EMBO/sd-character-level-ner huggingface dataset:
Dnmt2 methylates C38 of tRNA in the bone marrow and modulates the stability and fragmentation of substrate tRNAs.Figure 4Bisulfite sequencing maps of 454 reads showing the average methylation status of individual cytosines (boxes) within tRNAs from bone marrow (BM), mesenchymal stromal cells (MSC), and in vitro differentiated adipocytes and osteoblasts. Methylation levels are indicated on a color scale from green (unmethylated) to red (completely methylated). Representative Northern blot showing tRNA(Gly) and tRNA(Asp) fragmentation in bone marrow from 8-day-old and adult mice. tRNA(Ser) was included as a Dnmt2 non-substrate tRNA, and 5S rRNA was included as a loading control. tRNA fragment coverage for the Dnmt2 substrates tRNA(Gly) and tRNA(Asp), and the non-substrate tRNA(Ser) in bone marrow, as determined by deep sequencing. Wild-type coverage is represented in red, and Dnmt2−/− coverage is indicated in blue. The plots illustrate the specific increase of 5′-halves from tRNA-Gly and 3′-halves from tRNA-Asp fragments in Dnmt2 mutants.
Figure 6A Schematic tracing of presynaptic boutons from vHIP neurons using AAV-DIO-GFP virus. B Representative examples of GFP-labeled presynaptic boutons. Scale bars: 2.5 μm. C Quantification of presynaptic bouton density and weighted size in the mPFC (PL and IL). Mock: n= 57 axons from 6 mice; ZIKV: n= 63 axons from 7 mice. D Quantification of presynaptic bouton weighted size in the mPFC (PL and IL). Mock: n= 2974 boutons from 6 mice; ZIKV: n= 3486 boutons from 7 mice. Data information: All data were presented as mean ± SEM. *P<0.05, **P<0.01, Two-tailed unpaired t-test unless described otherwise. E Trans-synaptic tracing of vHIP-derived neurons in the mPFC. AAV1-hSyn-Cre injection in the ventral hippocampus (vHIP) of Ai14 mouse followed by examination of neuronal types in the mPFC. F Representative confocal images of co-labeled tdTomato and neuronal markers including CaMKII, PV, CAL, and SST in the mPFC. Scale bar, 50 μm. G Quantification of the percentages of mPFC neuron subtypes innervated by vHIP afferent axons. H Comparisons of percentages of vHIP-innervated mPFC (PL and IL) neuron subtypes. Mock, n=4 mice; ZIKV, n=7 mice. P= 0.5343 in CaMKIIa, 0.0117 in PV, 0.7491 in CAL, 0.4484 in SST, 0.1519 in other uncharacterized neuron subtypes. Data information: All data were presented as mean ± SEM. *P<0.05, **P<0.01, Two-tailed unpaired t-test unless described otherwise. I Schematic tracing of a retrograde dye CTB488-labeled vHIP neurons projecting towards mPFC. CTB488 was injected into the mPFC of vGLUT1-Cre::Ai14 mice followed by IHC staining in vHIP. J Representative confocal images of CTB488-labeled tdTomato-positive neurons in the vHIP. White arrowheads indicate CTB488 single positive neurons. Scale bars: 200 μm. K Quantification of the percentage of tdTomato;CTB488 double-positive cells out of total CTB488-positive cells. Mock, n= 7 mice; ZIKV, n=7 mice. Data information: All data were presented as mean ± SEM. *P<0.05, **P<0.01, Two-tailed unpaired t-test unless described otherwise.
– TLR8 recognizes UR/URR motif containing RNA.Figure 3A, B Cytokine release of 3 days differentiated (ddi) THP1 cells challenged with (A) ORNs or (B) bacterial RNA fractions (tot, total; S, Svedberg; r, ribosomal). C Responsiveness of Tlr8-/--3ddiTHP1 cells challenged with heat inactivated (hi) bacteria (triangle, decreasing doses; *, p ≤ 0.05; **, p ≤ 0.01) upon TLR2 blockade (T2.5). D Responsiveness of 3ddiTHP1 cells upon challenge with human (Hs) mitochondrial (mt) RNA (triangle, increasing doses). E-H NF-B driven relative luciferase activity (Rel. luc. act.) of hTLR8+ HEK293 cells or cytokine release by PBMCs all transfected with RNAs indicated and additional uridine (U) if indicated (n.p., not performed; vector, empty plasmid).
