EMBO/sd-character-level-ner huggingface dataset:
Enhanced induction of p53‐dependent apoptosis in response to irradiation in LGR5hi cells compared to LGR5lo cells A mRNA gene expression analysis by microarray.Figure 6A mRNA gene expression analysis by microarray. The figure shows the number of differentially expressed genes (#dg) and the median absolute fold change (med. Abs. FC) for genes that are regulated in both LGR5hi cells and LGR5lo cells in response to IR (n = 4-5 samples per group). NIR, non‐irradiated; IR, irradiated. Wilcoxon test. B, C LGR5hi and LGR5lo cells were freshly isolated from 3‐month‐old LGR5‐GFPki mice 6 h after 12 Gy γ‐irradiation (IR) or from non‐irradiated (NIR) mice (n = 3 mice per group). (B) qPCR analysis of relative mRNA expression of apoptosis‐related genes compared to GAPDH. Mean values ± SEM are given. Unpaired two‐tailed Student's t‐test. B, C LGR5hi and LGR5lo cells were freshly isolated from 3‐month‐old LGR5‐GFPki mice 6 h after 12 Gy γ‐irradiation (IR) or from non‐irradiated (NIR) mice (n = 3 mice per group). (C) Representative Western blot analysis of cell lysates for the expression of phospho‐p53 and cleaved caspase‐3. D, E Three‐month‐old mTerc+/+ mice were γ‐irradiated with 12 Gy. Small intestinal tissue was collected at 3 h after IR (n = 3 mice per group). (D) Representative picture of TUNEL staining. Arrowheads and numbers indicate ISPC positions in the crypts. Scale bar: 20 μm. (E) Percentage of TUNEL‐positive ISPCs at indicated positions in basal crypts. Mean values ± SEM are given. Unpaired two‐tailed Student's t‐test. F-P Two‐month‐old LGR5‐GFPki, p53+/+ mice and LGR5‐GFPki, p53−/− mice were exposed to 12 Gy γ‐irradiation (n = 4 mice per group). Basal crypts were isolated at 24 h after IR. (F-H) Flow cytometry analysis of the survival rate of LGR5+ cells (F), LGR5hi and LGR5lo cells (G), and LGR5hi‐high and LGR5hi‐low cells (H) of irradiated mice (24 h after IR) compared to non‐irradiated mice (NIR). Mean values ± SEM are given. Unpaired two‐tailed Student's t‐test. (I-P) Representative FACS plots of LGR5+ cells in freshly isolated basal crypts from irradiated and non‐irradiated mice of indicated genotypes. Note the stronger rescue of survival rate of LGR5hi‐high than the LGR5hi‐low cells upon p53 deletion. NIR, non‐irradiated; IR, irradiated.
MSCs efficiently transfer mitochondria in a dose-dependent manner to T and B lymphocytes.Figure 1Schematic representation of the experimental design used for co-culture experiments. B Representative confocal microscopy of human PBMCs stained with DAPI (white arrows) co-cultured with previously labeled UC-MSC with MitoTracker-Green (top image) and specific antibody for human CD90 (bottom image). C Percentage of mitochondrial transfer (MitoT) on total human CD45+ cells after 24 hrs co-culture with MitoTracker-Green labeled bone marrow (BM), menstrual (Mens) or umbilical cord (UC)-MSCs (MSC:PBMC ratio 1:25) (n=3 biological replicates, with 3 different donors of BM, Mens and UC-MSCs). D FACS analysis of MitoT on T CD3+ (n=4), B CD19+ (n=4) and natural killer (NK) CD56+ (n=3) cells after 24 hrs co-culture with MitoGreen labeled UC-MSCs (ratio 1:25) (Biological replicates, with different donors of UC-MSCs). E FACS analysis of MitoT on CD3+CD4+ or CD3+CD8+ T cell populations after 24 hrs co-culture with labeled UC-MSCs (ratio 1:25) (n=3 biological replicates, with 3 different donors of UC-MSCs). F Representative FACS plots of MitoT on CD45+CD3+ T cells at increasing amounts of UC-MSC. G Percentage of MitoT on CD45+CD3+ human cells after 24 hrs co-culture with increasing ratios of UC-MSC:PBMCs (n=5 biological replicates, with 5 different UC-MSCs and 5 different PBMCs). H FACS analysis of MitoT on CD45+CD3+ T cells after short times of co-culture with UC-MSCs at two different temperatures, at a ratio 1:25 (n=3 biological replicates, with 3 different UC-MSCs and 3 different PBMCs). I Mean fluorescence intensity (MFI) of MitoTracker-Green+ UC-MSCs at baseline or after 72 hours PBMC co-culture. Representative FACS plot (left panel) and average of MFI (n=4 biological replicates, with 4 different UC-MSCs) (right panel). J Confocal microscopy of human PBMCs after co-culture with MitoTracker-Green stained UC-MSCs. Images showed MSC-mitochondria (green) inside the CD3+ T cells (red). K qPCR analysis of human β2-microglobulin (B2M), human specific mitochondrial (Mito) and mouse specific (β-actin) gene expression levels in UC-MSCs, mouse derived spleen cells or FACS sorted CD45+ mouse cells after 24 hrs co-culture with UC-MSCs (n=3 biological replicates). n/d = not detected. L In vivo experimental design for MitoT in Balb/c mice transplanted with MitoTracker Green labeled MSCs. M FACS analysis of in vivo MitoT from UC-MSCs to Balb/c mouse mesenteric lymph node and spleen cells, 24 hours after intraperitoneal transplantation of 5*106 MitoTracker-labeled MSCs or without cells (control) (n=1).
