EMBO/sd-character-level-ner huggingface dataset:
Disease stage-dependent gene expression pattern in myeloid cells during experimental CNV.Figure 8a) Trajectory analysis reveals alterations in gene expression pattern over time and disease state (d0, d3, d7). Above: Trajectory heat map depicting loss of homeostatic microglial gene expression signatures (P2ry12, Tmem119, Csf1r, SiglecH) and induction of genes involved in cytoskeleton modifications (Actb, Tuba1c), cell cycle progression (Cdk1, Ptma) and antigen-presentation (Cd74, H2-Aa). Highlighted molecules (asterisks) were confirmed by immunohistochemistry shown in b-d). Below: Trajectory plot showing the trajectory followed by myeloid cells upon CNV progress. b) Above: P2ry12 expression is shown by trajectory analysis and represented in a t-SNE plot. Expression is high in homeostatic microglia (see Fig. 5 d, e, h). Below: Typical immunohistochemistry for P2RY12 (green) in unlesioned and lesioned areas. Arrows indicate lost P2RY12 expression. Asterisks mark microglial cells that maintained P2RY12 expression. Iba1 immunohistochemistry (red) to show microglia. Pictures are representative of two mice. Scale bars represents 25 µm. c) Above: Tmem119 expression visualized by trajectory analysis and represented in a t-SNE plot. High signals in homeostatic microglia. Below: TMEM119 immunohistochemistry (green) by Iba1+ (red) microglia inside lesioned and non-lesioned areas in the retina and RPE. Arrows indicate microglia with lost TMEM119 expression whereas asterisks mark microglial cells that maintained TMEM119 expression. Picture is representative of two mice. Scale bars represents 25 µm. d) Above: Ptma expression presented by a t-SNE plot and its trajectory analysis over the disease course. Ptma expression correlates with proliferation of rMGs at d7 during CNV in Cx3cr1CreERT2:Rosa26-tdTomato mice. Below: Representative immunofluorescence of proliferating EdU+(white) Iba1+ (red, Tomato) microglia at sites of CNV on d7 (arrow) next to an EdU-Tomato+Iba1- cell (asterisks) after EdU treatment from d0 to d6. Arrowhead points towards an intravascular myeloid EdU-Tomato-Iba1+ cell. Typical pictures from three mice obtained from one independent experiment are shown. Scale bar represents 25 µm.
Cross‐linking of Beclin and PtdIns 3‐kinase.Figure 2(A) Total cell lysates were treated with DSP (lanes 3, 4, 7 and 8) or its solvent, dimethylsulfoxide (lanes 1, 2, 5 and 6), at 4°C for 2 h. Proteins were solubilized with SDS and subjected to immunoprecipitation using protein A‐immobilized anti‐PtdIns 3‐kinase (PtdIns 3‐K) (lanes 1-4) or Beclin (lanes 5-8) antibodies. Immunoprecipitates were eluted with 1× SDS sample buffer (62.5 mM Tris-HCl pH 6.8, 2% SDS, 10% glycerol, a trace amount of Bromophenol blue). After treatment with 5% 2‐mercaptoethanol, proteins were separated by SDS-PAGE, followed by detection by immunoblotting with anti‐Beclin (upper panels) and anti‐PtdIns 3‐kinase (lower panels) antibodies. (B) Cell lysates prepared from HeLa cells were subjected to subcellular fractionation by centrifugation at 100 000 g for 1 h. The membrane (lanes 1 and 3) and supernatant (lanes 2 and 4) fractions were subjected to SDS-PAGE, followed by immunoblotting with anti‐PtdIns 3‐kinase (lanes 1 and 2) and anti‐Beclin (lanes 3 and 4) antibodies. (C) Membrane (lanes 1-4; 100 000 g, 30 min, pellet) and soluble fractions (lanes 5-8; 100 000 g, 30 min, supernatant) prepared from HeLa cells were treated with DSP (lanes 3, 4, 7 and 8) or dimethylsulfoxide (lanes 1, 2, 5 and 6). Proteins were solubilized with SDS and subjected to immunoprecipitation using protein A-immobilized anti-PtdIns 3-kinase antibodies. Immunoprecipitates were separated by SDS-PAGE under a reducing condition, followed by detection by immunoblotting with Beclin antibodies. F and E indicate flow-through and elute fracions, respectively.
