EMBO/soda-nel huggingface dataset:
Microglial PD-1 modulates neuroinflammation and Alzheimer's disease pathology by  astrocytic PD-L1 (2021)
Figure 5

Fig. 5: PD-1 deletion reduces phagocytosis and CD36 expression in microglia

A) In vitro phagocytosis of FAM-Aβ1-42 by wild type or PD-1-/- microglia for up to 6 h (mean +/-SEM of a technical quadruplicate, Student's t-test, ** p<0.01, *** p<0.001, representative result of an n=2 biological replicate).

B) In vitro phagocytosis of FAM-Aβ1-42 by wild type microglia cultured in astrocyte-conditioned medium (ACM) from wild type or PD-L1-/- astroglial cultures (mean +/-SEM of a technical quadruplicate, Student's t-test, * p<0.05, *** p<0.001, representative result of an n=2 biological replicate)

C) Flow cytometry analysis of microglia from wild type and PD-1-/- mice for the expression of CD36 and CD11b.

D) Quantification of microglial in vivo phagocytosis in female APP/PS1 and APP/PS1 PD-1-/- mice at 8.5 month of age after intraperitoneal injection of methoxy-XO4 (biological replicates with n=5 for APP/PS1 and n=4 for APP/PS1 PD-1-/- mice, Student's t-test (t = 2.1808, df = 6.9829), * p<0.05).

E-G) Relative expression of the cell surface marker (E) CD36 (biological replicates with n=5 for APP/PS1 and n=4 for APP/PS1 PD-1-/- mice, ANOVA (DF=1, F= 0.002, p=0.94), Tukey's HSD, * p<0.05, *** p<0.001), (F) CD11b (biological replicates with n=5 for APP/PS1 and n=4 for APP/PS1 PD-1-/- mice) and (G) CD45 on methoxy-XO4+ versus methoxy-XO4- microglia in APP/PS1 and APP/PS1 PD-1-/- mice normalized to methoxy-X04-negative cells from APP/PS1 mice (biological replicates with n=5 for APP/PS1 and n=4 for APP/PS1 PD-1-/- mice)

Data information: (D-G) central bands represent median, boxes show interquartile range (IQR) and whiskers define the +/-1.5xIQR.
Microglial PD-1 modulates neuroinflammation and Alzheimer's disease pathology by  astrocytic PD-L1 (2021)
Figure 6

Fig. 6: Transcriptome analysis of PD-1-/- microglia

A) Top 5 enriched KEGG pathways of differentially expressed genes (abs(fold change) > 2, p < 0.01, Benjamini-Hochberg adjusted) in PD-1-/- vs. wild type microglia.

B) Different expression of inflammasome-related genes (abs(fold change) > 1.5, p < 0.05, Benjamini- Hochberg adjusted) in PD-1-/- vs. wild type microglia.

C) Different expression of complement system-related genes (abs(fold change) > 1.5, p < 0.05, Benjamini-Hochberg adjusted) in PD-1-/- vs. wild type microglia.

Expanded View Figure legends
Alternative oxidase-mediated respiration prevents lethal mitochondrial cardiomyopathy (2018)
Figure 1

Figure 1: AOX expression prolongs the survival of cIII deficient Bcs1lp.S78G mice without affecting growth or whole-body metabolism.

A. Schematic presentation of the multiorgan manifestations, described in this study or previously, in homozygous Bcs1lc.A232G (GRAC) mice, and the time points of the investigations performed in this study.

B. Survival curves of Bcs1l homozygotes without (GRAC) and with (GROX) alternative oxidase (AOX) expression (n=18-21/group). The median survival of GRAC mice was 210 days and of GROX mice 589 days with no gender difference.

C. Weight of mice from P50 to P200 (n>10/group) and at P600 (n=4-6/group).

D. Blood lactate to glucose ratio at P200 (n=8/group).

E-G. Dual energy X-ray absorptiometry (DEXA) analysis (n=10/group) of (E) lean mass, (F) fat mass and (G) bone mineral density at P98.

H, I. Indirect calorimetric measurement (n=10/group) of (H) heat production and (I) respiratory exchange ratio at P77.