Fig.Figure 3 Fig. 3 Network derived from large-scale fractionation predicts 48 protein communities. Integration of experimental elution data, known functional associations and predicted interaction interfaces from homologous proteins allow the creation of a high-quality network with interconnected protein complexes (Appendix Figs. S8-S10). Here, known protein complexes are shown in blue and other physically associated proteins in grey, predicted interactions of complexes as grey lines and cross-links as red lines, cross-links between different subunits of a heteromultimeric complex are represented with red loops (see insert). Communities containing multiple complexes are highlighted with yellow; numbering and naming of complexes and communities is described in the legend of Appendix Fig. S9.
Differential behavior of inter-follicular and follicular stem cells A.Figure 4Schematic representation of cell sorting and single culture conditions. GFP+ bulge hair follicle stem cells (HF-SCs, CD34+ CD49fhigh) and GFP+ basal keratinocytes (b-KC, CD49fhigh) from ear skin of Co-mT/mG and DKO*-mT/mG mice were collected at day 10 after tamoxifen induction, sorted by FACS and cultured at low density (2000 cells/cm2) during 3-5 days as primary cultures. Time lapse imaging of 6 fields was performed during 48 hours, once small colonies of 5-10 cells were observed. Average number of live and dead (TOPRO-3+ Dapi+) primary keratinocytes derived from HF-SCGFP at different time points of time lapse capture during 48 hours. There is a slight increase over time of live HF-SCs from DKO* mice when compared to control cells, and no difference in average of dead cells. n = 4 fields from two independent experiments. Average number of live and dead (TOPRO-3+ DAPI+) primary keratinocytes derived from basal keratinocytesGFP show a significant increase in DKO* basal live keratinocytes at 12 hours, followed by a significant increase in cell death at 48 hours. Schematic representation of cell sorting and co-culture conditions. GFP+ and Tom+ bulge HF-SCs (CD34+ CD49fhigh) and GFP+ and Tom+ basal keratinocytes (CD49fhigh) from ear skin of Co-mT/mG and DKO*-mT/mG collected at day 10 after tamoxifen induction were sorted by FACS-sorting and co-cultured (same proportion of GFP and Tom cells) at low density (2000 cells/cm2) during 3-5 days. When small colonies of 5-10 cells were formed, time lapse imaging of 6 fields was performed during 48 hours. Average number of live primary keratinocytes from co-cultures of GFP+ with Tomato+ bulge HF-SCs at different time points of time lapse capture during 48 hours. Co-culture of HF-SCGFP with HF-SCTom from DKO*-mT/mG mice shows a significant increase of HF-SCTom, while the HF-SCTom from DKO*-mT/mG mice show a normal growth when they are co-culture with HF-SCGFP from control mice. Average number of live primary keratinocytes from co-cultures of GFP+ with Tomato+ basal keratinocytes at different time points of time lapse capture during 48 hours. Co-culture of b-KCGFP with b-KCTom from DKO*-mT/mG mice show significant increase of b-KCTom, while these b-KCTom from DKO*-mT/mG mice show a normal growth when they are co-culture with b-KCGFP from control mice. Average total number of live Tomato+ basal keratinocytes (from DKO* or control mice) at different time points cultured in the bottom part of trans-well system cultures with GFP+ basal keratinocytes (from DKO* or Control mice) cultured in the upper trans-well (Tw). n = 5 fields from three independent experiments. Tom+ basal keratinocytes significantly increased when these were cultured with mutant GFP+ basal keratinocytes in the trans-well in comparison with control GFP+ basal keratinocytes.
