EMBO/sd-character-level-ner huggingface dataset:
In vitro reconstitution of branched- and linear- actin networks assembly from purified actins.Figure 4A (Top) Snapshots of actin networks assembled from three different actins sources: Act_Sc, Act_N2 and Act_Ca. (Bottom left) Quantification of actin fluorescence on beads. Data are presented as mean +/- SD (n = 52 for Act_Sc branched, n = 70 for Act_N2 branched, n = 81 for Act_Ca branched, n = 32 for Act_Sc linear, n = 34 for Act_N2 linear and n = 47 for Act_Ca linear). **P<0.01, ***P<0.001 (Kruskal-Wallis test, with multiple comparisons). (Bottom right) In vitro actin network deviation indexes. B (Left) Snapshots of representative actin networks assembled in the presence of 600 nM Alexa-488-labeled ADF/cofilin (replacement of unlabeled ADF/cofilin). (Right) Quantification of ADF/cofilin's fluorescence intensity relative to actin. Data are presented as mean +/- SD (n = 34 for Act_Sc branched, n = 28 for Act_N2 branched, n = 43 for Act_Ca branched, n = 24 for Act_Sc linear, n = 15 for Act_N2 linear and n = 27 for Act_Ca linear). ***P<0.001 (Kruskal-Wallis test, with multiple comparisons). C (Left) Snapshots of representative actin networks assembled in the presence 1 µM Alexa 488-tropomyosin. (Right) Quantification of tropomyosin's fluorescence intensity relative to actin. Data are presented as mean +/- SD (n = 45 for Act_Sc branched, n = 63 for Act_N2 branched, n = 57 for Act_Ca branched, n = 22 for Act_Sc linear, n = 32 for Act_N2 linear and n = 27 for Act_Ca linear). *P<0.05, ***P<0.001 (Brown-Forsythe and Welch ANOVA tests, with Dunnett's T3 multiple comparisons tests).
Figure 2(A) Total to oxidized glutathione ratio (GSH/GSSG) in mock infected or HIV-1 infected CD4+ T-cells (7 dpi, n=4 biological replicates; mean±SEM). representative picture (B) of ROS content in HIV-1 infected or mock infected primary CD4+ T-cells at 7dpi (productive infection) and 14 dpi (latent infection). ROS content was measured by immunofluorescence, HIV-1 DNA+ cells were identified by FISH quantification of ROS content in HIV-1 infected or mock infected primary CD4+ T-cells at 7dpi (productive infection) and 14 dpi (latent infection). ROS content was measured by immunofluorescence, HIV-1 DNA+ cells were identified by FISH (D). The black dash in (C) indicates the median. Box plots in (D) depict median and 25-75 percentile, while whiskers extend from min to max. n= number of cells. Scale bar = 2μm. (E) Subcellular localization of Nrf2 in CD4+ T-cells infected with HIV-1 or mock infected (3dpi) as analyzed by biochemical fractionation and western blot; Whole Cell Extract (WCE), Cytoplasm (Cyt), Nucleoplasm (Nuc). (F) Nuclear localization (left) and content (right) of Nrf2 in HIV-1 RNA+ and HIV-1 RNA- cells (7dpi) as measured by combining immunofluorescence and HIV-1 RNA FISH. Black dash indicates median, n= number of cells. Scale bar = 2μm Time course of the mRNA levels of main downstream antioxidant targets of Nrf2 during the transition from productive (3-9 dpi) to latent (14 dpi) infection, as measured by qPCR Data in (G) are depicted as mean±SEM of 3 biological replicates. raw data were first normalized using 18S as housekeeping control and then expressed as Log2 fold mRNA expression in infected vs mock infected cells Trx= thioredoxin; NQO1= NAD(P) H [quinone] dehydrogenase 1; HMOX-1= heme oxygenase 1; G6PD= glucose-6-phosphate dehydrogenase; GCLC= glutamate-cysteine ligase ; TrxR1= thioredoxin reductase 1. Time course of the protein levels of main downstream antioxidant targets of Nrf2 during the transition from productive (3-9 dpi) to latent (14 dpi) infection, as measured by western blot GCLC and HMOX-1 in (H) were probed upon membrane stripping. (I) Relative mRNA expression of Nrf2 downstream antioxidant targets in J-Lat 9.2 cells left untreated (latent) or treated for 24h with 10µM TPA (reactivated). Data are depicted as mean±SEM of 3 biological replicates. raw data were first normalized using 18S as housekeeping control and then expressed as Log2 fold mRNA expression in TPA-reactivated vs latent cells Trx= thioredoxin; NQO1= NAD(P) H [quinone] dehydrogenase 1; HMOX-1= heme oxygenase 1; G6PD= glucose-6-phosphate dehydrogenase; GCLC=