Statistics: The survival data was analyzed using Log-rank (Mantel-Cox) test (p<0.0001). Bar graphs represent mean±SD. The data (D-I) were analyzed using Kruskal-Wallis and Mann-Whitney U tests for selected comparison. Significant differences between groups (p value) are indicated on graphs.
Alternative oxidase-mediated respiration prevents lethal mitochondrial cardiomyopathy (2018)
Figure 2

Figure 2. AOX prevents lethal cardiomyopathy and progression of renal tubulopathy to kidney atrophy.

A. Hematoxylin-eosin-stained cross sections of heart and kidney at P200.

B. Weight of heart and kidney at P200 (n=4/group).

C-D. Sirius Red staining (C) for fibrosis in liver, heart and kidney at P200. And (D) Quantification of fibrosis in myocardium, kidney cortex and liver (n=5-7).

E-F. Echocardiography data (n=4-6/group) showing (E) ejection fraction (EF) and fractional shortening (FS). (F) Systolic (LV Vol;S) and diastolic (LV Vol;D) left ventricle volume in mice.

G. Expression of cardiac hypertrophy and fibrosis-associated genes in the pre-symptomatic (P150) heart (n=6/group).

H, I. 24 h excretion of (H) albumin (I) creatinine in urine at P200 (n=4/group).

J, K. Liver enzymes (J) alanine aminotransferase (ALT) and (K) alkaline phosphatase (ALP) in plasma at P200 (n=8/group).

Bar graphs represent mean±SD. Statistics: one-way ANOVA followed by Tukey's test (for graphs B, D, G, H, J, and K), One-way ANOVA followed by unpaired t-test with Welch's correction for graph E and F, and Mann-Whitney U tests (for graph I).
Alternative oxidase-mediated respiration prevents lethal mitochondrial cardiomyopathy (2018)
Figure 3

Figure 3. AOX ameliorates cerebral astrogliosis and maintains mitochondrial ultrastructure in rescued tissues.

A. GFAP staining for cerebral astrocytes, the barrel field of the primary somatosensory cortex (S1BF) highlighted with brackets.

B. Mitochondrial ultrastructure in cardiomyocytes, kidney tubular cells and hepatocytes at P200 as visualized by electron microscopy. Scale bar 1 µm.

C-F. (C) Average cross-sectional area of mitochondrion, (D) number of cristae per mitochondrion, (E) cristae thickness, and (F) average distance between cristae in mitochondria. (n=3mice/group)

Bar graphs represent mean±SD. Statistics for graphs (C-F): One-way ANOVA followed by Tukey's test.
Alternative oxidase-mediated respiration prevents lethal mitochondrial cardiomyopathy (2018)
Figure 4

Figure 4. AOX mitigates cardiac, but not hepatic or renal, metabolic stress-related gene expression changes.

A-C. Principal component analysis of transcriptome data from (A) heart, (B) kidney and (C) liver.

D-F. Heat map visualization of pathway enrichment analysis (Reactome database). Full pathway analysis is provided in Appendix Table S1. Benjamin-Hochberg FDR-corrected p values are color-labelled as indicated for the three comparisons.

G-K. Gene expression of major transcriptional regulators of energy metabolism (PPAR-α, HIF-1α), mitochondrial biogenesis (PGC-1α, TFAM) and stress responses (ATF4) in heart at P150. (n=6/group)

L. Expression of cIII stress signature genes in heart, kidney, and liver. (n=6/group)

Box plot (G-K) whiskers represent quartiles, maximum and minimum value (relative fold change 'FC' to WT). Statistics: one-way ANOVA followed by Tukey's test.
Alternative oxidase-mediated respiration prevents lethal mitochondrial cardiomyopathy (2018)
Figure 5

Figure 5. AOX has little effect on cardiac energy metabolites but attenuates proline accumulation at onset of disease.

A. Significantly altered metabolites (FDR<0.2; P<0.05, n=5/group) in presymptomatic (P150) heart tissue of GRAC (red) and GROX (green) mice.

B. Significantly altered metabolites (FDR<0.2; P<0.05, n=5/group) in liver tissue of GRAC (red) and GROX (green) mice.

C-F. Concentrations of the TCA cycle intermediates (C) fumarate and (D) malate, (E) adenylate energy charge and (F) NADH/NAD+ ratio (n=5/group).