CryoEM structures of the SARS-CoV-2 S glycoprotein.Figure 3.Two orthogonal views of the closed SARS-CoV-2 S trimer cryoEM map. C. Atomic model of the closed SARS-CoV-2 S trimer in the same orientation as in panel A. D-E. Two orthogonal views of the partially open SARS-CoV-2 S trimer cryoEM map (one SB domain is open). F. Atomic model of the closed SARS-CoV-2 S trimer in the same orientation as in panel D. The glycans were omitted for clarity.
Figure 5(A) Effect of methylxanthines on wt lifespan. Average median lifespans ± SEM from all replicates indicated beneath representative curve. (B) Locomotory activity of 10-day-old gas-1 mutant nematodes treated with vehicle (DMSO) or methylxanthines. Values represent arbitrary units from 3 biological replicates with 4 technical replicates each, 35 worms per technical replicate. Error bars: mean ± SEM, **p value< 0.01, ***p value< 0.001, one-way ANOVA, Dunnett's post-hoc correction. (C) Effect of methylxanthines on the survival of gas-1 mutants. Average median lifespans ± SEM from all replicates indicated beneath representative curves. (D) Pharyngeal pumps/minute in young adult gas-1 mutant nematodes treated with vehicle or methylxanthines. Error bars: mean ± SEM, n=18. ****p value< 0.0001, one-way ANOVA, Dunnett's post-hoc correction. (E-F) Immunoblot analysis of (E) KIN-1 phospho-substrates in young adult gas-1 mutant nematodes treated with vehicle (DMSO) or methylxanthines (actin loading control). (E-F) Immunoblot analysis of (F) phospho-AAK-2 in young adult gas-1 mutant nematodes treated with vehicle (DMSO) or methylxanthines (actin loading control). (G) Left panel: FRAP in vehicle or PTX-treated gas-1 animals. Each point: mean recovery of 14 bleached regions (2 regions per animal, 7 animals per condition) ± SEM. Right panel: maximum (total) recovery per condition ± SEM. **p value< 0.01 unpaired t test. For all immunoblot panels: numbers correspond to band densitometries across biological replicates and for all lifespan curves: the legend shows Mean median lifespan ± SEM across all biological replicates. (H) Schematic drawing showing the critical biological processes underlying the lifespan extension of age-1; gas-1 mutant nematodes. OXPHOS deficient nematodes display decreased lifespan, diminished ATP production, fragmented mitochondrial network and a phosphoproteome signature associated with enhanced IIS signaling. As a result of IIS inhibition, AMPK and PKA drive a catabolic shift underlying lifespan extension of nematodes carrying OXPHOS defects. Supplementation of xanthine derivatives stimulates survival of mitochondrial mutant animals.