Figure 9(A) Representative western blots showing the levels of SQR, TST, ETHE1 and SUOX protein from the brain of 5 wild-type (WT) and five mutant (Mut) mice (B-E) Relative levels of proteins normalized to Vinculin.
IL-1β and TNF release in response to AdV and h9C12 monoclonal antibody is TRIM21 dependent (A) Schematic showing the h9C12 antibody mutants that no longer engage Trim21 (H433A) or FC receptors (LLAA).Figure 4Schematic showing the h9C12 antibody mutants that no longer engage Trim21 (H433A) or FC receptors (LLAA). Primed or (C) un- primed HMDM were stimulated with AdV (50 000 pp/cell) +/- h9C12 (20 μg/ml) for 16h. Ctrl or Trim21- deficient THP-1s were stimulated for 4 h with 1000 U/ml IFNα and TRIM21 expression measured by immunoblot. WT or TRIM21- deficient THP-1s were stimulated with AdV (50 000 pp/cell) and 20 μg/ml h9C12 or 20 mg/ml IVIg or with 200 ng/well HT-DNA or 10 μM Nigericin for 16h, and IL-1β measured in the supernatant by ELISA. WT or TRIM21- deficient THP-1s were stimulated with AdV (50 000 pp/cell) and 20 μg/ml h9C12 or 20 mg/ml IVIg or with 200 ng/well HT-DNA or 10 μM Nigericin for 16h, and TNF measured in the supernatant by ELISA. WT mice were injected i.v. with 2.5ug WT or H433A h9C12 antibody, then the next day injected i.v. with 2.5 x 1011 pp AdV-GFP. 4h later spleens were harvested and neutrophil influx measured by flow cytometry. HMDM were stimulated with AdV-GFP (250 pp/cell) in the presence of h9C12 antibodies at indicated doses. Infection was measured after 24h by flow cytometry. Infection relative to AdV alone is shown graphed on the left (n = 4 mean ± s.e.m). Representative plots and a comparison of the different h9C12 mutants at 0.2 ug/ml are also shown in the graph on the right(n = 4,mean ± s.e.m.).
Virions.Fig. 3viral particles in the ultrathin sections under electron microscope at 200 kV, sample from viral infected Vero E6 cells
Figure 2 (A) Confocal images of ALIX (red), Asl (white), α-tubulin (green) and Hoechst (blue) in wild type (w1118), Sas-6c02901, DSpdG02143, cnnHK21 and γ-Tub23CPI NBs (scale bars, 5 μm). The insets show higher magnification of the boxed centrosomes (B) The centrosomal fluorescence intensity of ALIX was measured and the graph shows the average intensity (relative to w1118 intensity) of three separate experiments (±S.E), where in total 78 w1118, 73 Sas-6c02901, 45 DSpdG02143, 61 cnnHK21 and 26 γ-Tub23CPI NBs were analyzed. Statistically significant differences are indicated (*p<0.05 and **p<0.01, Student's t-test) (C) GST or GST-ALIX were incubated with in vitro translated 35S-labelled Cnn or DSpd-2 proteins. Binding was detected by autoradiography (upper panel), and expression of GST and GST-ALIX was visualized by Ponceau S-staining (lower panel) (D) HeLa cells were transfected with control, Cep192 or Cep215 siRNA. Efficient depletion of Cep192 and Cep215 was confirmed by Western blotting analysis, which also showed unchanged expression levels of ALIX. Unspecific bands, below Cep192 and above Cep215, are indicated with * *. (E) Control, Cep192 or Cep215 siRNA-transfected HeLa cells were immunostained with anti-ALIX (red), anti-α-tubulin (green) and Hoechst (blue). Confocal micrographs are presented and the insets show the indicated centrosomes (arrows) at higher magnification (scale bars, 5 μm) (F) The centrosomal ALIX fluorescence intensity was measured and the graph shows the average intensity (relative to control siRNA cells) of three independent experiments (±S.E), where in total 41 control siRNA, 40 Cep192 siRNA and 39 Cep215 siRNA-transfected cells were analyzed, **p< 0.01, Student's t-test
