EMBO/sd-character-level-ner huggingface dataset:
The essential role of miR-27a and miR-24 in ESC differentiation.Figure 3A. The genomic location of miR-23a~27a~24-2 cluster. B,C. Northern blot analysis of the expression of miR-23a, miR-27a and miR-24 during EB formation (B) and RA induced differentiation of V6.5 ESCs (C). The heatmap was representive of the results of quantitative analysis. D,E. The expression of primary miR-23a~27a~24-2 transcript during EB formation (D) and RA induced differentiation (E). Error bars indicate s.d. (n=3). D,E. The expression of primary miR-23a~27a~24-2 transcript during EB formation (D) and RA induced differentiation (E). Error bars indicate s.d. (n=3). F. Q-PCR analysis of miR-23a, miR-27a and miR-24 expression in ESCs (JM8A3) and a variety of C57BL/6J adult tissues. Data are shown as mean + s.d. (n=3). G. Relative protein levels of Oct4, Sox2 and Nanog in miRNA mimic transfected Dgcr8-/- ESCs and wild- type ESCs. H,I. Oct4, Nanog, Foxa2, Gata2 and Nestin immunofluorescence staining of mimic transfected Dgcr8-/- (H) and wild- type ESCs(I).
Pattern emergence upon spatial perturbation.Figure 2 Figure 2 Pattern emergence upon spatial perturbation. A. Even and odd Mathieu functions in an elliptical geometry. The 0.even mode shows the symmetry of the basal state of the system. Here no homogeneous steady state exists. Note the similarity between the 0th and the 2nd even mode. B. Simulations of Min pattern formation from an initially homogeneous state. Dimensions of the cells shown are 6.5x2x1 μm3, 9x2x1 μm3, and 9x5x1 μm3. All cells show an initial striped pattern, which persists in both cells of 9 μm length throughout the simulation period. C. Simulations analogous to the experiments shown in Fig. 1B, with the same cell dimensions as in Fig. 2B. The left-hand column depicts the spatially perturbed MinD attachment profile, showing gradients along the diagonal lines of the rectangles. With these attachment profiles, the Min distributions in the three cells quickly evolve into longitudinal, striped, and transverse patterns, respectively.
Fig.Figure 6His-Activin A-Atto647N was imaged free in solution with no cells (A) or with parental HEK293T cells (B), ACVR2A/B dKO cells (C), parental HEK293T cells pre-incubated with 1.5 μg/mL of rhACVR2A Fc and 15 μg/mL of rhACVR2B Fc (D), HOM1 cells Number and brightness (N&B) analysis was performed on raster-scanned images, and a representative experiment is shown. Panels i, Brightness (B) vs intensity 2D plots. Red rectangular box, pixels that contribute to brightness of free, dimeric His-Activin A-Atto647N in solution. Green rectangular box, pixels that contribute to His-Activin A-Atto647N clustered into putative tetramers. Blue and pink rectangular boxes, pixels that contribute to His-Activin A-Atto647N in higher order clusters. Panels ii, Selection map of the image captured reconstructed with the pixels in red, green, blue and pink rectangular boxes shown in the upper panels (i). Panels iii, B histograms, frequency distribution of the number of pixels across various brightness of His-Activin A-Atto647N of the selected image. Bav indicates the average brightness for each image. Panels iv, cumulative B histograms from a representative experiment with 4 independent regions of interest. His-Activin A-Atto647N was imaged free in solution with , HOM1 cells (E), HOM1 ACVR2A/B dKO cells (F) and HOM1 cells pre-incubated with 1.5 μg/mL of rhACVR2A Fc and 15 μg/mL of rhACVR2B Fc (G). Number and brightness (N&B) analysis was performed on raster-scanned images, and a representative experiment is shown. Panels i, Brightness (B) vs intensity 2D plots. Red rectangular box, pixels that contribute to brightness of free, dimeric His-Activin A-Atto647N in solution. Green rectangular box, pixels that contribute to His-Activin A-Atto647N clustered into putative tetramers. Blue and pink rectangular boxes, pixels that contribute to His-Activin A-Atto647N in higher order clusters. Panels ii, Selection map of the image captured reconstructed with the pixels in red, green, blue and pink rectangular boxes shown in the upper panels (i). Panels iii, B histograms, frequency distribution of the number of pixels across various brightness of His-Activin A-Atto647N of the selected image. Bav indicates the average brightness for each image. Panels iv, cumulative B histograms from a representative experiment with 4 independent regions of interest. H. Average brightness of His-Activin A-Atto647N in conditions (A-G). Means ± SEM are plotted. Each point represents one analyzed image, collected from 2-3 biological replicates. Data information: In H and I the p values are from one-way ANOVA with Tukey's post hoc correction. *, p<0.05; **, p<0.01; ***, p<0.001; ****, p<0.0001. I. Average brightness of His-Activin A-Atto647N in ICR-B169 and HSJD-DIPG-007 cells alone or with pre-incubation with 1.5 µg/mL of rhACVR2A Fc and 15 µg/mL of rhACVR2B Fc. Source data is in Fig. EV4B-F. Means ± SEM are plotted. Each point represents one analyzed image, collected from 2-4 biological replicates. Data information: In H and I the p values are from one-way ANOVA with Tukey's post hoc correction. *, p<0.05; **, p<0.01; ***, p<0.001; ****, p<0.0001.