Figure 1A. GAPDH smFISH and IF analysis using anti-G3BP antibody (G3BP) and anti-Dcp1b antibody (DCP1) in A549 cells (RL-WT) or A549 RL-KO cells either mock transfected (mock) or transfected with poly(I:C) (PIC) for 5 hours. Scale bar 10 microns. B. Graph of the fraction of G3BP foci with different volumes in PIC treated RL-WT and RL-KO cells. C. Number of DCP1 foci per cell in PIC treated RL-WT and RL-KO cells relative to the number in mock treated cells. D. Average volume of individual Dcp1 foci in mock and PIC treated RL-WT and RL-KO cells.
Generating mice deficient in Filamin A editing.Figure 2(A) Scheme showing the FLNA wild-type (wt) allele, the targeting vector and the FLNAΔECS neo-allele. The loxP flanked PGK-neo cassette in the targeting vector replaced the editing complementary sequence (ECS) using homologous recombination, which was then deleted using Cre recombinase. Right, southern blotting analysis screened for positive clones shown. The positions of loxP sites, restriction enzyme (EcoRI) and Southern blotting probe are also indicated. (B) Sequencing electropherograms show the average FLNA editing levels in wt and FLNAΔECS tissues (Stomach, colon, heart, lung and tibial artery). Editing levels were checked in tissues from 3 independent mice and the value below the chromatogram depict the average value of 3 replicates. Data information: Arrows indicate the FLNA editing site. (C) FLNA mRNA expression levels measured by qPCR in wt and FLNAΔECS colon tissue showed no difference. Data information: data shown as mean ± SD from three independent experiments. ** P<0.05 (D) FLNA protein levels measured by western blotting are identical in wt and FLNAΔECS stomach tissue. Y-axis represents FLNA expression normalized to tubulin levels. Data information: data shown as mean ± SD from three independent experiments. ** P<0.05 (E) FLNA editing levels (%) in wt whole dorsal aorta, tunica media and tunica adventitia. Tunica media consisting of smooth muscle cells show the highest FLNA RNA editing. Data information: Arrows indicate the FLNA editing site. data shown as mean ± SD from three independent experiments. ** P<0.05
Lowering expression of α1-tubulin leads to fewer microtubules and aberrant sporozoite shapes A.Figure 4Complementation of α1-tubulin(-) parasites with different α1-tubulin constructs lacking introns (α1∆introns) or featuring only codon-modified exons (α1cm&∆introns). Scanning electron micrographs show the aberrant shape of the α1cm&∆introns-expressing sporozoites. See also Fig EV5. Scale bar: 1 μm. Fluorescence images showing the SiR-tubulin stained microtubules, nuclei (Hoechst) and cytoplasmic shape (mCherry) of the parasite lines. Note the small amount of DNA (black arrowhead) and the reduced level of SiR-tubulin fluorescence in the α1cm&∆introns-expressing sporozoites (lower graph). See also Fig S6. Quantitative data was derived from 96-120 sporozoites per line. Box plots represent 50% (boxes) and 95% (bars) of data with horizontal bar showing the median. Scale bar: 5 μm. **** indicates p<0.0001, Kruskal-Wallis-test. Note that α1cm&∆introns-expressing sporozoites do not express mCherry; their autofluorescence (auto-fluo) indicates an aberrant shape. qRT-PCR analysis of the three parasite lines at day 7 and 12 after infection. Error bars correspond to the standard deviation from two biological replicates. TEM reveals α1cm&∆introns-expressing sporozoites with smaller (A and B) and larger (C) diameters than wild type sporozoites. See also Fig S7. Scale bar: 0.2 µm. α1cm&∆introns-expressing sporozoites show a reduced number of subpellicular microtubules (green arrowheads) by TEM. Cartoon showing that sporozoites cannot form in the absence of microtubules, while lower numbers allowed formation of aberrantly shaped parasites. Note that subpellicular microtubules are exclusively polymerized from the apical polar ring.