a) Lysates of cells transfected with the indicated siRNAs blotted for NDP52, TBK1, IKKε and proliferating cell nuclear antigen (PCNA).figf6(a) Lysates of cells transfected with the indicated siRNAs blotted for NDP52, TBK1, IKKε and proliferating cell nuclear antigen (PCNA). Control, a nonspecific band detected by the IKKε antibody. (b,c) Numbers of bacteria recovered from HeLa cells transfected with the indicated siRNAs and infected with S. Typhimurium. At the indicated time points after infection, cells were lysed and bacteria counted on the basis of their ability to form colonies on agar plates. Mean and s.d. of triplicate HeLa cultures and duplicate colony counts. (d) Numbers of bacteria recovered from HCT116 cells expressing the indicated shRNAs and infected with S. Typhimurium. At the indicated time points cells were lysed and bacteria counted by flow cytometry. Mean and s.d. of triplicate cultures. (e) HeLa cells transfected with the indicated siRNAs were infected with S. Typhimurium expressing green fluorescent protein (GFP). At the indicated time points after infection, cells were scored according to the number of bacteria they contained. (f) HeLa cells transfected with the indicated siRNAs were infected with S. Typhimurium expressing GFP. At the indicated time points after infection, cells were stained for ubiquitin and infected cells were scored if they contained ubiquitylated bacteria. All data represent at least two independent experiments. *P 0.05, Student's t-test.
Figure 6(A) The amount of subgenomic E RNA in the panel of clinical samples used to infect Vero E6-TMPRSS2 cells was measured using a previously-published primer-probe set (Wölfel-sgE) (B) For a subset of 85 NP swab samples for which sufficient material remained, SARS-CoV-2 negative-strand E RNA levels were determined by strand-specific RT-PCR with tagged primers. Data information: Culture supernatants were harvested when cells displayed CPE or at day 7 if no CPE was observed earlier, clarified, and presence (blue square) or absence (yellow circle) of infectious SARS-CoV-2 determined by immuno-focus assay.
Phosphoinositide regulates LAPTM4B interaction with PIPKIγi5Co‐immunoprecipitation (co‐IP) of full‐length or N‐terminus‐deleted LAPTM4B with PIPKIγi5 in HEK293 cells.Figure 5A Co‐immunoprecipitation (co‐IP) of full‐length or N‐terminus‐deleted LAPTM4B with PIPKIγi5 in HEK293 cells. B GST‐tagged LAPTM4B N‐terminus (93 amino acids) and T7‐tagged PIPKIγi5 C‐terminus (223 amino acids) were purified from E. coli for in vitro GSTpull‐down assay. C Co‐IP of full‐length or N‐terminal deletion mutants of LAPTM4B with PIPKIγi5 in HEK293 cells. ∆25: amino acids 1-25 deleted. D Schematic diagram of LAPTM4B with polybasic motif (PBM) magnified and PBM mutants specified. E Coomassie Brilliant Blue staining of purified wild‐type and mutated LAPTM4B N‐termini. F Purified LAPTM4B N‐termini from (E) were used in PIP strips assay. LPA, lysophosphatidic acid; LPC, lysophosphocholine; PE, phosphatidylethanolamine; PC, phosphatidylcholine; S1P, sphingosine 1‐phosphate; PI3,4,5P3, PtdIns(3,4,5) P3, phosphatidylinositol (3,4,5)‐trisphosphate; PA, phosphatidic acid; PS, phosphatidylserine. GST pull‐down assay of wild‐type or 6RQ mutant of LAPTM4B N‐termini with PIPKIγi5 C‐terminus. H Co‐IP of wild‐type or PMB‐mutated LAPTM4B with wild‐type or kinase dead PIPKIγi5 in HEK293 cells. I Top: GSTpull‐down assay of LAPTM4B N‐terminus and PIPKIγi5 C‐terminus with increased concentration of PtdIns(4,5) P2 addition. Bottom: Quantification of the relative amounts of PIPKIγi5 C‐terminus bound to LAPTM4B N‐terminus in top panel (mean ± SD; n = 4). J LAPTM4B N‐terminus binds multiple phosphoinositides including PtdIns(4,5) P2 in liposome‐binding assay. K Top: GSTpull‐down assay of LAPTM4B N‐terminus and PIPKIγi5 C‐terminus with 0.5 μM addition of different phosphoinositides. Bottom: Quantification of the relative amounts of PIPKIγi5 C‐terminus bound to LAPTM4B N‐terminus in top panel (mean + SD; n = 4). L Top: GSTpull‐down assay of wild‐type or 6RQ mutant of LAPTM4B N‐terminus and PIPKIγi5 C‐terminus with 0.5 μM addition of PtdIns(4,5) P2. Bottom: Quantification of the relative amounts of PIPKIγi5 C‐terminus bound to LAPTM4B N‐terminus in top panel (mean + SD; n = 4).