x promotes cell proliferation.Figure 3A, Top: Scheme of the sites targeted by sgRNA in the generation of AGO1x mutant cell lines. Bottom: Western analysis of lysates from mutant MDA-MB-231 cells showing the depletion of AGO1x and the comparable total AGO1 levels across the cell lines. Three independently collected sets of cells were lysed and analysed in parallel. After calculating the levels of AGO1x and AGO1 relative to GAPDH, values corresponding to individual lanes were normalized to the average over the replicates of the control line. These numbers are indicated below the blots. The P-values of the t-tests comparing the expression AGO1x in mutant (W1A and W6A) and control (CTRL) lines were W1A-CTRL: 0.0008, W6A-CTRL: 0.0002, and for AGO1 W1A-CTRL: 0.1227, W6A-CTRL: 0.1173. B, Phase contrast images of control and mutant cell lines at 24 and 48 hr after plating equal numbers (10'000) of cells in individual wells of a six well plate. C, Impedance-based mean (+/- s.d.) cell indices at the indicated time points after seeding equal numbers of CTRL (n=6), W1A (n=5) and W6A (n=5) cells. From 6 hours on, there is a statistically significant difference between control and the two mutant cell lines (P < 0.001, two tailed t-test). D, Impedance-based mean (+/- s.d.) cell indices as a function of time for control (n=3), W1A (n=4) and W6A (n=3) cells grown in electronically monitored Boyden chambers. From 12 hours on, there is a statistical significant difference between control and the two mutants (P < 0.005, two tailed t-test). E, Mean (+/- s.d.) number of colonies obtained in a soft agar colony formation assay of control, W1A and W6A mutant cell lines (all n=4, P-value determined using an unpaired two-tailed t-test). F, Mean (+/- s.d.) number of spheres obtained from a sphere formation assay of control, W1A and W6A mutant cell lines (all n=8, P-values computed as in E). G, AGO1x staining of tissue sections from breast cancers, classified as low-proliferating or high-proliferating based on Ki-67 staining. The upper and lower panel represents two different tissue sections. H, Mean (+/- s.d.) percentage of AGO1x positive cells in breast tumors with low (n=7) and high (n=9) proliferation index, assessed by the expression level of Ki-67. P-value was determined using an unpaired two-tailed t-test.