Figure 5Lyzs-Cre and p38γ/δLyzs-KO mice were fed a ND or a HFD for 10 weeks. (A) Representative H&amp;amp;E and oil red stained liver sections (Scale Bar: 50μm) Lyzs-Cre and p38γ/δLyzs-KO mice were fed a ND or a HFD for 10 weeks. (B) liver triglycerides (n=5-10). Lyzs-Cre and p38γ/δLyzs-KO mice were fed a ND or a HFD for 10 weeks. (C) Plasma ALT at the end of the diet period. Lyzs-Cre and p38γ/δLyzs-KO mice were fed a ND or a HFD for 10 weeks. (D) Glucose tolerance measured at the end of the diet period. Blood glucose concentration was measured in mice given an intraperitoneal glucose injection (1g/kg) after overnight fasting (n=5-10). Lyzs-Cre and p38γ/δLyzs-KO mice were fed a ND or a HFD for 10 weeks. (E) Basal blood glucose in overnight-fasted ND and HFD-fed Lyzs-Cre and p38γ/δLyzs-KO mice (n=5-10). Data are means ± SEM. (n=5-10) *P<0.05; **P<0.01; ***P<0.001 refers to p38γ/δLyzs-KO versus Lyzs-Cre; ##P<0.01; ###P<0.001 refers to ND versus HFD (2-way ANOVA coupled to Bonferroni's post tests or Newman-Keuls post test for liver triglycerides).
Figure 3IE were isolated and binding to HBMEC and HDMEC was determined under flow conditions in the absence and presence of 5 µg/ml αICAM-1 or αCD36 antibody or 50 µg/ml rEPCR. Number of IE bound per mm2 EC surface was determined. IE were isolated and binding to HBMEC and HDMEC was determined under flow conditions in the absence and presence of 5 µg/ml αICAM-1 or αCD36 antibody or 50 µg/ml rEPCR. Number of IE bound per mm2 EC surface was determined. Using the same data, percentage inhibition by αICAM-1 antibody was calculated relatively to binding in the absence of antibody. Using the same data, percentage inhibition by rEPCR was calculated relatively to binding in the absence of inhibitor. Using the same data, percentage inhibition by αCD36 antibody was calculated relatively to binding in the absence of antibody.
Figure 5(A) A scheme of the hCMV uORF2 reporter mRNAs. Wild-type hCMV uORF2 (light blue) or its mutant (dark blue) was inserted at the end of the sfGFP ORF. Asterisks indicate stop codons. (B) Western blotting to detect the sfGFP levels at 6 hpf. Tubulin (Tub) was detected as a loading control. (C) qRT-PCR analysis of hCMV uORF2 reporter mRNA levels in wild-type embryos at 6 hpf relative to 2 hpf. Data information: bar charts show the average of three independent biological replicates. Error bars show SD. Individual data points are shown as dots. P-values were calculated using two-sided Student's t-test. (D) Poly(A) tail analysis of hCMV uORF2 reporter mRNAs at 2 and 6 hpf. The developmental stages are shown above as hpf. The lane labeled A0 shows the 3´ UTR fragment without a poly(A) tail. (E) Quantification of the PAT assay at 6 hpf in (D). (F) Scheme of human and zebrafish Znf598 proteins. The sgRNA target site is indicated by an arrowhead. Genome sequences of wild-type and znf598 mutant zebrafish are shown below. The predicted amino acid sequence of mutated Znf598 is shown in red. The asterisk indicates the premature stop codon. (G) qRT-PCR analysis of hCMV uORF2 reporter mRNA levels in MZznf598 embryos at 6 hpf relative to 2 hpf. (H) qRT-PCR analysis of hCMV uORF2 reporter mRNA levels in wild-type embryos, MZznf598 embryos and MZznf598 embryos injected with a mRNA encoding Myc-tagged full-length (full) or RING domain deleted (∆RING) Znf598. mRNA levels at 6 hpf relative to 2 hpf are shown. Data information: bar charts show the average of three independent biological replicates. Error bars show SD. Individual data points are shown as dots. P-values were calculated using two-sided Student's t-test. (I) Western blotting to detect the Myc-tagged Znf598 proteins at 6 hpf. Tubulin (Tub) was detected as a loading control. (J) Results of PACE in MZznf598 zebrafish embryos. Black circles show the relative stability of the reporter mRNAs (averages of two biological replicates) in MZznf598 embryos. Gray circles show the relative stability of reporter mRNAs in wild-type embryos The stability of a codon-tag reporter with a UGA stop codon was set to one. Error bars represent maximum and minimum data points. The relative effect of each codon on mRNA stability in wild-type embryos is shown as a color gradient from red (destabilizing) to blue (stabilizing).