Amnion single cell transcriptomeA.Figure 5A. In Brachyury (T)-Cre; mTmG embryos, mesoderm-derived cells express membrane eGFP (mG, green) and the other cells membrane tdTomato (mT, grey). B. Scheme of amnion isolation. C. Uniform Manifold Approximation and Projection (UMAP) (Left, color-coded) and RNA velocity (Right, black arrows show streaming directions from unspliced to spliced RNA) of amnion cells where 6 unsupervised clusters were identified. D. UMAP of eGFP, Krt8, Cdh1 and Cdh2. E. EB K8-eYFP embryo stained for eYFP (yellow) and E-cadherin (white) (n=7) (Top). Dashed white area zoom on an amnion region which highlights the two cell layers (Bottom). F. Dot plot of specific gene expression in amnion clusters. G. Immunostaining for Cdx2 (n=20). H. Immunostaining for Nrp1 (n=8) and Col1a1 (n=7), primarily expressed in Amnion Mesenchyme. I. Immunostaining for Epcam (n=12), Cldn7 (n=7), and Lrp2 (n=6), primarily expressed in Amnion Ectoderm. J-K. Sections of C57BL/6J wild-type and KtyII-/- E7.5 embryos immunostained for Nrp1 (K) (n=8 and 2, respectively), and Lrp2 (L) (n=3 and 2).
Figure 2Metabolite enrichment analysis in latently infected cells as compared to their uninfected counterparts (A) and cells reactivated from latency and compared to latently infected cells (B) Jurkat T-cells cells, as compared to their uninfected counterparts. The top enriched pathways were ordered according to p values obtained by Q statistics for metabolic datasets performed with Globaltest (MetaboAnalyst) (Xia et al, 2009). Heatmaps of glycolytic metabolites in latently infected Jurkat T-cells as compared to their uninfected counterparts or Jurkat T-cells with HIV-1 reactivated by TNF as compared to latently infected cells. Data are displayed as Log2 fold change expression. Adjusted P values (q values) were calculated by the Benjamini-Hochberg false discovery rate. Relative ratios of NADH/NAD+ (D) and ATP/ADP (E) in latently infected and reactivated cells. Data were normalized using the matching uninfected control.
Figure 2Relative quantification of the differentiation markers LOR and IVL in HEKn transfected with uc.291 si-RNA (1) and (2) or SCR control siRNA and collected after 3 days of differentiation. Data shown represent the mean ± s.d.; n=4 different human donors; ** p<0.01, Student's t-test. WB showing ΔNp63α, Klf4, K10 and Loricrin (Lor) protein levels in HEKn transfected cells as described in (A). β-actin was shown as loading control. One representative experiment of three is shown. IF staining of late differentiation markers Loricrin and Filaggrin in uc.291-depleted, si-uc.291(1) and si-uc.291(2), organotypic human epidermis compared to scramble control one (SCR); haematoxylin/eosin (H&E) staining shows the organotypic epidermis structure (left panels); scale bars: 50μm. The magnified boxes show expression of Loricrin and Filaggrin in the granular layer; scale bars: 10μm. Dotted lines represent the insert border. One representative experiment of three is shown. IF staining and confocal analysis of p63 in the organotypic human epidermis obtained as described in (C). Dotted lines represent the insert border. One representative experiment of three is shown. Scale bars: 50μm Box plot showing quantification of p63 positive nuclei on total nuclei. Each central line represents the median; each box represents Q1 (quartile 1, below the median) and Q3 (quartile 3, above the median); the whiskers represents minimum (Q1-1,5*IQR) and maximum (Q3+1,5*IQR). IQR=interquartile range; 16, 15, and 19 images were analysed for SCR, si-uc.291 (1), and si-uc.291 (2) respectively; *p<0.05; ** p<0.01, Student's t-test. WB showing cyclin-dependent kinase inhibitor 1B (p27Kip1) and ∆Np63α protein levels in HEKn cells transfected with uc.291 si-RNA (2) or SCR control siRNA and collected in proliferating (0 day) or differentiating (1, 2, 3 days of differentiation, DD) conditions. β-actin was used as loading control. The numbers below the bands represent the densitometric analysis normalized for the loading control. One representative experiment of three is shown. Schematic representation of the expression of selected markers in organotypic human epidermis upon uc.291 depletion.