Figure 6(A) Left: differentially accessible regions (DARs) between WT Treg cells and Tet2/3 DKO Treg cells. 2,270 regions are more accessible in WT Treg cells (upper panel); 1,201 regions are less accessible in WT Treg cells (lower panel). Middle: 5hmC distribution assessed by CMS-IP in the DARs shown on the left. Right: DNA methylation status assessed by WGBS in DARs shown on the left: 11,303 CpGs in 2,270 regions more accessible in WT Treg cells (upper panel); 4,785 CpGs in 1,201 regions less accessible in WT Treg cells (lower panel). CD4+ naïve T cells were used as a reference. P-values are calculated using analysis of variance and boxplots show the median and quartiles with the whiskers extending to the most extreme data point within 1.5 times the interquartile range. (B) Left: differentially accessible regions (DARs) between TGFβ iTregs and TGFβ + RA + VitC iTregs. 174 regions are more accessible in TGFβ + RA + VitC iTregs (upper panel); 144 regions are less accessible in TGFβ + RA + VitC iTregs (lower panel). Middle: 5hmC distribution assessed by CMS-IP in the DARs shown on the left. Right: DNA methylation status assessed by WGBS in DARs shown on the left: 1,123 CpGs in 174 regions more accessible in TGFβ + RA + VitC iTregs (upper panel); 678 CpGs in 144 regions less accessible in TGFβ + RA + VitC iTregs (lower panel). P-values are calculated using analysis of variance and boxplots show the median and quartiles with the whiskers extending to the most extreme data point within 1.5 times the interquartile range.
Figure 7Principle component analysis of RNAseq samples from biological replicates of PP-iNKT, spleen iNKT, and PP CD4+ cells Differential expression analysis comparing transcriptomes of spleen iNKT cells versus PP-iNKT cells. Heatmap of FPKM values from genes uniquely upregulated in PP-iNKT cells as compared to both spleen iNKT and PP CD4 cells. Red indicates higher expression and blue indicates lower expression. Spleen and PP cells from Vα14 TN mice were stained with the indicated antibodies, analyzed by flow cytometry and gated on CD1d (PBS57) tetramer+CD3+ cells. Quantification of data from D, n=11 mice per group. Mann-Whitney test. Error bars are SEM.
Figure 4A, B. DAPI staining of pollen in cycb1 mutants, including pollen configurations found in cycb1;1-/- cycb1;2+/- cycb1;3+/- cycb1;4-/- mutants (B). Scale bars: 5 μm. C. Quantification of DAPI-stained pollen configurations in different cycb1 mutant combinations, n = 420-616 pollen grains per genotype. Different letters indicate significant differences in the proportion of abnormal pollen (uni- and bicellular) in a Chi-squared test followed by the Marascuilo procedure to identify significant pairwise comparisons. D. Alexander staining of mature pollen indicating pollen viability. Scale bars: 5 μm. E. Quantification of Alexander-stained pollen viability, n = 403-498 pollen grains per genotype. Different letters indicate significant differences in the proportion of dead pollen in a Chi-squared test followed by the Marascuilo procedure to identify significant pairwise comparisons.
Time course of membrane alterations upon HCV infection.Figure 3(A) Images of mock-infected high-pressure-frozen and freeze-substituted Huh7.5 cells. (B-E) Micrographs of high pressure frozen freeze-substituted Huh7.5 cells harvested 16, 24, 36 and 48 h after infection with Jc1 (100 TCID50/cell). Yellow squares indicate the areas that are shown at higher magnification on the right of each subpanel. Note the time-dependent increase of complexity of HCV-induced membrane alterations. ER, endoplasmic reticulum; m, mitochondria; MVB, multi-vesicular bodies; LD, lipid droplet; DMV, double membrane vesicle; MMV, multi membrane vesicle; DMT, double membrane tubule (labeled with a black arrow).
Figure 6CENP-C crosslinks CENP-A nucleosomes, recruits other CCAN components and pre-conditions chromatin for formation of a functional centromere.
Fig.Figure 6(A) Validation of anti-phasic Rac1 expression in mouse (mRac1, red) and human enteroids (hRac1, black) by qRT-PCR. (B) Quantitative analysis of fluorescent intensity from SYTOX orange in Rac1-KD bHIEs. (C-D) Quantitative analysis of fluorescent intensity from SYTOX orange in control Rac1 f/f;CAGGCre-ERTM without tamoxifen (C) or Rac1 f/f;CAGGCre-ERTM with tamoxifen-treated (D) enteroids.
