4°C overnight. It was then washed in 1X PBS to eliminate the serum and transferred to a new 24-well plate for digestion for 45 min at 37°C with 500 µL trypsin-EDTA. The epidermal digestion reaction was stopped by adding FBS and the dermal digestion reaction was stopped by adding 6 mM EDTA in PBS (6% FBS). The cells in the digested samples were dissociated by pipetting up and down and the suspension was filtered through a 35 µm-mesh filter. Samples were centrifuged (350 x g, 10 min, 4°C) and imme- diately stained in PBS (supplemented with 0.04% BSA) with anti-CD45 PE (#304008, Biolegend) and anti-HLA-ABC APC (#311409, Biolegend) antibodies and Aqua Live-Dead stain, on ice for 30 min. The cells were then sorted, in microtubes, with a BD FACS Aria II sorter (BD Biosciences), into live CD45⁺ and CD45⁻ fractions. Cells from each fraction were then pooled in a specific ratio (20,000 epidermal CD45⁻ cells, all sorted epidermal CD45⁺ cells, 10,000 dermal CD45⁻ cells, and all sorted dermal CD45⁺ cells) and trans- ferred immediately to the 10X Genomics single-cell RNA sequencing platform.

## Single-cell RNA-sequencing

Single-cell RNA sequencing (scRNA-seq) was performed with different strategies, as explained below. First, to study the bone marrow, we enriched the fresh bone marrow aspirate of two healthy individuals and P2 in CD34⁺ cells as described above in the bone marrow section. Second, for studies of blood leukocyte subsets in the basal state, we used freshly isolated PBMCs from P2, P3, and two healthy donors for library preparation. Third, for enrichment in myeloid cells, we sorted the HLA-DR⁺ monocytes and DCs from the fresh PBMCs of P2, P3, and the two healthy controls (same controls as for the previous experiment on the blood). Cells were stained with a mixture of Abs against cell surface markers and Aqua Live/Dead Cell Stain (same staining as above), and were then sorted in microtubes with a BD FACS Aria II sorter (BD Biosciences). Cells were sorted by gating on HLA-DR⁺Lin⁻ (CD3, CD19, CD20, and CD56) cells. The freshly sorted cells were then immediately subjected to RNA sequencing. The Abs used for cell sorting were anti CD3-FITC (#555332, BD), anti-CD19 FITC (#555412, BD), anti-CD20 FITC (#560962, BD), anti-CD56 FITC (#345811, BD), anti-CD11c APC (#559877, BD), anti-HLA-DR PE (#555812, BD), anti-CD1c APC-Cy7 (#331520, Biolegend), anti-CD14 BV711 (#563155, BD), anti-CD123 PE-Cy7 (#306010, Biolegend), anti-CD14 Pacific Blue (#558121, BD), and anti-CD16 BV650 (#563692, BD) antibodies.

Fourth, we used fresh cells obtained from digested biopsy specimens from the healthy skin and lesions of P2 and P3 (as described in the skin digestion section) for scRNA-seq. Library preparation and sequencing were performed in the same manner for all ap- proaches. Briefly, the scRNA-seq libraries were generated with either the Chromium Next GEM Single-Cell 3' Reagent Kit v3.1 or the Chromium Next GEM Automated Single-Cell 5' Kit v2 with Feature Barcoding (10X Genomics) according to the manufacturer's protocol. The purified libraries were sequenced on a NovaSeq 6000 (Illumina) machine. We aimed to achieve a median depth of 50000 reads per cell for gene expression.

## Single-cell RNA-sequencing analysis

The data generated during this study were analyzed in an integrative approach. For the PBMC analysis, we added the data previously obtained for four other controls available in the in-house pipeline of the laboratory (data for four adult controls from different batches of experiments added to the data for the two healthy controls in the experiment), and publicly available control PBMC datasets downloaded from the 10X Genomics web portal (https://support.10xgenomics.com/single-cell-gene-expression/datasets). For the skin analysis, patients were compared with data for four healthy in-house controls acquired at different time points.