Figure 6A Time lapse images of E. coli fabA(Ts) cells expressing FOF1 a-mNG. For corresponding controls, see Appendix Fig S10. Cells were grown at 30°C and shifted from 30°C to non-permissive 37°C. See Movie EV2 for full time lapse. B Images of E. coli fabA(Ts) co-expressing WALP23-mScarlet-I and FOF1 a-mNG grown at permissive 30°C or shifted to non-permissive 40°C. For fluorescence intensity correlations, see Appendix Fig S8D. For corresponding images of E. coli fabA(Ts) cells co-expressing inner membrane marker FOF1 a-mNG and outer membrane marker OmpA-mCherry, see Fig EV5. C Images of FOF1 a-mCherry-expressing E. coli fabA(Ts) grown at 30°C or shifted to non-permissive 37°C and stained with Laurdan. For fluorescence intensity correlations, see Appendix Fig S8E. D Growth behaviour of FOF1 a-mNG expressing E. coli fabA(Ts) after shift from 30°C to non-permissive 37°C and upon recovery through exogenous supplementation with UFA oleate (cis-Δ9-C18:1). The corresponding reversible segregation of FOF1 a-mNG is shown above the growth curve. For further controls, see Appendix Fig S11A. For a comparable fluorescence time lapse analysis of mNG-labelled glucose permease PtsG, see Appendix Fig S12. E Fatty acid composition of E. coli fabA(Ts) cells (same cell batch as in D) upon growth at 30°C, upon depletion of UFA by incubation at 37°C for 120 min and upon recovery by oleate supplementation for additional 120 min. For detailed analyses, see Appendix Fig S11B-D.
Figure 3(A) The cellular stiffness of ALDH- and ALDH+ 4T1 or MCF-7 cells sorted by flow cytometry were determined by AFM. n = 100. Data information: Mann-Whitney test The data represent mean ± SD. (B) The stiff or soft cells were isolated from ALDH- or ALDH+ 4T1 cells by microfluidic chip. Then, these cells were injected into the mammary fat pads of NSG or BALB/c mice mice (100 cells/mouse). Twelve weeks later, the tumor formation was recorded. n = 10. The 100 stiff ALDH+ or soft ALDH- 4T1 cells were injected into the mammary fat pads of NSG mice. Eight weeks later, the lung sections were H&E stained. The lung metastatic mice were counted (C) Scale bar, 0.5 mm. n = 5 mice with metastatic tumor. The 100 stiff ALDH+ or soft ALDH- 4T1 cells were injected into the mammary fat pads of NSG mice. Eight weeks later, the lung sections were H&E stained. metastasis index was calculated (D). Scale bar, 0.5 mm. n = 5 mice with metastatic tumor. Data information: two-tailed Paired Student's t test data represent mean ± SD. (E) The side-population (SP) or non-SP (NSP) from 4T1 or MCF-7 cells were sorted by flow cytometry, and then the cellular stiffness of those cells were measured by AFM. n = 100. Data information: Mann-Whitney test The data represent mean ± SD. (F) The 100 soft NSP or stiff SP- 4T1, MCF-7, B16 or MP-1 cells were injected into the mammary fat pads (4T1and MCF-7) or subcutaneous tissue (B16 and MP-1) of NSG mice. The tumor formation in the lungs was recorded. The 100 soft NSP or stiff SP- 4T1 or MP-1 cells were injected into the mammary fat pads (for 4T1) of NSG mice or by tail vein into NSG mice (for MP-1). The tumor formation in the lungs was recorded (G). Scale bar, 0.5 mm. n = 5 mice with metastatic tumor. The 100 soft NSP or stiff SP- 4T1 or MP-1 cells were injected into the mammary fat pads (for 4T1) of NSG mice or by tail vein into NSG mice (for MP-1). Six to eight weeks later, mice were sacrificed and the lung sections were H&E stained. The metastatic micronodules in the lung were counted and the metastasis index was calculated (H). Scale bar, 0.5 mm. n = 5 mice with metastatic tumor. Data information two-tailed Paired Student's t test The data represent mean ± SD.