NOD2 deletion exacerbates diet-induced bacterial translocation, inflammation, and impairs insulin action in the liver.Figure 4A-C Quantification of bacterial DNA in the gonadal white adipose tissue (A) (WAT,n = 14-16 mice for all groups), mesenteric adipose tissue (B) (MAT,n = 10-15 mice for all groups), and liver (C) (n = 10-12 mice for all groups) of chow-fed and 16 week HFD-fed WT and NOD2−/− mice, *P = 0.0002. D Quantification of NOD2 transcripts in hepatocyte and non-hepatocyte cell populations of liver from chow-fed (n = 10) and HFD-fed (n = 6) WT mice, *P = 0.006. E Quantification of macrophage and inflammatory markers in liver of chow-fed (n = 5 in both genotypes) or HFD-fed (n = 11 in both genotypes) WT and NOD2−/− mice (n = 6 for IL-6 analysis), *P = 0.02, **P = 0.0001, #P = 0.02, and ##P = 0.002. F Representative liver IHC for the macrophage marker F4/80 (representative of n = 5 WT and n = 6 NOD2−/− mice). Scale bar equals 50 μm. G Quantification of hepatic triglycerides (TAG) in HFD-fed WT (n = 6) and NOD2−/− (n = 6) mice, *P = 0.0002. H Quantification of hepatic G6P transcript levels in chow-fed (n = 5) and HFD-fed (n = 11) WT and NOD2−/− mice, *P = 0.02. I, J Quantification (I) and immunoblots (J) of insulin-stimulated pFOXO1Ser256 in liver lysates after vena cava injection of insulin (0.5 IU/kg) in HFD-fed WT (n = 4) and NOD2−/− (n = 4) mice, *P = 0.0498. I, J Quantification (I) and immunoblots (J) of insulin-stimulated pFOXO1Ser256 in liver lysates after vena cava injection of insulin (0.5 IU/kg) in HFD-fed WT (n = 4) and NOD2−/− (n = 4) mice, *P = 0.0498. K Blood glucose and quantification of the AUC during a 120-min pyruvate tolerance test (PTT, 2.0 g/kg pyruvate, i.p.) in HFD-fed WT (n = 6) and NOD2−/− mice (n = 6), *P = 0.0005.
Figure 8A-C Quantification of FRET efficiency (percentage) in junctions of HUVEC expressing VE-cad FL (A, B) or VE-cad FL-TS (C). HUVEC were stimulated 4h with TNFα prior to adding IL-8 for 4 min, followed by flow with buffer alone (A) or together with PMNs (B, C). The connected data points represent FRET efficiency of same junctions compared before and after addition of flow/PMNs. D Quantification of FRET efficiency (percentage) in junctions of HUVEC expressing VE-cad FL or VE-cad FL-TS. Cells were either exposed to flow and PMNs or to flow alone, fixed using 4 % PFA and washed with PBS. FLIM measurements were performed at sites of transmigration (PMNs) or at junctions without PMNs (Flow). E Representative images of HUVEC expressing VE-cad FL at a site of PMN transmigration (left micrographs) or without PMN (right micrographs). Maximum intensity projection of a Z-stack of YPet fluorescence (VE-cadherin in green) and CellTracker DeepRed (PMN in red) is shown for the upper micrographs. The part of the PMN above the HUVEC monolayer is encircled in yellow and the part underneath in blue. The amplitude averaged lifetime of YPet per pixel (FLIM measurement) of the same cells is shown in the bottom micrographs. F HUVEC were grown on collagen-coated polyacrylamide gels of varying physiologic stiffness of 0.2 kPa and 20 kPa, and stimulated with TNFα for 17 h prior to adding HL60-derived neutrophils for 20 min. VE-cadherin was immunoprecipitated and precipitates as well as total cell lysates were analyzed by Western blotting for indicated antigens. Relative quantifications of pY731 blot signals adjusted to the amount of precipitated VE-cadherin are shown on the right
SPIN90 binds to the front side of Arp2/3 complex.Figure 3 A. Ribbon diagram of Arp2/3 complex with bound SPIN90 from the co-complex structure. SPIN90 is shown in pink with cylindrical helices. Conventional designations of "Front", "Back" and "Bottom" views are indicated in right panel  B. Ribbon diagram showing front side of Arp2/3 complex and bound SPIN90 with Fo-Fc electron density map contoured at 2.1 σ calculated without phases from SPIN90 (chain M)  C. Model of bovine Arp2/3 complex with bound WASP CA segments modeled based on crosslinking/mass-spectrometry, homology modeling, x-ray crystallography and FRE The complex is in an identical orientation to the right panel in (A)  D. Surface representation of model described in (C) showing the convention for front, back and bottom views of Arp2/3 complex