Cells were subjected to preprocessing with CellRanger. Approximately 10,000 cells were sequenced per sample. Data were filtered manually with common quality-control metrics and integrated with Harmony.¹⁰⁰ We performed two sequential graph-based
clustering analyses. The first-round clustering identified general cell subsets, whereas the second-round clustering identified T-lymphocyte cell subsets at a sufficiently high resolution for analysis. For the first round of clustering, we identified clusters with the SingleR pipeline guided by MonacolimmuneData¹¹⁵ for PBMCs and BlueprintEncodeData¹²¹,¹²² for skin cells. In the second round of clustering, we used MonacolimmuneData.¹¹⁵ Cell type-specific marker gene expression was then assessed manually. Clusters were visualized by uniform manifold approximation and projection (UMAP). Gene expression in single cells was quantified with Seurat. Pseudobulk analysis¹²³ was performed by aggregating all reads from cells assigned to a given cluster, as previously described.¹⁰⁶ Differential expression analysis was performed with DESeq2.¹⁰⁷ P-values were obtained with DESeq2 and the false-discovery rate (FDR) was calculated by the Benjamini-Hochberg procedure; FDR < 0.05 was used as the cutoff. All analyses were conducted in R v.4 (http://www.R-project.org/).

For the BM CD34⁺ HSPCs, data were processed and scRNAseq data were analyzed according to a published pipeline.¹²⁴ We filtered on cells expressing CD34 to remove contaminants, due to the low purity of the CD34⁺ cells selected by the magnetic approach for P2 patient. We analyzed a total of 15,164 CD34⁺ cells, with a mean of 14,451 genes detected (SD=143.1). The single-cell RNAseq data are available from Biostudies EMBL-EBI (S-BSST1333). Each individual cell in our dataset was annotated by the Cell-ID method and with reference BM HSPC signatures (*p*<0.01).⁴²,¹²⁴

## Whole-blood activation assay

Whole-blood samples from P2, P3 and three healthy donors were collected into heparin-containing collection tubes. Samples were diluted 1:2 in RPMI 1640 supplemented with 100 IU/mL penicillin and 100 µg/mL streptomycin (Thermo Fisher Scientific). Stimulation was performed in a 48-well plate. Briefly, 1 mL of blood was used per well and per treatment. Samples were incubated with medium alone, with live BCG (*M. bovis*-BCG, Pasteur substrain) at a MOI of 20, or with BCG plus IL-12 (20 ng/mL; #219-IL, R&D Systems), BCG plus IL-23 (R&D Systems), or PMA (Merck) for 48 hours at 37°C under an atmosphere containing 5% CO₂. The supernatants were collected for cytokine determinations. The IFN-γ and TNF contents of the supernatants from the whole-blood stimulation assay were determined in a LEGENDplex™ assay (BioLegend) according to the manufacturer's instructions. Samples were analyzed by flow cytometry on a Gallios flow cytometer. Data were analyzed with LEGENDplex Cloud-based Data Analysis Software (BioLegend).

## BCG stimulation assay

PBMC samples from P2, P3, healthy donors, and a GATA2-haploinsufficient patient were dispensed into a U-bottomed 96-well plate at a density of 2 × 10⁵ cells per well, in 200 µL lymphocyte medium per well. Cells were incubated in the presence or absence of live BCG, at a multiplicity of infection of 1, with or without recombinant human IL-12 (500 pg/mL, R&D) or recombinant human IL-23 (100 ng/mL, 1290-IL R&D Systems). After 40 hours of stimulation, GolgiPlug (BD Biosciences, 555029; 1:1,000 dilution) was added to each well to inhibit cytokine secretion. After another eight hours of incubation, the cells were collected by centrifugation for flow cytometry analysis. Cells were stained with the Zombie NIR Fixable Viability Kit (BioLegend; 1:2,000 dilution) at room temperature for 15 minutes, and then stained on ice for 30 minutes with a surface-staining panel containing FcR blocking reagent (Miltenyi Biotec; 1:50 dilution), anti-CD3-Alexa Fluor 532 (eBioscience, Clone: UCHT1, 58-0038-42; 1:50 dilution), anti-γδTCR-FITC (eBioscience, Clone:B1.1, 11-9959-41; 1:50 dilution), anti-Vδ2-APC/Fire 750 (BioLegend, Clone:B6, 331419; 1:100 dilution), anti-CD56-BV605 (BioLegend, Clone: 5.1H11, 362537; 1:100 dilution), anti-CD4-BV750 (BD Biosciences, Clone: SK3, 566356; 1:800 dilution), anti-CD8a-Pacific Blue (BioLegend, Clone: SK1, 344717; 1:100 dilution), anti-Vα7.2 TCR-APC (BioLegend, Clone: 3C10, 351708; 1:100 dilution), anti-Vα24-Jα18-PE/Cy7 (BioLegend, Clone: 6B11, 342912; 1:100 dilution), anti-CD20-BV785 (BioLegend, Clone: 2H7, 302356; 1:200 dilution) and anti-PD-1-PE (eBioscience, Clone: MIH4, 12-9969-42; 1:100 dilution) antibodies. Cells were fixed by incubation with 2% paraformaldehyde in PBS on ice for 15 minutes. Cells were then permeabilized/stained by incubation overnight at -20°C in the permeabilization buffer from the Nuclear Transcription Factor Buffer Set (BioLegend), with an intracellular cytokine panel containing FcR blocking reagent (Miltenyi Biotec; 1:50 dilution), anti-IFN-γ-BV711 (BioLegend, Clone: 4 S.B3, 502540; 1:50 dilution), anti-TNF-BV510 (BioLegend, Clone: MAb11, 502950; 1:50 dilution) and anti-IL-10-PE/Dazzle594 (BioLegend, Clone: JES3-19F1, 506812; 1:50 dilution) antibodies. As a positive control, cells in a separate well were stimulated by incubation with PMA (Sigma; 25 ng mL⁻¹) and ionomycin (Sigma; 500 nM) for one hour without GolgiPlug followed by a further 7 hours with GolgiPlug (for intracellular cytokine staining). Cells were acquired with an Aurora cytometer (Cytek). Data were analyzed with FlowJo.