Figure 6DNA methylation levels of the HES5 promoter region of SH97Q cells treated with DHT and RG108. HES5 mRNA levels of SH97Q cells treated with DHT and RG108. DNA methylation levels of the Hes5 promoter region in DHT-treated NSC97Q cells treated with or without RG108. mRNA levels of Hes5 (n = 3) in DHT-treated NSC97Q cells treated with or without RG108. (E) Hes5 mRNA levels in the spinal cords of AR-97Q mice treated with or without RG108 (n = 3). (F, G) Hes5 immunoreactivity of spinal motor neurons in AR-97Q mice treated with or without RG108 (n = 3).
Figure 2(A) The molecular surface of hCAP-G is shown in orange. hCAP-H is shown as a green ribbon model. The four major contact sites (I, II, III and IV) are boxed. (B-E) Zoomed-in views of sites I-IV. Residues of hCAP-G and hCAP-H are labeled in white or black and green, respectively.
Figure 3(A) List of top hits of EZH2-bound proteins identified by TAP-MS. The number of peptides and coverage of each protein are indicated. (B) Reciprocal co-IP of endogenous EZH2 and eIF4G2 or PABP1 in C4-2 cells. (C) Co-IP of endogenous EZH2 with eIF4G2 or PABP1 from C4-2 cell lysate pre-treated with RNaseA in 37oC for 30 min. The effectiveness of RNaseA treatment was monitored by RT-PCR analysis of the presence of GAPDH mRNA. (D) C4-2 cells were transfected control (siC) or EZH2-specific siRNA and lysed for polysome fractionation. RNA was extracted from even-number fractions followed by RT-qPCR analysis for p53 and GAPDH mRNA. The β-ACTIN mRNA was used as an internal control. Data shown as means±SD (n=3). Statistical significance was determined by two-tailed Student's t-test. * P<0.01; NS, no significance. (E) C4-2 cells were transfected with control (siC) or EZH2-specific siRNAs in combination with empty vector, EZH2 WT or different deletion mutants followed by polysome fractionation and RT-qPCR. β-ACTIN mRNA was used as an internal control. Data shown as means±SD (n=3). * P<0.01.
SesB hypomorphic flies have an enhanced autophagy phenotype, and epistasis analysis indicates that SesB acts downstream of Dcp-1.Figure 8.(A) Ovaries from fed SesBOrg flies show reduced Ref(2) P staining in degenerating midstage egg chamber (arrow). (B) Ovaries from fed w1118 and SesBOrg were analyzed by Western blotting using Ref(2) P. Actin served as a loading control. Results are representative of three independent experiments (n = 3). (C) SesBOrg flies expressing GFP-mCherry-DrAtg8a in the germline showed increased autophagosomes and autolysosomes in nondegenerating (left) and degenerating (right) midstage egg chambers. Insets show autophagosomes and autolysosomes (left) and autolysosomes (right) in SesBOrg midstage egg chambers. (D) Quantitation of percentages of autolysosomes (autolysosomes/total autophagic structures) in nondegenerating ovaries from fed w1118 and SesBOrg flies. 10 egg chambers were manually examined per genotype. Error bars represent the means ± SD. Statistical significance was determined using a two-tailed Student's t test (***, P < 0.0001). (E) Degenerating midstage egg chambers (arrows) from fed SesBOrg flies are TUNEL positive. (F) Degenerating midstage egg chambers from starved control SesBOrg/FM7a;Dcp-1Prev1/Dcp-1Prev1flies contained persisting nurse cellnuclei and increased Ref(2) P. n = 117 ovarioles examined. Box represents zoomed image. (G) Degenerating midstage egg chambers from fed SesBOrg;Dcp-1Prev1flies contained condensed and fragmented nurse cellnuclei and low levels of Ref(2) P. Box represents zoomed image. n = 156 ovarioles examined. (H) In response to starvation, Dcp-1 acts to negatively regulate the levels of SesB. SesB is a negative regulator of autophagy, and reduced SesB levels mediated by Dcp-1 lead to decreased ATP and induction of autophagic flux.