G-I. (G) Proline concentration in heart tissue at P200 (# below detection limit) and mRNA expression of proline synthesis-related genes (n=6/group)) (H) Aldh18a1 and (I) Pycr1 at P150.

Bar graphs (A-G) represent mean±SD, and box plots of mRNA expressions represent quartiles, minimum, and maximum value (relative fold change 'FC' to WT). Statistics for graphs (A-B, F, and H): one-way ANOVA followed by Tukey's test.
Alternative oxidase-mediated respiration prevents lethal mitochondrial cardiomyopathy (2018)
Figure 6

Figure 6. AOX modifies RISP assembly and rescues mitochondrial respiration in tissue-specific fashion.

A. Blue native PAGE analysis of RISP and CORE1 assembled into free cIII2 and supercomplexes (SCs).

B-C. RISP per CORE1 ratio in (B) SC1 and (C) free cIII2.

D-F. Spectrophotometric assay of (D) cI, (E) cII, and (F) cIII activity in isolated mitochondria at P200 (n=4-6/group).

G. CIV activity as measured using an oxygraph.

H. n-Propyl gallate-sensitive AOX mediated cI and cII-linked state 3 respiration (n=7/group, Mann-Whitney U test).

I and J. CI-linked (I), and cI- and cII-linked (J) state 3 respiration in isolated heart, kidney and liver mitochondria (n=8/group). We used malate, pyruvate and glutamate to generate NADH for the cI. Subsequently, we added the CII substrate, succinate, to obtain CI&CII-linked respiration in presence of saturating ADP.

Bar graphs represent mean±SD. Statistics: one-way ANOVA followed by Tukey's test. Significant differences between groups (p value) are indicated on graphs.
Alternative oxidase-mediated respiration prevents lethal mitochondrial cardiomyopathy (2018)
Figure 8

Figure 8. AOX normalizes cardiac nitric oxide-related gene expression.

A. Heatmap visualization of the expressions of selected nitric oxide (NO)-related genes.

B-E. Gene expression of Ca2+ channel genes (B) ryanodine receptor, RYR2, (C) sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA2 or ATP2A2), (D) phospholamban (PLN), and (E) endothelial nitric oxide synthase NOS3 (n=6/group).

F and G. Expression of neuronal nitric oxide synthase (NOS1) mRNA (F), and (G) protein with a representative Western blot. WT brain was used as positive control in the 1st lane (G).

H. Cardiac gene expression of dimethylarginine dimethylaminohydrolase 1 (DDAH1) (n=6/group).

I. Cardiac citrulline to arginine ratio (n=5/group).

J. Cardiac protein nitrotyrosine quantified from Western blots.

K. Nitrite concentration in heart tissue (n=4/group).

Box plots for mRNA expression (B-E, and H), represent quartiles, minimum and maximum value (relative fold change 'FC' to WT). Bar graphs (F,I, and K) represent mean±SD. Statistics: one-way ANOVA followed by Tukey's test for graphs B-F, and H. Significant differences between groups (p value) are indicated on graphs.
Human chromosome-specific aneuploidy is influenced by DNA-dependent centromeric features (2019)
Figure 4

Figure 4. Faithful chromosome segregation is biased in favor of centromeres carrying a high frequency of CENP-B/CENP-C molecules.

A. Schematic of RPE-1 chromosome spreads used in experiment (B) and (C).

B. Representative immunofluorescence FISH (IF-FISH) images to measure CENP-B intensity at specific chromosome in untreated cells. Scale bar represents 5 µm.

C. Bar plot showing the normalized CENP-B fluorescence intensity at every chromosome over the mean on each metaphase spread (n > 37 per chromosome) ± SEM. Acrocentric chromosomes 13, 14, 21 and 22 were not analyzed and are marked by a line. One-way ANOVA with post-hoc Tukey's multiple comparison test shows high diversity between chromosomes. t-test against the mean was used to estimate the statistical significance for each chromosome. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.