## T-cell proliferation assay

This experiment was performed as for the purposes of diagnosis, in the diagnostic unit of Necker Hospital, Paris (AP-HP).

## In-vitro and ex-vivo CD3⁺TCRαβ⁺ CD4⁺ T-cell polarization experiment

Cryopreserved PBMCs were labelled with antibodies directed against CD45RA (BV605, clone HI100, #562886, BD Horizon), and CCR7 (AF700, clone 150503, #561143, BD Pharmingen), CD3 (BV421, clone UCHT1, #562426, BD Horizon), CD8 (BUV395, clone RPA-T8, #563795, BD Horizon), CD4 (APC, clone RPAT4, #300514, BioLegend), TCRαβ (PECy7, clone IP26, #306720, BioLegend), and TCRγδ (PE, clone B1.1, #12-9959-42, ebioscience). Naïve (defined as CD45RA⁺CCR7⁺) and memory (defined as CD45RA⁻CCR7⁺⁻) CD3⁺TCRαβ⁺CD4⁺ T cells were isolated (>98% purity) with a FACS Aria cell sorter (BD Biosciences). Isolated cells were then cultured with T-cell activation and expansion beads (TAE, anti-CD2/CD3/CD28, Miltenyi Biotec) + IL-2 (50 IU/mL, #IL002, Merck) alone (TH0) or under TH1 (IL-12 [50 ng/mL, # 219-IL-005, R&D Systems]), or TH17 (TGF-β1 [2.5 ng/mL, #100-21C-10,
clustering analyses. The first-round clustering identified general cell subsets, whereas the second-round clustering identified T-lymphocyte cell subsets at a sufficiently high resolution for analysis. For the first round of clustering, we identified clusters with the SingleR pipeline guided by MonacolimmuneData¹¹⁵ for PBMCs and BlueprintEncodeData¹²¹,¹²² for skin cells. In the second round of clustering, we used MonacolimmuneData.¹¹⁵ Cell type-specific marker gene expression was then assessed manually. Clusters were visualized by uniform manifold approximation and projection (UMAP). Gene expression in single cells was quantified with Seurat. Pseudobulk analysis¹²³ was performed by aggregating all reads from cells assigned to a given cluster, as previously described.¹⁰⁶ Differential expression analysis was performed with DESeq2.¹⁰⁷ P-values were obtained with DESeq2 and the false-discovery rate (FDR) was calculated by the Benjamini-Hochberg procedure; FDR < 0.05 was used as the cutoff. All analyses were conducted in R v.4 (http://www.R-project.org/).