| Changes in Liquid-like properties of viroplasms during the course of infection.Figure 2(a) Fluorescence recovery after photobleaching (FRAP) of EGFP-tagged replication factories after 4 HPI (early infection) and 12 HPI (late infection). Scale bar, 10 μm. (b) Fluorescence Intensities after FRAP of EGFP-tagged replication factories after 4 HPI (blue) and 12 HPI (red) shown in (a). Each data point represents mean±SD intensity values calculated for multiple NSP5-EGFP-tagged granules in 5 RV-infected cells. (c) Sphericity of NSP5-EGFP-containing granules during RV infection. Each data point represents mean±SD sphericity values calculated for NSP5-EGFP-NSP5-tagged granules in cells detected in 15 frames. Data were recorded for 320 min immediately after 4 HPI when multiple NSP5-EGFP granules could be detected in RV-infected cells. (d) Calculated volumes of NSP5-EGFP-tagged granules formed in RV-infected cells after 4 HPI as shown in (c). The mean values decrease due to de novo formation of multiple smaller NSP5-EGFP granules that continuously assemble in cells between 4 HPI (t=0 min) and 9 HPI (t=300 min). The shaded area represents the 95% confidence interval for each point.
Pharmacological inhibition of iNOS reduces C26-induced derangement of amino acids and impairment of energy production.Figure 4Scores scatter plot of partial least square discriminant analysis (PLS-DA) model of metabolites from saline, C26, and C26 + GW treated mice. Metabolomic data were range-scaled and mean-centered. Pathway Analysis using MetaboAnalyst 4.0 Software comparing significantly altered pathways from saline to C26 as well as C26 to C26 + GW. Pathways are ranked by their significance and filtered based on a Pathway Impact Score >0.1. Metabolomic data were range-scaled and mean-centered. P-values were obtained using GlobalTest and the -log(p-value) corresponding to a p-value of 0.05 is indicated by the red dashed line. (left) Western blot analysis of pThr172-AMPK (pAMPK) and total AMPK (AMPK) in quadriceps muscle (n = 3). (right) Quantification of the pAMPK to AMPK ratio relative to the saline treated control (n = 3). Relative concentrations of metabolites involved in glycolysis. Relative concentrations of amino acids. Relative concentrations of odd-chain acylcarnitines. Relative estimated activity of CPT1, CPT2, LCADH, and β-oxidation.
is co-expressed with ZEB1 and ZNF596 in GBM specimens Representative images of RNAscope for LINC00115 and IHC for ZEB1 and ZNF596 expression in two GBM specimens, Sample #1 (positive) and Sample #2 (negative).Figure 8Representative images of RNAscope for LINC00115 and IHC for ZEB1 and ZNF596 expression in two GBM specimens, Sample #1 (positive) and Sample #2 (negative). Scale bar, 50 μm. Images are representative of two independent experiments. Correlation of detected expression between ZEB1 or ZNF596, and LINC00115 from a separate cohort of a total of 75 paraffin-embedded clinical GBM samples. Statistical analysis was performed by Chi-square test. Kaplan-Meier survival analysis of patients with GBM tumors expressing high or low levels of LINC00115 with high or low levels of ZEB1 or ZNF596 expression. Median survival (in months): LINC00115hi/ZEB1hi, 5.4; LINC00115lo/ZEB1lo, 27.9; LINC00115hi/ZNF596hi, 4.2; LINC00115lo/ZNF596lo, 23.8. Statistical analysis was performed by log-rank test. A working model of TGF-β-activated LINC00115-driven GSC tumorigenicity and self-renewal through sequestering miR-200s from its binding targets, resulting in enhanced both ZEB1 signaling and ZNF596/EZH2/STAT3 signal axis.