D. Scatter plot showing a significant negative correlation between the mean of CENP-B IF-FISH signal (n > 37) and the mis-segregation rate (from Fig. 1F) (r = Spearman rank coefficient). The lines represent linear regression with 95% confidence band.

E-F. Bar graphs report the sum of the normalized reads counts of different HOR arrays (see Table EV2) representing CENP-C binding following CUT&RUN, sequencing and centromere mapping. Cells were treated for 6 h with IAA to deplete CENP-A. Error bars represent the SEM of three independent experiments and the dashed line represents the mean. Acrocentric chromosomes 13-14, 21 and 22 were marked by a line. t-test against the mean was used to estimate the statistical significance for each chromosome. *p < 0.05; **p < 0.01; ***p < 0.001.

G. Scatter plot showing a significant positive correlation between the mean of CENP-C reads (n = 3) and CENP-B reads (n = 3) (r = Spearman rank coefficient). Data from chr 1, 5 and 19 were excluded from the analysis in addition to chr 13, 14, 21 and 22 to better assess correlation without the FISH correction (as in sup figure EV2 I). The lines represent linear regression with a 95% confidence interval.

H. Scatter plot showing a significant negative correlation between the mean of CENP-C reads (n = 3) and the mis-segregation rate (from Fig. 1F) (r = Spearman rank coefficient). The lines represent linear regression with 95% confidence interval. Data from chr 13, 14, 21 and 22 were excluded from the analysis. The lines represent linear regression with a 95% confidence interval.
Human chromosome-specific aneuploidy is influenced by DNA-dependent centromeric features (2019)
Figure 5

Figure 5. Chromosome-specific aneuploidy arises following perturbation of centromere strength.

A. Schematic of the centromere strength model in the RPE-1 CENP-AAID CENP-B KO mutant. After CENP-A depletion a reduced bias in chromosome mis-segregation is expected.

B. Representative image of centromere 11 (red) and centromere X (green) FISH on RPE-1 CENP-AAID CENP-B KO mutant in control (top) or IAA treated condition (bottom). Scale bar represents 5 µm. Bar graphs show automatic FISH quantification of chromosome mis-segregation in the indicated cell lines with and without IAA addition for 24 h. Error bars represent the SEM of three independent experiments (n> 200 cells per experiment). Dunnett's multiple comparisons test was used to compare conditions. Grey lines separate independent experiments. Fold changes between grey and red bars are also indicated. C. Schematic of the centromere strength model with the microtubule destabilizer BAL27862 compound.

D. Representative image of centromere 17 (red) and centromere X (green) FISH on RPE-1 cells treated with the BAL27862 compound. Scale bar represents 5 µm.

E. Bars represent the frequency of chromosome mis-segregation into micronuclei after 28 h treatment with the BAL27862 drug at indicated concentration. Error bars represent the SEM of three independent experiments, dashed line represents the mean. n > 40 micronuclei were analyzed. Dunnett's multiple comparisons test, *: p<0.05. DMSO was used at the same concentration of BAL27862.
Human chromosome-specific aneuploidy is influenced by DNA-dependent centromeric features (2019)
Figure 6

Figure 6. Different level of CENP-B across centromeres dictate chromosome segregation outcome.

A. Representative image of CENP-B box FISH and mFISH stained RPE-1 metaphase spread cells. Chromosome 3 homologs are indicated by white arrows with single CENP-B box chromosomes signal magnified in insets. Scale bar represents 5 µm.

B. Representative image of CENP-B box (red) and centromere 3 (green) FISH on RPE-1 interphase nucleus after 48 h IAA treatment. CENP-B box signal at individual homologs are magnified in insets. Scale bar represents 5 µm.

C. Bar plots represent the quantification of the CENP-B box FISH intensity at the chromosome 3 homolog contained in the main nucleus versus the homolog present in the micronucleus as represented in (B). Error bars represent the SEM of three independent experiments. n = 69 cells. Unpaired t test, ****p < 0.0001.

D. Schematic of the experiment shown in (E-G). ΔN-CENP-B protein was fused to dCas9-doxycycline (Dox) inducible protein and was inserted at a single genomic locus in DLD-1 CENP-A-/EA male cell line expressing a gRNA targeting centromere Y.