For the BM CD34⁺ HSPCs, data were processed and scRNAseq data were analyzed according to a published pipeline.¹²⁴ We filtered on cells expressing CD34 to remove contaminants, due to the low purity of the CD34⁺ cells selected by the magnetic approach for P2 patient. We analyzed a total of 15,164 CD34⁺ cells, with a mean of 14,451 genes detected (SD=143.1). The single-cell RNAseq data are available from Biostudies EMBL-EBI (S-BSST1333). Each individual cell in our dataset was annotated by the Cell-ID method and with reference BM HSPC signatures (*p*<0.01).⁴²,¹²⁴

## Whole-blood activation assay

Whole-blood samples from P2, P3 and three healthy donors were collected into heparin-containing collection tubes. Samples were diluted 1:2 in RPMI 1640 supplemented with 100 IU/mL penicillin and 100 µg/mL streptomycin (Thermo Fisher Scientific). Stimulation was performed in a 48-well plate. Briefly, 1 mL of blood was used per well and per treatment. Samples were incubated with medium alone, with live BCG (*M. bovis*-BCG, Pasteur substrain) at a MOI of 20, or with BCG plus IL-12 (20 ng/mL; #219-IL, R&D Systems), BCG plus IL-23 (R&D Systems), or PMA (Merck) for 48 hours at 37°C under an atmosphere containing 5% CO₂. The supernatants were collected for cytokine determinations. The IFN-γ and TNF contents of the supernatants from the whole-blood stimulation assay were determined in a LEGENDplex™ assay (BioLegend) according to the manufacturer's instructions. Samples were analyzed by flow cytometry on a Gallios flow cytometer. Data were analyzed with LEGENDplex Cloud-based Data Analysis Software (BioLegend).

## BCG stimulation assay

PBMC samples from P2, P3, healthy donors, and a GATA2-haploinsufficient patient were dispensed into a U-bottomed 96-well plate at a density of 2 × 10⁵ cells per well, in 200 µL lymphocyte medium per well. Cells were incubated in the presence or absence of live BCG, at a multiplicity of infection of 1, with or without recombinant human IL-12 (500 pg/mL, R&D) or recombinant human IL-23 (100 ng/mL, 1290-IL R&D Systems). After 40 hours of stimulation, GolgiPlug (BD Biosciences, 555029; 1:1,000 dilution) was added to each well to inhibit cytokine secretion. After another eight hours of incubation, the cells were collected by centrifugation for flow cytometry analysis. Cells were stained with the Zombie NIR Fixable Viability Kit (BioLegend; 1:2,000 dilution) at room temperature for 15 minutes, and then stained on ice for 30 minutes with a surface-staining panel containing FcR blocking reagent (Miltenyi Biotec; 1:50 dilution), anti-CD3-Alexa Fluor 532 (eBioscience, Clone: UCHT1, 58-0038-42; 1:50 dilution), anti-γδTCR-FITC (eBioscience, Clone:B1.1, 11-9959-41; 1:50 dilution), anti-Vδ2-APC/Fire 750 (BioLegend, Clone:B6, 331419; 1:100 dilution), anti-CD56-BV605 (BioLegend, Clone: 5.1H11, 362537; 1:100 dilution), anti-CD4-BV750 (BD Biosciences, Clone: SK3, 566356; 1:800 dilution), anti-CD8a-Pacific Blue (BioLegend, Clone: SK1, 344717; 1:100 dilution), anti-Vα7.2 TCR-APC (BioLegend, Clone: 3C10, 351708; 1:100 dilution), anti-Vα24-Jα18-PE/Cy7 (BioLegend, Clone: 6B11, 342912; 1:100 dilution), anti-CD20-BV785 (BioLegend, Clone: 2H7, 302356; 1:200 dilution) and anti-PD-1-PE (eBioscience, Clone: MIH4, 12-9969-42; 1:100 dilution) antibodies. Cells were fixed by incubation with 2% paraformaldehyde in PBS on ice for 15 minutes. Cells were then permeabilized/stained by incubation overnight at -20°C in the permeabilization buffer from the Nuclear Transcription Factor Buffer Set (BioLegend), with an intracellular cytokine panel containing FcR blocking reagent (Miltenyi Biotec; 1:50 dilution), anti-IFN-γ-BV711 (BioLegend, Clone: 4 S.B3, 502540; 1:50 dilution), anti-TNF-BV510 (BioLegend, Clone: MAb11, 502950; 1:50 dilution) and anti-IL-10-PE/Dazzle594 (BioLegend, Clone: JES3-19F1, 506812; 1:50 dilution) antibodies. As a positive control, cells in a separate well were stimulated by incubation with PMA (Sigma; 25 ng mL⁻¹) and ionomycin (Sigma; 500 nM) for one hour without GolgiPlug followed by a further 7 hours with GolgiPlug (for intracellular cytokine staining). Cells were acquired with an Aurora cytometer (Cytek). Data were analyzed with FlowJo.

