EMBO/sd-character-level-ner huggingface dataset:
Cell death analysis shows that miR-181a/b silencing protects SH-SY5Y cells from FCCP treatment. N≥7 independent experiments.
C Immunohistochemical localization of CD206+ perivascular macrophages (arrows) in hippocampal CA1 of Siglechdtr/dtr mice. Sections prepared 2 days after administration of PBS or DT were stained with anti-Iba1 (green), anti-CD206 (red) and anti-CD31 (blue) antibodies. Scale bar, 50 µm. D Number of perivascular macrophages in hippocampal CA1 of Siglechdtr/dtr mice 2 days after PBS or DT administration (n = 5 animals per group, two-tailed unpaired Student's t-test).
(b) Binding of myosin VI to Tom1 requires the WWY motif. Bottom, the mammalian two-hybrid assay was used to test binding of full-length Tom1 and Tom1L2 or various truncated versions of Tom1, annotated by amino acid numbers, against wild-type myosin VI tail or the myosin VI ΔWWY or ΔRRL tail. The graph shows the mean values from n = 2 independent experiments. Top, Tom1 domain structure and truncation mutants. VHS, Vps-27, Hrs and STAM; GAT, GGA and Tom1.
A. TBK1 activity is required for TLR3- and TLR4-stimulated mTORC1 catalytic activity: RAW264.7 macrophages were pre-treated with amlexanox ([50 μM] or 100 μM]) (2 hr.), rapamycin [20 ng/mL] (30 min), or Ku-0063794 [1 μM] (30 min) and treated -/+ poly (I:C) [30 μg/ml] or LPS [100 ng/ml] (60 min). Raptor was immunoprecipitated (IP), and IPs and whole cell lysate (WCL) were immunoblotted (IB) as indicated.
(A) Anti-acetyllysine Western blot analyses and total protein stains of E. coli cell extracts. Cellular protein extracts before and after induction of His6-MBP-GNAT expression were separated on 12 % acrylamide gels and immunoblotted by using an anti-acetyllysine antibody or stained with Coomassie Dye. GNAT1, 2, 5, 6, 7 and 10 as well as His6-MBP control were expressed in E. coli BL21(DE3) pLysS. GNAT3 and 4 were expressed in Rosetta(DE3). In the total protein stains, recombinant GNAT protein constructs were highlighted by blue arrowheads. Protein expression before (ni) and after (i) induction with IPTG is indicated. The luminescence signal, indicating acetylated proteins, was usually recorded after 40-120 seconds. Since MBP-GNAT4 expression resulted in a saturated signal, the luminescence was additionally recorded for 10 seconds (indicated by an asterisk).
3-DZA inhibits localization of YTHDC1 (F) and WTAP (G) in nSBs. HeLa cells were exposed to thermal stress in the presence of 3-DZA (100 µM) and stained as described above. Scale bar: 10 µm.
A Agarose gel image illustrating PCR amplicons of ltaS (LBA0447), slpB, and slpX deletions in NCK2187.
(F) FACS assay of interactions of galectin-9 with wild -type M. marinum (Mm_WT) and MMAR-5356 and MMAR-5357 mutants of M.marinum treated with Tet. The blank control was Mm_WT staining with APC-anti-rabbit antibody alone. Data shown are representative of n=3 independent experiments.
Immunohistochemical detection of PLN in paraformaldehyde (PFA)-fixed testis from wild type and SPPL2c-/- mice. Cryosections were stained using a rabbit monoclonal PLN antibody or normal rabbit IgG as negative control and diaminobenzidine as peroxidase substrate.
(C) GFP-LC3-positive vesicles were detectable in 293/GFP-LC3 cells after siRNA depletion of β′-, α-, and β-COP but only basal levels of GFP-LC3-positive vesicles are observed in control siRNA-depleted cells.
(e) β-galactosidase staining of HeLa cells transfected with a control luciferase siRNA or VHR siRNA and cultured for three days. Note the increased number of blue-staining granules in many cells. (
(A) Representation of the vectors used in this study. WTFlx Ndufs3 cDNA was expressed from rAAV9-Ndufs3. An rAAV9-eGFP expressing viral preparation was used as a control.
(c) Representative images of an MCF10A E7+Bcl2 cell-in-cell structure formed in soft agar. The arrow marks LC3 recruitment around internalized cells. Scale bar, 5 μm.
F. 293T cells were transfected with 2 μg of FLAG-tagged RIG-I and 1, 5, or 10 μg of V5-tagged HDAC6. At 36 h post-transfection, whole cell lysates were prepared for co-immunoprecipitation analysis of V5-tagged HDAC6 and acetylated K909 (on RIG-I wild-type). Samples were analyzed by western blotting with a K909 acetylation-specific antibody.
Droplet formation of mCherry-fused recombinant proteins of the S3-LCEs. Representative images are shown below the schematic of the mCherry fusion protein. Detailed information about the indicated S3-LCEs is shown in Fig EV4B. The recombinant proteins (20 nM) were incubated with 75 mM NaCl in droplet formation buffer.