Target profiles of MicV and VrrA A) V.Figure 2V. cholerae ΔvrrA ΔmicV strains carrying pBAD-micV, pBAD-vrrA or an empty vector control (pCtr) were cultivated to early stationary phase (OD600 of 1.5) in LB. Cells were treated with L-arabinose (0.2% final conc.) and RNA samples were collected at the indicated time points after induction. Northern blot analysis was performed to determine VrrA, MicV and ompT levels. 5S rRNA served as loading control. For comparison, RNA samples of a wild-type strain carrying pCtr were collected during various growth phases, which indicated ~18-fold and ~7-fold higher levels of VrrA and MicV expressed from the pBAD plasmids, respectively (see Source data for quantifications). Venn-diagram summarizing the RNA-seq results: RNA samples were collected from V. cholerae ΔvrrA ΔmicV strains carrying pBAD-micV, pBAD-vrrA or an empty vector control. Depicted are genes displaying a fold-change of ≥3 and FDR-adjusted p-value ≤1E-8 obtained from MicV-expressing conditions (blue) or vrrA-expressing conditions (green). Genes regulated by both sRNAs (fold-change ≥3 in one condition, fold-change ≥ 2.0 in the other) are depicted in light-green. V. cholerae ΔvrrA ΔmicV strains carrying the indicated reporter plasmids (x-axis) and either an empty vector control (pCtr), the pMicV, or the pVrrA plasmid, were cultivated in M9 and GFP fluorescence was measured. Fluorescence of the control strains was set to 1. The target genes were classified according to (B): regulated by both sRNAs V. cholerae ΔvrrA ΔmicV strains carrying the indicated reporter plasmids (x-axis) and either an empty vector control (pCtr), the pMicV, or the pVrrA plasmid, were cultivated in M9 and GFP fluorescence was measured. Fluorescence of the control strains was set to 1. The target genes were classified according to regulated only by MicV V. cholerae ΔvrrA ΔmicV strains carrying the indicated reporter plasmids (x-axis) and either an empty vector control (pCtr), the pMicV, or the pVrrA plasmid, were cultivated in M9 and GFP fluorescence was measured. Fluorescence of the control strains was set to 1. The target genes were classified according to regulated only by VrrA
Model of how neoplastic transformation corrupts the physiological function of EZH2 in glioma.Figure 6During embryonic development, cell-extrinsic cues establish lineage-specific chromatin landscapes which support normal brain function. PRC2 maintains cell identity in the forebrain by repressing spinal cord-specifying HOX genes and allowing expression of EMX2, a critical regulator of neurogenesis and inhibitor of neural stem cell proliferation. Upon activation of oncogenic signalling by driver mutations, cell-intrinsic changes lead to PRC2 redistribution on chromatin and, as a consequence, to a switch in the expression of homeotic genes. With an altered identity, cells lose physiological brakes that restrain their proliferation and glioma develops. Since maintenance of the rewired transcriptional programs is required for sustaining malignant cellular properties, glioma cells become dependent on PRC2 and thus vulnerable to EZH2 inhibition.
a) Electron micrographs of striatal neurons from control and HD94 mice grown over a wild-type rat astrocyte monolayer.figf3(a) Electron microscopy of striatal neurons from control and HD94 mice grown over a wild-type ratastrocyte monolayer. Higher magnification fields show the double membrane and clear content of the cytosolic vesicles. Top right: number of vacuoles per cell profile (13-17 cell profiles in triplicate experiments; *P 0.05). Arrows, enlarged electron-clear vesicles. (b) Electron micrographs of striatal cells from mutant huntingtin knock-in (111Q-htt) and wild-type (7Q-htt) mice. (c) Electron micrographs of lymphoblasts from unaffected controls (UC) and people with HD, maintained in the presence or absence of serum. Arrows, enlarged electron-clear vesicles. (d) Immunogold for LC3 in striatal cells from mutant huntingtin knock-in mice (111Q-htt), striatal neurons from HD94 mice and lymphoblasts (lymph) from people with HD. Full fields and more details of autophagic vacuoles in Supplementary Figures 10 and 11.