## T-cell proliferation assay

This experiment was performed as for the purposes of diagnosis, in the diagnostic unit of Necker Hospital, Paris (AP-HP).

## In-vitro and ex-vivo CD3⁺TCRαβ⁺ CD4⁺ T-cell polarization experiment

Cryopreserved PBMCs were labelled with antibodies directed against CD45RA (BV605, clone HI100, #562886, BD Horizon), and CCR7 (AF700, clone 150503, #561143, BD Pharmingen), CD3 (BV421, clone UCHT1, #562426, BD Horizon), CD8 (BUV395, clone RPA-T8, #563795, BD Horizon), CD4 (APC, clone RPAT4, #300514, BioLegend), TCRαβ (PECy7, clone IP26, #306720, BioLegend), and TCRγδ (PE, clone B1.1, #12-9959-42, ebioscience). Naïve (defined as CD45RA⁺CCR7⁺) and memory (defined as CD45RA⁻CCR7⁺⁻) CD3⁺TCRαβ⁺CD4⁺ T cells were isolated (>98% purity) with a FACS Aria cell sorter (BD Biosciences). Isolated cells were then cultured with T-cell activation and expansion beads (TAE, anti-CD2/CD3/CD28, Miltenyi Biotec) + IL-2 (50 IU/mL, #IL002, Merck) alone (TH0) or under TH1 (IL-12 [50 ng/mL, # 219-IL-005, R&D Systems]), or TH17 (TGF-β1 [2.5 ng/mL, #100-21C-10,
Peprotech], IL-1β [50 ng/mL, #200-01B-10, Peprotech], IL-6 [50 ng/mL, #200-06-20, PeproTech], IL-21 [50 ng/mL, #200-21, Peprotech], IL-23 [50 ng/mL, #200-23-10, PeproTech]) polarizing conditions. After five days of polarization, the cells were stimulated with PMA (100 ng/mL, #P8139-1MG, Sigma Aldrich)-ionomycin (750 ng/mL, #I0634-1MG, Sigma Aldrich) for 6 h, with brefeldin A (10 mg/mL, #B7651-5MG, Sigma Aldrich) added after the first 2 h of incubation. For the assessment of intracellular cytokine production, cells were incubated with conjugated monoclonal antibodies directed against IFN-γ (BUV737, clone 4S.B3, #564620, BD Horizon), TNF (PerCP, clone Mab11, #502924, BioLegend), IL-9 (PE, clone MH9A3, #560807, BD Pharmingen), IL-13 (BV421, clone JES10-5A2, #563580, BD Horizon), IL-4 (AF488, clone 8D4-8, #500710, BioLegend), IL-17A (BV510, clone BL168, #512330, BioLegend), IL-17F (BV650, clone O33-782, #564264, BD Horizon), IL-2 (BV750, clone MQ1-17H12, #566361, BD Horizon), IL-21 (eF660, clone eBio3A3-N2, #50-7219-42, Thermo Fisher Scientific), and IL-22 (PerCP e710, clone 22URTI, #46-7229-42, eBioscience). All cells were also stained with the Zombie UV fixable viability dye kit (#423107, BioLegend). Cells and beads were acquired on a BD FACSymphony A5 Cell Analyzer (BD bioscience) and analyzed with FlowJo Software. A previously described gating strategy was used (109).