HTT Is Modified by Both SUMO-1 and SUMO-2.Figure 3(A) HeLa cells transfected with His-tagged HTTex1p (46Q) or 46QP-K6,9,15R (3R) along with GFP-SUMO-1 or GFP-SUMO-2, lysed under denaturing conditions, and nickel purified (Ni-NTA). Unmodified HTT-46Q is indicated by the arrow and SUMO-modified HTT by the boxed region. The lysine mutant (3R) serves as a negative control. Ni-NTA represents nickel-purified His-tagged HTT, and WC TCA represents 10% of the whole-cell lysate expression of HTT and myc-actin (transfection control). HTT is modified by SUMO-1 (left) and SUMO-2 (right). (B) SUMO isopeptidases (SENP1, SENP2, SENP3, SENP5, and SENP6) modulate SUMO-modified HTT when overexpressed together with HTT (46QP-H4 or 3R) and SUMO-1 (GFP-SUMO-1). SENP1, SENP2, and SENP6 decrease HTT SUMOylation. Graph depicts quantitation of western blot using the Odyssey Infrared Imaging System (LI-COR) to calculate the ratio of HTT purified versus the HTT modified by SUMO multiplied by 100. (C) Titration of SUMO-1. Denaturing nickel purification of HTTex1p (46QP-H4) following transfection with decreasing amounts of SUMO-1 reduces the amount of SUMO-modified HTT to undetectable levels. The Ni-NTA blot displayed in the gray scale shows purified HTTex1 and SUMO-modified HTTex1 using HTT antibody. Note that 0.5 μg of SUMO-1 (¼ the amount of SUMO-1 cDNA ) was used for identifying the SUMO-1 E3 ligase for HTTex1p. Graph depicts quantitation of western blot using the Odyssey Infrared Imaging System to calculate the ratio of HTT purified versus the HTT modified by SUMO multiplied by 100. Note that all experiments were performed in triplicate, and a representative figure is shown.
Figure 4Column graphs representing the percentage of CD4 T cells from NSCLC patients or age-matched healthy donors as represented in the graph, expressing the indicated cytokines after T cell stimulation with anti-CD3/anti-CD28 antibodies. Data on total CD4 (left graph), CD28+ subsets (center graph) and CD28negative subsets (right graph) are shown. Error bars correspond to standard deviations and bars represent means from 5 independent biological replicates (patients). representing CD4 T cells expressing only one cytokine representing CD4 T cells expressing only cytokine, two CD4 T cells expressing only three cytokines simultaneously
Temporal resolution of protein content in seedlings during the dark-to-light transition.Figure 7A Immuno-blots showing the levels of PAP8, PAP5, Histone H3 (H3), Rubisco (RbcL), and HY5 ; D, Dark; Ph, photomorphogenic growth conditions from germination on. *, HY5 is detected as a modified form (+ 6 kDa); the arrows indicate the two different post-translational modified forms of HY5 for which accumulation occurs upon light exposure. B Relative protein contents normalized to histone H3 during the transition from skotomorphogenesis to photomorphogenesis, we propose a phase of photo-initiation corresponding to the 5 minutes of light allowing PAP8 and PAP5 to rise to nearly 100% of their maximum. The first macroscopic signs of greening are indicated around 3 hours while the photosynthetic apparatus bursts at 8 hours to be fully accumulated at 48 hours.