Figure 3 (A) Flow cytometric analysis of B220+CD11c+NK1.1+ cells in the lung of TCM-stimulated mice. Values in the dot plots present ratios for total cells gated KikGR red and KikGR green cell  (B) Representative flow cytometric analyses of the relocation of B220+CD11c+NK1.1+ cells in lungs that were photoconverted in TCM-stimulated KikGR mouse liver (gated in red polygonal region). Percent is shown for total cells (upper panel in B), and for gated KikGR red or KikGR green cells (lower panels in B). Region where the photoconverted cells locate is magnified in the right. Yellow arrow shows the photoconversion direction. Three independent experiments. LTCM stands for TCM derived from LLC
d post sham OP or PH, treated with saline (control) or the Rho kinase inhibitor Fasudil for 1 h.Figure 6Fluorescence stainings for the apical marker CD13 on liver tissue sections from mice at 2 d post sham OP or PH, treated with saline (control) or the Rho kinase inhibitor Fasudil for 1 h. Shown are maximum projections of 50 µm stacks in the PV area. Quantification of BC diameter within 18 zones along the CV-PV axis (zone 1, peri-central; zone 18, peri-portal) from mice at 2 d post sham OP (green) or PH (red), treated with saline (control, solid line) or Fasudil (dashed line). Diameter was measured from 3D BC network reconstructions of image stacks as representatively shown in (a). The zones directly adjacent to the CV and PV were excluded from the analysis (~ 1 cell layer). Mean ± s.e.m, n=7-8 mice per condition from 3 independent experiments. BC diameter of sham operated mice treated with saline vs. Fasudil, p= 6.59*10-8; BC diameter of PH mice treated with saline vs. Fasudil, p=2.31*10-14. Fluorescence stainings for YAP, PCNA and with the nuclear marker DAPI on liver tissue sections from untreated mice or animals at 2 d post sham OP or PH, treated with saline (control) or Fasudil for 1 h. Indicated regions (dashed rectangle) are shown as magnifications in insets. Quantification of the mean nuclear YAP intensity within 10 zones along the CV-PV axis, (zone 1, peri-central; zone 10, peri-portal) from IF images as representatively shown in (c). Data was normalized to untreated animals. Mean ± s.e.m, n=7-8 mice per condition from 3 independent experiments. Nuclear YAP intensity of sham operated mice treated with saline vs. Fasudil, p=0.13 (n.s.) ; nuclear YAP intensity of PH mice treated with saline vs. Fasudil, p=0.0004.
a) Mutations in FRS on ERK2 decrease colocalization of ERK2 with ATG5–ATG12, LC3 and WIPI1.f6(a) Mutations in FRS on ERK2 decrease colocalization of ERK2 with ATG5-ATG12, LC3 and WIPI1. Immunofluorescence (IF) showing colocalization (depicted as white pixels by 'colocalization finder application') of WT-ERK2-HA, FRS ERK2 mutants (L198A-, L232A-, L198A/L232A-, Y261A-ERK2-HA) or common docking (CD) mutant (D319N-ERK2-HA) with ATG5-ATG12 (panels 1-6), LC3 (panels 7-12) or WIPI1 (panels 13-18) in EGF-treated NIH/3T3 cells. ERK2 is stained in red, and autophagy proteins are stained in green. Scale bar, 10 μm. The bars represent mean±s.e.m. **P0.01, ****P0.0001 compared with WT-ERK2-transfected cells; Student's t-test, 50 cells analysed from n=2. ( (b) Mutations in FRS on ERK2 decrease colocalization of ERK2 with nuclearATG5-ATG12 or LC3. IF showing colocalization (white pixels) of WT-ERK2-HA (red), FRS ERK2 mutants (L198A-, L232A-, L198A/L232A-, Y261A-ERK2-HA) or common docking (CD) mutant (D319N-ERK2-HA) with ATG5-ATG12 (panels 1-6) or LC3-II (panels 7-12) in EGF-treated NIH/3T3 cells. Scale bar, 5 μm. The bars represent mean±s.e.m. **P0.01, ***P0.001, ****P0.0001 compared with WT-ERK2-transfected cells; Student's t-test, 50 cells analysed from n=2. Arrows depict colocalization. (c) ATG5-ATG12 conjugation is required for ERK phosphorylation. Immunoblots for P-ERK, total ERK and β-actin in NIH/3T3 cells transfected with WT ATG5 or the conjugation-defective ATG5 K130R mutant and treated with EGF (10 min). The bars represent mean±s.e.m. *P0.05 compared with WT ATG5-transfected cells; Student's t-test, n=3.