E. Immunoblot shows expression of the (ΔN)CENP-B-dCas9 after doxycycline induction.

F. Representative images show a micronucleus or the main nucleus containing the Y chromosome (red) detected by FISH in no-doxycycline or doxycycline treated cells respectively. Chromosome X is shown in green. Scale bar represents 5 µm.

G. Bar plot shows the mean (n ≥ 3) frequency of micronuclei containing the chromosome Y or chromosome X ± doxycycline treatment for 72 h and IAA for 48 h ± SEM. n > 200 cells with a micronucleus. Mann-Whitney test. *p=0.0173.
Human chromosome-specific aneuploidy is influenced by DNA-dependent centromeric features (2019)
Figure 7

Figure 7. Abundance of CENP-B boxes influences segregation fidelity in mouse but has a weaker impact in a cancer model system.

A. Schematic of the experiment shown in (B).

B. Table summarizing whole-chromosome aneuploidy (fold over the mean) using the indicated methods to measure chromosome mis-segregation rate. Bold numbers represent significant statistical difference from the mean for each method. Orange to dark red gradient highlights chromosomes that mis-segregate (with at least one method) at a significantly higher rate compared to the mean level (weak chromosomes). Light green to dark green gradient highlights chromosomes that mis-segregate (with at least one method) at a significantly lower rate compared to the mean (stronger chromosomes). Data obtained with only one method were excluded. Im. St. = image stream.

C. Plot shows the karyotype distribution of DLD-1 and RPE-1 cells in untreated or Auxin-treated condition

D. Schematic of the experiment shown in (E).

E. Representative image of a chromosome spreads in DLD-1 cells following mFISH and subsequently CENP-B boxes FISH. Inset shows the supernumerary (SN) chromosome. Scale bar represents 10 µm.

F. Bar plot shows the mean variation in CENP-B boxes in DLD-1 cells from data from E (n = 10 cells) and whole-genome sequencing and centromere mapping ± SD.

G-H. Scatter plot showing a significant positive correlation between the indicated features in DLD-1 cells (n = 1). The lines represent linear regression with 95% confidence band. Data from chr 13, 14, 21, 22, Y and SN were excluded from the analysis.

I. Schematic of the experiment described in (J) and (K).

J. Bars show frequency of micronuclei with or without siRNA treatment. Error bars represent the SEM of three independent experiments (n > 170 cells per replicate).

K. (Left) Representative image of CENP-B box (red) staining after siRNA against CENP-A/HJURP for 48 h. CENP-B box signals at individual micronuclei are magnified in insets. Scale bar represent 5 µm. (Right) Bar graph represents the mean of the CENP-B box FISH intensity contained in the main nucleus versus the one present in the micronucleus ± SEM. Color dots represent independent experiments. (n > 43 micronuclei and 840 main nucleus) Mann-Whitney U test: ***p = 0.0001.
Human chromosome-specific aneuploidy is influenced by DNA-dependent centromeric features (2019)
Figure 8

Figure 8. DNA-dependent centromeric feature and chromosome size influence chromosome-specific aneuploidy.

A-B. Scatter plot showing (A) a significant positive or (B) a negative correlation between chromosome length and the mis-segregation rate (from Fig. 1F) or CENP-C (n = 3) reads over chromosome length and mis-segregation rate (r = Spearman rank coefficient). The lines represent linear regression with 95% confidence interval. Data from chr 13, 14, 21 and 22 were excluded from the analysis. Numbers indicate chromosomes with low or high mis-segregation frequency.

C. Model of how chromosome segregation is influenced by centromere size and chromosome length.

Figure Legend for EV Figures
Notch and EGFR regulate apoptosis in progenitor cells to ensure gut homeostasis in Drosophila (2019)
Figure 2

Figure 2. Physiological caspase-dependent apoptosis of enteroblast cells in the adult midgut

(A) Schematic representation of the apoptotic pathway in Drosophila.

(B) Lineage tracing using the initiator caspase sensor act-DBS-S-QF (14 days of tracing) combined with QUAS-FLP and act>FRT>lacZ are visualized by ßGalactosidase (ßGal, green). ISCs are labelled by anti-Dl (red, insets). Red arrowhead points to the ISC. Gut is counterstained with DAPI (blue) and DAPI (grey, in the inset).