a) In silico prediction of miR-196 and IRGM mRNA interactions showed differences in binding within the seed region.figf1(a) In silico prediction of miR-196 and IRGM mRNA interactions showed differences in binding within the seed region. (b) IRGMC mRNA was significantly enriched in miR-196B complexes. Extracts of cells expressing FLAG-tagged-AGO1 and transfected with biotinylated miR-196B or control miR-20 as indicated and then with IRGMC or IRGMT plasmids were submitted to tandem affinity purification (immunoprecipitation with FLAG antibodies followed by affinity purification on streptavidin beads). IRGM mRNA variants were quantified using quantitative RT-PCR; results are presented as the ratio between miR-196B and miR-20 (non-relevant miRNA) pull-downs and the mean of three independent experiments ± standard deviation (s.d.). IP strep, immunoprecipitation straptavidine. (c) HEK293 cells (IRGMC/C) were transfected with either FLAG-tagged IRGMC or FLAG-tagged IRGMT plasmids and co-transfected with miR-196B. Immunoblotting with an IRGM antibody revealed the specificity of the downregulation effect mediated by the miRNA IRGM mRNA interaction. Quantification of the immunoblot signals are presented as IRGM expression relative to actin (mean of at least three independent experiments ± s.d.).
Figure 3(A) Experimental design to assess the integration of 4D-derived neurons. (B) Fluorescence pictures (left) of immunolabeled GFP+RFP- (4D-) and GFP+RFP+ (4D+) apical dendrites of superficial granule neurons and quantifications (middle and right) of spine density and total dendritic length (box and whiskers) and 3D-Sholl (line graph profile) of apical dendrites starting from the soma (drawings) as the mean number of intersections at 10 μm intervals. (C) 3D reconstruction of multi-channel confocal stacks acquired from a RFP+ apical dendrite of a granule cell showing co-localization with the pre- and post-synaptic markers VIAAT (granule neuron) and gephyrin (mitral cell), respectively (magnification shown in insets). (D) Anti-RFP immunogold labeling of a cell in the superficial granule cell layer of a 4D+ mouse. Inset shows a representative RFP+ synapse (out of >10 analyzed from 4D+ mice). Current clamp recordings showing examples of spontaneous barrages of synaptic potentials (E), Inset in E (4D+ cell) is magnified (bottom). Current clamp recordings showing examples of repetitive spiking in response to depolarizing current steps (F-G) of 4D- (top) and 4D+ (bottom) mice. Note in G that the lag preventing spike initiation is longer at lower currents (green) and shorter at higher (black), suggesting the presence of an A-type K current typical of granule cells in 4D- vs. 4D+. (H) Box and whiskers plots representing electrophysiological properties of neurons derived from 4D- and 4D+ NSC (black and red, respectively) including from left to right: spike number, resting membrane potential (Vrest), input resistance (Rin) and lag to spiking of the recorded superficial granule cells (see Fig. EV3 for additional parameters).
Characterization of renal SIRT7 expression in C57BL6 mice.Figure 4A, B Sirt7 mRNA (A) and protein (B) abundance in microdissected renal tubules normalized to Rpl26 or Ponceau, respectively. Enrichment for each segment was previously validated by immunoblotting against segment-specific markers as indicated in material and methods (Spirli et al, 2019). C Immunolocalization of SIRT7 and ATPase in the kidneys of C57BL6 mice with metabolic acidosis (SIRT7, red; ATPase, green; DAPI, Blue). Scale bar: 20 μm. Controls of the specificity of antibodies using kidney of SIRT7-deficient mice are provided in Appendix Fig S4. D KCC4 and SIRT7 protein levels in kidneys from control mice or mice with metabolic acidosis. Values are the mean ± SEM.