## Skin tissue processing and imaging mass cytometry (IMC) data acquisition

We used 3 µm-thick sections of formalin-fixed paraffin-embedded blocks of tissue from cutaneous biopsies performed on patients (warts and normal skin), healthy relatives (normal skin) and healthy donors (normal skin). Slides were first baked for 2 h in a slide oven until all the visible wax had disappeared. They were then dewaxed in a meta-xylene bath for 20 min and rehydrated in a series of descending concentrations of ethanol (100%, 70% and 50%; 10 min each). Slides were washed for five minutes in water and then incubated in antigen retrieval solution for 30 min at 96°C followed by 10 min at room temperature. They were then washed for 10 min in pure water and 10 min in PBS. A PAP pen was used to encircle the samples on each slide, and the slides were then blocked by incubation with 100 µL superblock solution per sample for 45 min in a hydration chamber. Staining was achieved in two steps: a mixture of antibodies diluted in PBS-BSA 0.5% was first applied to the slides, which were then incubated overnight at 4°C in a hydration chamber. The slides were then washed with PBS for 10 minutes and incubated with a second mixture at room temperature for 1.5 h. Our 29-metal antibody IMC panel was designed to evaluate the immune infiltrate present in healthy skin and at the site of infection. It made it possible to study several cell subsets simultaneously, including APCs (DCs, macrophages, monocytes, Langerhans cells), lymphocytes (CD4, CD8, Trm, γδ, Treg, and B cells), neutrophils and keratinocytes (Abs summarized in STAR Methods). Each antibody used in our mixture was previously validated on tonsils, by IMC for conjugated antibodies purchased directly from Fluidigm and by both immunofluorescence and IMC for in-house conjugated antibodies. All conjugations were performed according to the Fluidigm protocol. The stained slides were washed for 16 min in 0.2% Triton-X100 and 16 min in PBS. They were then stained with Intercalator-Ir in Maxpar PBS 1:400 for 30 min at room temperature in a hydration chamber, washed in pure water for 4 min and air-dried for at least 20 min. Finally, IMC data corresponding to pre-established regions of interest (ROIs) were acquired as raw .mcd files with the Hyperion imaging system (Fluidigm).

## IMC data analysis

IMC data preprocessing and segmentations were performed with the Steinbock toolkit.¹²⁵ The preprocessing steps led to the generation of a single .tif file per ROI. Segmentation masks were generated with the deep learning model Deepcell using the iridium channel as the nuclear signal and vimentin, pancytokeratin, CD45, E-cadherin and HLA-DR as cytoplasmic signals. Signal spillover was subsequently compensated as previously described.¹²⁶ Segmented and compensated data were then analyzed with Qupath software.¹²⁷ In Qupath, cells with an area of 15 µm² were filtered out from downstream analysis. In each remaining cell, various signal data were calculated for each channel (mean, median, minimum and maximum, giving the range, and standard deviation). Based on the vimentin, cytokeratin, CD45, CD4, E-cadherin, CD68, CD8a, CD3 and Langerin channels for an image of healthy skin from a healthy donor, we manually annotated about 10 to 15 cells of each of the following cell subsets: CD4⁺ T cells, CD8⁺ T cells, macrophages, Langerhans cells, other CD45⁺ cells, keratinocytes and other CD45⁻ cells. These annotations were then used to train a random forest cell classifier directly in Qupath. This classifier was then applied to other images with occasional adjustments based on new annotations. In parallel, for quantification, we defined two ROIs in each image: the epidermis, identified with the pancytokeratin and E-cadherin channels, and the 200 µm of subepidermal dermis. This approach enabled us to analyze the composition of the epidermis and the subepidermal dermis accurately for each sample.

## Mouse papillomavirus (MmuPV1) infection

All mouse work was approved by the Institutional Animal Care and Use Committee of Pennsylvania State University's College of Medicine (COM), and all methods were performed in accordance with guidelines and regulations. The mice were housed (2–5 mice/cage) in sterile cages within sterile filter hoods in the COM BL2 animal core facility. Infectious mouse papillomavirus (MmuPV1) was isolated from lesions on the tails of mice from our previous studies.¹²⁸,¹²⁹ In brief, papillomas scraped from the tails of the mice were homogenized in phosphate-buffered saline (1×PBS) with a Polytron homogenizer (Brinkman PT10-35) at maximum speed for three minutes, with chilling in an ice bath. The homogenate was spun in a bench centrifuge at 10,000 rpm, the supernatant was decanted into Eppendorf tubes, and the resulting mouse papillomavirus suspension was stored in glycerol at -80°C (1:1,V/V). C57BL/6, Rag1⁻/⁻, and Flt3lg⁻/⁻ (4–6 weeks old) mice were obtained from Jackson Laboratories and reared in the animal core facility of PSUCOM. Three female Flt3lg-heterozygous mice in the B6 background were purchased from the Mutant Mouse Resource & Research Centers