a) Immunoelectron microscopy analysis of HeLa cells transfected with Flag–Dvl2 under nutrient-rich or starvation conditions.figf3(a) Immunoelectron microscopy analysis of HeLa cells transfected with Flag-Dvl2 under nutrient-rich or starvation conditions. The arrowheads indicate Flag-Dvl2. Scale bars, 1 μm (left); 250 nm (right). (b) Localization of endogenous Dvl proteins (red) and GFP-LC3 (green) in GFP-LC3 stable HeLa cells in nutrient-rich medium or under starvation (starv.) conditions with BFA1 for 4 h. The right panel shows the percentage co-localization between Dvls and GFP-LC3-positive dots. Scale bar, 5 μm. (c) Immunoelectron microscopy analysis of HeLa cells transfected with Flag-Dvl2 and GFP-LC3 under starvation conditions. Arrows indicate Flag-Dvl2; arrowheads indicate GFP-LC3. Scale bar, 250 nm. (d) Dvl2-LC3 interaction in vivo. IB, immunoblotting; IP, immunoprecipitation; S, starvation; WCL, whole-cell lysate; N, nutrient-rich. (e, f) Dvl2 interacts directly with LC3 (e) and GABARAP (f) but not with GATE-16 in vitro. GST pulldown assay using in vitro-translated Flag-Dvl2 and immobilized GST or the indicated GST fusions. Precipitated proteins were detected with anti-Flag antibody. Coomassie staining of GST and GST-LC3 protein is shown. (e, f) Dvl2 interacts directly with LC3 (e) and GABARAP (f) but not with GATE-16 in vitro. GST pulldown assay using in vitro-translated Flag-Dvl2 and immobilized GST or the indicated GST fusions. Precipitated proteins were detected with anti-Flag antibody. Coomassie staining of GST and GST-LC3 protein is shown. (g) The LC-3-binding motif is important for the Dvl2-LC3 interaction. GST-LC3-immobilized beads were incubated with the lysates from the cells expressing wild-type (WT) Dvl2 or the W444A, I447A mutant (MUT); the pulldowns were then subjected to anti-haemagglutinin (anti-HA) immunoblotting. Uncropped images of blots are shown in Supplementary Information, Fig. S8.
TMX1, TMX3 and NFAT1 control melanoma proliferation and invasion A Proliferation (48 h) of melanoma cell lines measured after transient knockdown of TMX1 or TMX3.Figure 6Proliferation (48 h) of melanoma cell lines measured after transient knockdown of TMX1 or TMX3. Proliferation (48 h) of melanoma cell lines with stable TMX1 knockdown (two clones). Proliferation (24 h) of melanoma cell lines after transient knockdown of NFAT1. Proliferation of WM3734 48 h after transient knockdown of TMX1 and treatment with NAC (100 µM) or mTEMPO (100 nM). Transwell migration (48 h) of WM3734 after transient knockdown of TMX1 or TMX3. Transwell migration (48 h) of WM3734 after transient knockdown of TMX1 and treatment with NAC (100 µM) or mTEMPO (1 µM). Transwell migration (48 h) of melanoma cell lines after preventing NFAT-calcineurin interaction with dipyridamole (40 µM) or inhibition of calcineurin with cyclosporine A (CsA, 2 µM). Invasion of WM3734 with stable TMX1 knockdown (two clones) through Matrigel for 96 h. Mouse xenograft model to assess tumor growth of WM3734 with stable TMX1 knockdown (TMX1 kds; two clones) in immunodeficient mice. (I) Tumor growth in the first 19 days after injection; (J) the corresponding tumor volume quantified after 19 days and (K) after 45 days.