Figure 3Immunofluorescence analysis of CD20 and IGF2R expression (in red) in the TA muscles of three-month-old C57Bl6/J and mdx mice. Scale bars=75μm. (a). IGF2R (green) and sarcomeric actin (magenta) expression in the VM muscles of two DMD patients. Scale bars=25μm. (b). Representative WB analysis of CD20 and β-actin (c) expression in the TA and VM muscles of C57Bl6/J and mdx mice (n=10 mice per group). Densitometry analysis of WB data are expressed as the CD20/β-actin ratio, in arbitrary units in the lower panels One-way ANOVA. ****p<0.0001. Representative WB analysis of pSer+pThr and GADPH (d) expression in the TA and VM muscles of C57Bl6/J and mdx mice (n=10 mice per group). Densitometry analysis of WB data are expressed as the pSer+pThr/CD20 ratio in arbitrary units in the lower panels One-way ANOVA. ****p<0.0001. (e) Representative WB analysis of IGF2R and GAPDH expression in total protein lysates of the human (healthy and DMD) and murine (C57Bl6/J and mdx) TA and VM muscles (n=2 per healthy and DMD; n=10 mice per group); quantifications are expressed as the IGF2R/GAPDH ratio in arbitrary units. (f) Representative WB analysis of IGF2R expression in sarcolemma protein lysates of the TA and VM muscles of healthy, DMD, C57Bl6/J and mdx (n=2 per healthy and DMD; n=10 mice per group). Densitometry analysis of IGF2R expression, results are shown as a fold increase, as indicated in the lower panel in arbitrary units. One-way ANOVA. *p<0.05; ****p<0.0001. All values are expressed as the mean±SEM.
Figure 6A. Complex structures of the mouse chimeric proteasome (left) and the hβ5i-V31M/hβ6 mutant (right) with PR-924 visualize the kinked (k) orientation. The 2FO-FC electron density maps for the ligands bound to Thr1 of the chimeric β5 active sites are depicted according to Fig 5A. Steric hindrance of Met31 (green) with the bulky 3-methyl-1H-indene cap is indicated by black double arrows. B. Superposition of inhibitors bound to the mouse chimeric proteasome (left) and the hβ5i-V31M/hβ6 mutant (right). Compounds display the same binding modes (kinked (k) or linear (l)) to the mβ5i/mβ6 active site as to the corresponding human β5i/β6 subunit (Fig. 5) and to the hβ5i-V31M/hβ6 mutant. Note that the inhibitors are rotated by about 45° compared to panel A. C. Superposition of chimeric mammalian and WT yeast β5 active sites bound to the P3-d-Ala compounds PR-924 and 18. The nucleophilic Thr1 is depicted as well as the S1 pocket forming residues 31 and 45. Residue 31 impacts on the position of the 3-methyl-1H-indene cap of PR-924 and 18. Due to steric hindrance, the N-cap is shifted in the mouse and in the hβ5i-V31M/hβ6 chimera compared to the hβ5i/hβ6 mutant proteasome crystal structures (black double arrow). Note that the structures are rotated by 90° compared to panel B.
Figure 2(A) Western blot analyses of the knockout efficiency of Cdc20 in Cdc20f/f and Sp7-Cre;Cdc20f/f BMSCs. (B, C) The ALP staining (B) and ALP quantification (C) of Cdc20f/f and Sp7-Cre;Cdc20f/f BMSCs after 7 days osteogenic differentiation. (n=6) (D) The efficiency of CDC20 knockdown determined by western blot in hBMSCs. (E, F) The ALP staining (E) and ALP quantification (F) of NC and CDC20sh hBMSCs after 7 days osteogenic differentiation. (n=5) (G) Schematic representation of the CDC20 truncated mutant fragments. The boundaries of its WD40 domain (amino acids 171-471) were shown. (H, I) The ALP staining (H) and ALP quantification (I) of Vector and truncated fragments overexpression of CDC20 in CDC20sh hBMSCs after 7 days osteogenic differentiation. (n=5) (J, K) The H&E staining (J) and Masson's trichrome (K) of the histological sections from implanted NC and CDC20sh hBMSCs scaffold hybrids. Scale bar, 100 μm.