miR-155-induced autophagy promotes the elimination of intracellular mycobacteria.Figure 7(A and B) RAW 264.7 cells were transiently transfected with control or miR-155 mimic for 24 h, and then incubated with DMSO or 3-MA for 2 h. Protein levels of LC3 were detected by Western-blot in uninfected cells (A) Intracellular mycobacterial viability was determined at the indicated timepoints by CFU assay after BCG challenge for 1 h (B) (C and D) RAW 264.7 cells were transiently co-transfected with control or miR-155 mimic together with a control siRNA or Atg7 siRNA. The expression levels of Atg7 and LC3 were detected by Western-blot (C). Intracellular mycobacterial viability was determined by CFU assay at the indicated time after challenging with BCG for 1 h (D). Values of LC3-II/β-actin ratios are indicated below the representative blot. Data are shown as the mean ± SEM of three independent experiments. *, p<0.05; **, p<0.01; NS, not significant.
Figure 2(A) Representative H&E whole-lung section at 3 dpi in B6.K18-hACE2IP-THV transgenic mice inoculated i.n. with 0.3 × 105 TCID50 of SARS-CoV-2, compared to non-infected controls (NI). In the infected lung, less transparent, purple-red areas resulted from inflammatory lesions of the lung parenchyma. Scale bar: 500 μm. (B-E) Examples of lesions observed in the lungs of infected mice at higher magnification. The two top panels depict mild (B) (scale bar: 100 μm) and mild to moderate (C) interstitium thickening accompanied by dense inflammatory infiltrates predominantly localized in the vicinity of bronchioles (yellow stars) and also present around blood vessels (green stars) (scale bar: 200 μm). (D) Alveoli filled with a proteinaceous exudate containing a few cells (green arrows) (scale bar: 50 μm). (E) Discrete degenerative lesions of the bronchiolar epithelium, such as perinuclear clear spaces (blue arrows), hyper-eosinophilic cells with condensed nuclei (orange arrow) and some intraluminal fibrinous and cell debris (black arrow) in an overall well-preserved epithelium (scale bar: 20 μm). (F) Heatmap representing the histological scores for various parameters in infected mice at 3 dpi, compared to NI controls (n = 3/group). Statistical significance was evaluated by Mann-Whitney test (*= p < 0.05, **= p <0.01, ****= p < 0.0001, ns = not significant). (G) Representative NCoV-2-specific IHC image of the brain in NI control or SARS-CoV-2-infected mice at 3 dpi. Scale bar: 500 μm. (H) Closer view of a NCoV-2 positive area from an infected mouse. Scale bar: 500 μm.
Chaperone-mediated autophagy is impaired in CTNS-deficient mice.Figure 8Schematic representation of lysosomal isolation and in vitroCMA assay [adapted from Kaushik and Cuervo (2008)]. Lysosomes were isolated from livers of starved wild-type (WT) and Ctns−/− mice as described in Supplementary Materials and Methods and incubated at 37°C for 30 min with the CMA substrate GAPDH, in the presence or absence of ATP (necessary for CMA) and protease inhibitors (Prot. Inhib.). A fraction of the CMA reactions was then mixed with sample buffer and boiled at 95°C for 5 min, followed by SDS-PAGE and GAPDH and LAMP1 immunoblotting.Quantitative densitometry analysis of CMA activity performed in independent experiments using lysosomes isolated from a total of 11 WT and 10 Ctns−/− mice. Results are mean ± SEM. ***P = 0.0005; **P = 0.0041; N.S., not significant (unpaired t-test). The ability of lysosomal proteases to mediate GAPDH degradation was assessed by incubating Triton X-100-treated WT and Ctns−/− lysosomes with GAPDH for 30 min in acidic reaction buffer and compared with input (reaction with no lysosomes). Quantitative densitometry analysis of independent reactions performed with lysosomes from a total of 9 WT and 9 Ctns−/− mice shows no significant difference in protease activity. Results are mean ± SEM.