(C) Control GBE-Su(H)ReDDM> and (D) GBE-Su(H)ReDDM>p35, p35 midguts after 7 days of tracing.

(E-E'') Adult midgut klu>mCD8::GFP, GBE-Su(H)-lacZ stained for ß-Gal (red) and GFP (green). DAPI counterstaining in blue (E' and E''). The arrows point to examples of klu-negative Su(H)-lacZ-positive cells. The green label (asterisk, E) is in the overlying muscle.

(F-J) Adult midguts of (F) control kluReDDM> (G) kluReDDM>Debcl-RNAi, (H) kluReDDM>Drice-RNAi, (I) kluReDDM>Diap1-RNAi, (J) kluReDDM>Diap1 midguts after 7 days of tracing. The arrows point to examples of PH3+ labelled ISCs.

(K) Quantification of klu+ cells in the indicated midguts at day 7 in low demand (Low) or standard (Variable) culturing conditions (n=7, 5, 9, 7).

(L) Intestinal cell turnover measured by quantification of the number of newly renewed ECs (RFP-only cells: gut scored n=9, 13, 9, 13).

(M) Quantification of apoptotic progenitor cells (esg+) measured as GFP+ TUNEL+ cells (n= 9, 9, 9, 9).

(N) Quantification of klu+ cells and renewed ECs in kluReDDM>Diap1 posterior midguts (n=8) relative to 'Low' control in K and L, respectively.

Data information: (K-N) Box and whisker plot showing min to max values and the box is extending from the 25th to 75th percentiles. The line in the middle of the box is plotted at the median. P values (*, P<0.0261; **, P<0.005; ***, P<0.0021) calculated by one-way ANOVA with Bonferroni correction. Scale bars, 100µm, except in E, 50µm.
Notch and EGFR regulate apoptosis in progenitor cells to ensure gut homeostasis in Drosophila (2019)
Figure 3

Figure 3 Enteroblast life and death decision is governed by opposing Notch and EGFR activity

(A-E) Confocal images of representative midguts of the indicated genotypes using kluReDDM for tracing and age- and EB-specific transgene expression at day 7 after temperature shift. ISC mitosis was monitored by using PH3 labelling (white) in all guts. (B-E) Accumulation and loss of klu+ cells by loss (B and E) and gain (C and D) of N and EGFR signalling. In (C), the accumulation of EC-committed EBs (klu+ cells) in midguts with constitutively activated EGFR (kluReDDM> EGFRact) is accompanied with a non-autonomous increase in ISC mitosis. Inset in (C) shows labelling with Dl (blue), revealing symmetric ISC mitoses as reported previously (Jiang and Edgar, 2009).

(F) Quantification of the number of klu+ (EBs) in the indicated midguts (n= 21, 14, 5, 6, 8, 6, 4, 10, 12, 9, 13, 14).

(G) Quantification of apoptotic esg+ (TUNEL+ GFP+) cells in the indicated midguts (n= 28, 4, 7, 7, 16, 8, 8, 7, 10, 7, 20, 24). Quantification of PH3+ and EC renewals is in Fig EV2A.

(H-M) Confocal images of representative midguts of the indicated genotypes as in (A-E). (H and I) Rescue of the loss of EBs induced by gain of Notch (H-RNAi) or the loss of EGFR (EGFRDN) by expressing Debcl-RNAi. (J and K) Impact of gain of expression (J) and loss by RNAi expression (K) of lz using kluReDDM (see Fig EV2A for PH3+ quantification). White arrowheads point to examples of mitosis labelled by PH3+. (L and M) Impact of gain of expression (L) or loss by RNAi expression (M) of klu (M) using kluReDDM.

(N-R) Images of representative midguts of esgReDDM at day 7 after temperature shift of the indicated genotypes. PH3 labelling (white) detected rare mitoses in the mutant genotypes.

Data information: Scale bar, 100µm for all images. Note that impaired cell renewal reduces the intestinal width. (F, G) **, P< 0.01; ****, P< 0.0001, one-way ANOVA with Bonferroni correction.