Tamoxifen treatment decreases HIF-1A levels and proliferation and increases apoptosis in Suit-2 pancreatic cancer cells.Figure 7(A, E, F) Immunofluorescence images of control and tamoxifen treated suit-2 cells, scale bar is 20 μm. Panel A represents HIF-1A staining in hypoxia and non-hypoxia conditions, panels E and F show Ki67 and Cc3 staining as markers of proliferation and caspase-mediated apoptosis, respectively. Panel E: Red - F-actin, green - Ki67, blue - nuclei. Tamoxifen negatively regulates HIF-1A in hypoxia and non-hypoxia conditions. (B,C,D) Quantification of immunofluorescence staining in panels A, E, F. For quantification, eight fields of view (n>50 cells) per condition. Histogram bars represent mean ± s.e.m.; ***P<0.001, t-test. All panels include data collected during 3 independent experiments
K-Urm1 complex at 3.Figure 3Interaction analysis of WT and mutated CtUba4 with GST-CtUrm1 by GST pull-down in the presence of 1mM ATP. CtUba4C202K covalently linked to GST-CtUrm1 is marked by an asterisk. Comparison of the structures of the CtUba4C202K-CtUrm1 complex and apo CtUba4 focusing on the arrangement of Urm1 and RHD in relation to the AD dimers. The symmetry operator of the crystallographic two-fold axis is indicated. Overall structure of the CtUba4C202K-CtUrm1 complex. The AD dimer is shown as surface representation, the Uba4 linker (light blue), crossover loop (light brown) and Urm1 (salmon) are shown as cartoon. The C-terminus of Urm1 (dark red), Lys202 (green) and the formed covalent peptide bond between the two components (red) are highlighted. The ATP molecule (transparent red) is positioned in the nucleotide binding site based on the ATP-bound EcMoeB-MoaD complex (PDB ID: 1JWA). Uba4 and Urm1 molecules from the adjacent asymmetric unit which form the dimer are shown in transparent grey. Close up view of the CtUba4 active site and the CtUrm1 C-terminus (top). A superposition of the linker from apo CtUba4 structure is shown (bottom). Crucial residues are shown as ball-and-stick model and the tentative positions of Cys202 Cys305 are indicated.
Figure 2Representative images of PCNA-stained lung sections from KrasWT, KrasG12D and KrasG12D:Adam17ex/ex mice at 6 weeks post Ad-Cre (for KrasG12D) or vehicle (for KrasWT) inhalation. Scale bar, 100μm. Quantification of PCNA-positive cells/high power field (HPF) in the indicated mouse lungs (n = 6 per genotype). qPCR expression analyses of cell cycle regulation genes (normalized against 18SrRNA) in lungs from the indicated mice (n = 6 per genotype). * Representative immunoblots of KrasG12D and KrasG12D:Adam17ex/ex lung lysates with the indicated antibodies. Each lane is an individual mouse sample. Semi-quantitative densitometry of Myc protein levels (relative to Actin) in lung lysates from (D). n = 6 per genotype. Representative images (F) and quantification of CD45-positive cells/HPF (G) in CD45-stained lung sections from the indicated genotypes at 6 weeks post inhalations. In (F), scale bar, 100μm. In (G), n = 6 per genotype.
Figure 5(A) Primary CD4 T cells were infected with WT or ΔVpu HIV-1 (NLAD8 strain). After 24h, productively infected cells were enriched by magnetic depletion of CD4high cells, and cultured with the indicated E430G-mutated antibodies and with NHS or HIHS. The % of Gag+ was measured in live cells by flow cytometry at days 1, 2, 3 and 6. (B) HIV-1-infected primary CD4 T cells were cultured with the indicated antibodies (E430G) and with NHS or HIHS. The frequency of infected cells was assessed by flow cytometry in live cells at days 1, 2, 3 and 6. Numbers indicate the percentage of infected (Gag+) cells. One representative experiment is shown. (C) Frequency of live HIV-1-infected cells (NLAD8 WT or ΔVpu strain) 6 days after incubation with the indicated antibodies, and with NHS or HIHS. Each donor of CD4 T cells is represented by a different symbol. n=8 donors of CD4 T cells. (D) Elimination of HIV-1-infected cells (strain NLAD8 WT or ΔVpu) over culture with the indicated antibody (E430G), and NHS or HIHS. The frequency of % infected (Gag+) cells is determinined by flow cytometry. The mean relative percentage of infected cells compared to the HIHS condition is depicted. n=6-8 donors of CD4 T cells.
Figure 8RBM47 is an ISG and induced by viral infection. RBM47 binds to the 3'UTR of IFNAR1 mRNA, stabilizes it, and amplifies IFNAR1 signaling.