CRP-mediated transcriptional control of vcdRPA.Figure 2A. Northern blot analysis of the expression of VcdRP and Spot42 sRNA monitored at low cell density (OD600 of 0.1) and high cell density (OD600 of 2.0), in the absence (-) or presence (+) of cAMP (5mM f.c.) supplemented externally in LB medium. The solid triangle indicates the band corresponding to the full-length primary transcript. Probing with 5S rRNA confirmed equal loading. B. Relative fluorescence intensities of V. cholerae wild-type or mutants lacking crp or cyaA harboring a transcriptional reporter plasmid of vcdRP fused to mKate2. Cells were grown in LB to OD600 of 1.0 and fluorophore production was measured. The fluorescence of WT was set to 1. C. Electrophoretic mobility shift assay (EMSA) showing that CRP protein binds to the vcdRP promoter sequence. Migration of the [32P] end-labeled vcdRP promoter fragment in the absence and presence of increasing concentrations of purified CRP protein (indicated by the black triangle above the gel) was determined by native polyacrylamide gel electrophoresis and autoradiography. The open triangle indicates free DNA whereas the filled triangle indicates VcdRP in complex with CRP. Final concentrations of CRP used in each reaction from left to right (lanes 1-8) were: 0, 0.35, 0.7, 1.4, 2.1, 2.8, 3.5, and 4.2 µM. Radiolabelled DNA and CRP were incubated at 37°C for 20 minutes in buffer containing 0.2 mM cAMP, before being loaded on a 7.5 % non-denaturing polyacrylamide gel.
Figure 5Cartoon representation of the EschOTU crystal structure colored in a rainbow gradient from N- to C-terminus. The catalytic triad residues are marked on both the structure and the linear color gradient above, showing their positions with respect to each other and the overall OTU sequence. The black and grey arrows relate how the EschOTU fold is permutated with respect to other OTUs. The black open arrow marks the open N- and C-termini, while the closed grey arrow marks a closed loop. OTU subfamilies that follow this overall sequence topology are listed in the lower right. This arrangement is only observed in EschOTU. As in A, for the human OTUB1 structure (PDB 4DDG). The closed black arrow marks a closed loop, while the open grey arrow marks the open N- and C-termini. This arrangement is representative of the human Otubain, OTUD, and OTULIN subfamilies, as well as vOTUs. As in A, for the human Cezanne structure (PDB 5LRW). This arrangement is representative of the human A20 subfamily, viral PLP2, and Legionella ceg23. Cartoon representation of the CCHFV vOTU crystal structure (PDB 3PHW) colored in a rainbow gradient from N- to C-terminus. The catalytic triad residues are marked on both the structure and the linear color gradient above, showing their positions with respect to each other and the overall OTU sequence. The closed black arrow marks a closed loop, while the open grey arrow marks the open N- and C-termini. A schematic for the permutated vOTUP variant is shown to illustrate how it relates to the native sequence topology. Ub-PA activity-based probe assay for WT vOTU and sequence-permutated vOTUP. Strong reactivity is indicated with asterisks. Ub-KG(TAMRA) cleavage assay monitored by fluorescence polarization for WT vOTU and sequence-permutated vOTUP.