GFP-Aub iCLIP in GSCs and targeting by piRNAs, in Cbl mRNAs (A) GFP-Aub crosslinks in cultured GSCs (Ma et al, 2017), in Cbl mRNAs.Figure 5 Figure 5: GFP-Aub iCLIP in GSCs and targeting by piRNAs, in Cbl mRNAs (A) GFP-Aub crosslinks in cultured GSCs (Ma et al, 2017), in Cbl mRNAs. Thin boxes are 5'- and 3'UTRs, lines are introns, and thick boxes are exons. The coverage of Cbl mRNAs with iCLIP reads is shown in blue. Significant crosslink clusters are shown in red. (B, C) Sequences of crosslinked regions in 5'UTR (B) and 3'UTR (C) of CblL mRNA. The nucleotides (nt) in significant crosslink clusters are in red. The sequence, occurrence and origin of anti-sense piRNAs from ovarian and embryonic libraries are indicated. Mismatched nt are in blue. (D, E) Coverage of CblL (D) and CblS (E) mRNAs with ovarian and embryonic piRNAs. Significant crosslink clusters are indicated in red at the top of the graphs. (D, E) Coverage of CblL (D) and CblS (E) mRNAs with ovarian and embryonic piRNAs. Significant crosslink clusters are indicated in red at the top of the graphs.
TBK1 promotes growth factor-induced mTORC1 signaling in a stimulus-selective manner.Figure 2A. EGF increases mTORC1 signaling in a TBK1-dependent manner: TBK1+/+ and TBK1-/- MEFs were serum starved (20 hr.), pre-treated with Ku-0063794 [1 μM] (30 min), and stimulated -/+ EGF [25 ng/mL] (0-60 min). Whole cell lysate (WCL) were immunoblotted (IB) as indicated. B. Ectopic expression of TBK1 rescues mTORC1 signaling in TBK1-/- MEFs: TBK1-/- MEFs were transiently transfected with vector control (V), wild type (WT), or kinase dead (KD) Flag-TBK1, serum-starved (20 hr.), and analyzed as above. C. EGF-receptor signaling remains intact in TBK1-/- MEFs: TBK1+/+ and TBK1-/- MEFs were serum-starved (20 hr.) and stimulated -/+EGF [25 ng/mL] (0, 5, or 15 min). D. TBK1 is required for EGF-stimulated mTORC1 and mTORC2 signaling: TBK1+/+ and TBK1-/- MEFs were stimulated with EGF as in A. E. TBK1 is not required for insulin-stimulated mTORC1 signaling: TBK1+/+ and TBK1-/- MEFs were stimulated with insulin as in C. F. Side-by-side comparison of EGF- vs. insulin-stimulated mTORC1 and mTORC2 signaling in TBK1+/+ vs. TBK1-/- MEFs: MEFs were treated as in C. G. Pharmacologic TBK1 inhibition reduces EGF-induced mTORC1 signaling: HEK293 cells were serum starved (20 hr.), pre-treated with Ku-0063794 [1 μM] (30 min) or amlexanox [50 μM] (2 hr.), and stimulated -/+EGF [25 ng/mL] (30 min) or insulin (INS) [100 nM] (30 min). H. EGF but not insulin increases mTOR P-S2159: TBK1+/+ vs. TBK1-/- MEFs: MEFs were treated as in C.
Architecture and conservation of the Trs85 subunit The Trs85 subunit colored from N- (blue) to C-terminus (red).Figure 3The Trs85 subunit colored from N- (blue) to C-terminus (red). Several features are highlighted and described in the text. Surface view of Trs85 colored by ConSurf analysis. On the left is the conservation across model organisms and humans, on the right conservation was estimated based on more closely related species. The TRAPP core binding region is highly conserved.