Figure 4(A and B) View of the first normal modes of monomer A and monomer B in the human cGAS-DNA 2:2 complex in the resting and palmitoylation states during the simulation. The colors of monomer A and monomer B in (A) and (B) are consistent with those in fig. 3, (D) and (E). PC1: the first principal components in PCA. (C) Cross-correlated motions of the Cα atoms of the WT and palmitoylated cGAS proteins. (D) Immunoprecipitation (with an anti-Flag antibody) and immunoblot analysis of HEK293T cells transfected with plasmids encoding Flag-cGAS, HA-cGAS, and Myc-ZDHHC18 (WT or CS mutant) for 24 h. Data information: WCL: whole-cell lysate; IP: immunoprecipitation; Data are representative of at least two independent experiments. (E) Immunoprecipitation (with an anti-Flag antibody) and immunoblot analysis of HEK293T cells transfected with the indicated plasmids followed by the addition of DMSO, 2-BP (50 μM) or palmitic acid (100 μM). Data information: WCL: whole-cell lysate; IP: immunoprecipitation; 2-BP: 2-bromopalmitate. Data are representative of at least two independent experiments. (F) FRET assay of HeLa cells transfected with the indicated plasmids with the addition of DMSO, 2-BP (50 μM) or palmitic acid (100 μM). Scale bar: 8 μm. (G) FRET efficiency of cGAS-GFP and cGAS-mCherry in (F). Data information: 2-BP: 2-bromopalmitate. Data are representative of at least two independent experiments. Mean ± SEM from triplicates of technical replicates, unpaired t test; *, P˂0.01. ***, P˂0.001 (G (H) Immunoprecipitation (with an anti-Flag antibody) and immunoblot analysis of HEK293T cells transfected with Flag-cGAS and HA-cGAS (WT, C405S, C409S and C474S) expression plasmids. Data information: WCL: whole-cell lysate; IP: immunoprecipitation Data are representative of at least two independent experiments. (I) FRET assay of HeLa cells transfected with the indicated plasmids. Scale bar: 8 μm. (J) FRET efficiency of cGAS-GFP and cGAS-mCherry in (I). Data information: Data are representative of at least two independent experiments. Mean ± SEM from triplicates of technical replicates, unpaired t test; *, P˂0.01. ***, P˂0.001 J).
Figure 5(A) Effect of Tet2 and Tet3 single deficiency on osteoclastogenesis. TRAP-stained cells (left panel) and the number of TRAP-positive cells with more than three nuclei (right). Scale bar, 100 μm. (n = 6 (control and Tet2Rank-/-) and n = 3 (Tet3aRank-/-) biological replicates (B) μCT analysis of the femurs of 10-week-old control (n = 6), Tet2Rank-/- (n = 5), Tet3Rank-/- (n = 5), Tet2Rank-/-; Tet3aRank+/- (n = 6), Tet2Rank+/-; Tet3aRank-/- (n = 6) and Tet2Rank-/-; Tet3aRank-/- (n = 4) male mice (axial view of the metaphyseal region). (C) Histological analysis of the proximal tibias of 10-week-old control, Tet2Rank-/-, Tet3Rank-/-, Tet2Rank-/-; Tet3aRank+/-, Tet2Rank+/-; Tet3aRank-/- and Tet2Rank-/-; Tet3aRank-/- male mice (TRAP staining [red, osteoclasts]). (D) Parameters for osteoclastic bone resorption during bone morphometric analysis of 10-week-old control (n = 10), Tet2Rank-/- (n = 5), Tet3Rank-/- (n = 6), Tet2Rank-/-; Tet3aRank+/- (n = 6), Tet2Rank+/-; Tet3aRank-/- (n = 6) and Tet2Rank-/-; Tet3aRank-/- (n = 4) male mice. (E) Effect of Tet2 and Tet3 double deficiency on osteoclastogenesis. TRAP-stained cells (left panel) and the number of TRAP-positive cells with more than three nuclei (right). (n = 6 (control), n = 10 (Tet2Rank-/-; Tet3aRank+/-), n = 3 (Tet2Rank+/-; Tet3aRank-/-) and n = 4 (Tet2Rank-/-; Tet3aRank-/-) biological replicates