Microbial dysbiosis in OGT-deficient mice.Figure 5 A PCoA plot of unweighted UniFrac distance of bacterial communities in wildtype and Vil-Ogt KO mice that were singly housed at Yale or UMN (UMN KO n = 4, rest n = 5)  B Cladogram showing the most discriminative bacterial clades between wildtype and Vil-Ogt KO mice that were identified by LEfSe. Regions in red indicate clades that were enriched in Vil-Ogt KO mice, while regions in green indicate clades that were enriched in wildtype mice  C-H Antibiotic-treated C57BL/6 male mice were transplanted with fecal microbiota from wildtype or Vil-Ogt KO mice and then induced with DSS for colitis (WT n = 5, KO n = 7). (C) Daily changes in body weight  C-H Antibiotic-treated C57BL/6 male mice were transplanted with fecal microbiota from wildtype or Vil-Ogt KO mice and then induced with DSS for colitis (WT n = 5, KO n = 7) (D) Colitis scores  C-H Antibiotic-treated C57BL/6 male mice were transplanted with fecal microbiota from wildtype or Vil-Ogt KO mice and then induced with DSS for colitis (WT n = 5, KO n = 7) (E) Intestinal barrier functional assays measuring serum FITC-dextran at day 10  C-H Antibiotic-treated C57BL/6 male mice were transplanted with fecal microbiota from wildtype or Vil-Ogt KO mice and then induced with DSS for colitis (WT n = 5, KO n = 7) (F) H&E staining of colon tissues. Ulcer areas are designated by red circles. Scale bars = 200 μm  C-H Antibiotic-treated C57BL/6 male mice were transplanted with fecal microbiota from wildtype or Vil-Ogt KO mice and then induced with DSS for colitis (WT n = 5, KO n = 7) (G) Pathological scores of the mucosal injury  C-H Antibiotic-treated C57BL/6 male mice were transplanted with fecal microbiota from wildtype or Vil-Ogt KO mice and then induced with DSS for colitis (WT n = 5, KO n = 7) (H) RT-qPCR of inflammatory markers in the colon. (D: P = 0.0409; E: P = 0.0457; G: P = 0.032
Electron microscopy of macrophages 10 h after infection with HSV-1.figf4 Electron microscopy of macrophages 10 h after infection with HSV-1. (a) Arrows indicate membrane-coiled structures emerging from the nucleus of an infected cell. (b-d) Four-layered membrane structures formed by coiling of the nuclear membrane. (e,f) Glucose-6-phosphatase (black deposits) on autophagosome-like structures emerging from the nuclear envelope or free in the cytoplasm, and viral capsids in the cytoplasm engulfed in the lumen of an autophagosome-like compartment. N, nucleus; VP, viral particles. Scale bars, 1 μm (a), 0.25 μm (b-d,f) or 0.4 μm (e). Images are representative of three independent experiments with at least 100 cell profiles in each.
Figure 7A, In Rb1d/d mice with pre-implantation P4 treatment, the fragmented uterine epithelial cells with the cytoplasmic lipid droplets were engulfed by the trophoblast cells. Scale bar = 2µm. Arrowhead, cytoplasmic fragments engulfed by trophoblast; dotted line circle, lipid droplets in the cytoplasm; tr, trophoblast; le, luminal epithelium; s, stroma; red dotted line, stroma; green dotted line, luminal epithelium; blue dotted line, trophoblast. B, Pre-implantation P4 treatment rescued the expression of MFG-E8 on day 5 of pregnancy in Rb1d/d uteri. Scale bar = 100μm. Blue signal, nuclei stained by DAPI; yellow signal, MFG-E8; le, luminal epithelium; s, stroma; arrowhead, embryo. C, Pre-implantation P4 treatment recovered the expression of pRIP3, a mediator of necroptosis, on day 5 of pregnancy in Rb1d/d uteri. Scale bar = 100μm; blue signal, nuclei stained by DAPI; purple signal, pRIP3; le, luminal epithelium; s, stroma; arrowhead, embryo.
Figure 4(A) ChIP analysis of TAP-tagged Rad6 presence at ADH1, PMA1 and YEF3 coding regions (5' & 3') in WT or mog1Δ strains. Bar charts indicate the mean and standard deviation for at least three independent experiments. Significance of the differences was obtained using Student´s t-test (*, P value = 0.01-0.05; **, P value = 0.001-0.01, ***, P value = 0.0001-0.001, ****, P value <0.0001). (B), (C) and (D) Bre1-TAP, Rtf1-TAP and Rpb3 respectively, were analyzed as described in (A). (E) Bar charts showing the relative chromatin occupancy to 5'ORFs (left panel) or 3'ORFs (right panel) of each factor relative to Rpb3 occupancy at these gene positions.