T1-weighted DCE-MRI of an ALSP patient with the ΔA781_N783 CSF-1R variant. Colourmetric scale indicates the slope of the quantified Gd-BOPTA accumulation, as an indicator of BBB permeability.
C, Immunostaining of endogenous Trnp1 and DAPI in P19 cells and E14 cortical cells at 1 day in vitro (div).
ISC (eGFP+ve) cells were FACS-sorted from Lgr5-DTReGFP embryos at E16.5 and subjected to RNAseq analysis. Heatmap of differentially regulated genes in HE and KO ISCs at E16.5. Biological replicates for RNAseq experiments on ISC Lgr5-DTReGFP E16.5 independent embryonic pools, each obtained from independent litters (n= 2 HE, 2 KO)
(C) Double immunofluorescence for ETS1 and β-catenin in intestinal biopsies of healthy controls and inflamed mucosa from UC patients. Scale bar: 50 μm.
YTHDC1 accumulation within nSBs was arrested by METTL3 knockdown. HeLa cells were stained as described in Figure 1C. Scale bar: 10 µm. Box plot of relative intensities of YTHDC1 to HSATIII within nSBs in individual nuclei (n=30). The mean is indicated with X. The first and third quartiles are the ends of the box, the median is indicated with a vertical line in the box, and the minimum and maximum except for the outliers are the ends of the whiskers. The outliers are indicated with open circles. P-values (Kruskal-Wallis test, followed by Dunn's multiple comparison test) are shown above.
B Growth curves of the indicated strains shown as population doublings (pds) on the y axis and hours in liquid culture on the x axis. Before inoculation in liquid YES medium, single cells underwent approximately 25 pds to produce visible colonies on solid medium. For each strain, averages and SDs (error bars) of three biological replicates are shown. Arrows indicate times of harvesting and corresponding pds for experiments shown in (C) and (D).
Figure 3. CD22 is highly clustered on the surface of resting B cells(A-C) dSTORM analysis of CD22, CD19, IgM and IgD. Wild-type primary B cells were stained with Alexa647-conjugated antibody against CD22, Cy5-conjugated antibody against CD19 or IgD, or Cy5-conjugated Fab fragments against IgM and settled onto nonstimulatory coverslips. Cells were then fixed, imaged with dSTORM and analysed.(A) 2D (top and middle) and pseudo 3D (bottom) dSTORM images were reconstructed from single molecule localizations. The white dashed square is shown in magnification (middle) and pseudo 3D (bottom).(B and C) Quantification of the distribution of CD22, CD19, IgM and IgD with H function and Hopkins index. Error bars (Hopkins index) and thin lines (H function) denote mean ± SEM. Data are pooled from at least three experiments per receptor.*P < 0.001, **P < 0.0001 (Student's t-test).
(E) RNA Pol II ChIPseq and RNAseq signals of PMAIP1, CDKN1A, SESN2 and SESN3 in HGC-27 cells. Red bars represent the RNA Pol II ChIPseq signal. Orange or blue bars represent the RNAseq signals of HGC-27 cells overexpressing lncPSCA (lncP) or with silenced DDX5 (shD), respectively.
F. Whole cell lysates from HDAC6+/+ and HDAC6-/- MEFs were prepared and used in dsRNA pull-down assays. Pull-down samples were analyzed by western blotting with anti-RIG-I, anti-HDAC6, and anti-β-actin antibodies. Intensity of pull-downed RIG-I was quantified.
(B-Q) Changes in fluxes of key metabolic reactions highlighted in (A) compared to baseline fluxes (dashed line) based on personalized modelling, where 'SX' indicating flux obtained by the modeling of subject number 'X' as shown in Table EV13.
(B) Recruitment of endogenous LRRK2 and EGFP-Rab8A to M. tuberculosis (Mtb) was visualised by confocal microscopy. Scale bar = 5 μm.
(A-B) Representative confocal images of Gb3 accumulation, revealed by STX staining, in brain sections from CLN7Δex2 mice at 3 and 7.5 months of age compared with CLN7 WT mice. Scale bars: 80 µm.
(B) qPCR analysis of ATF3 and IL‐8 induction following infection of HeLa cells with Listeria WT or TKO for 1-4 h. Values are means±s.e.m. n=3.
Human PBMCs were cultivated for 48h in RPMI containing 80mg/dl glucose (NC) and supplemented with 10mM beta-hydroxybutyrate (BHB). T cell stimulation was performed through CD3/CD28 Dynabeads at a bead:cell ratio of 1:8. Pan T cells, CD4+ and CD8+ T cells were isolated through magnetic cell separation. Mitochondrial metabolism was analyzed for each subpopulation using a Seahorse XF96 Analyzer. (A-C) OCR, maximum respiration, spare respiratory capacity and basal respiration were measured in A pan T cells, B CD4+ T cells and C CD8+ T cells, n=9 (pan T), n=3/4 (CD4 unstimulated), n=7/8/9 (CD4 stimulated), n=5 (CD8 unstimulated) and n=8/7/7 (CD8 stimulated) individual experiments. Data depicted as mean ± SEM (OCR) and box plots with median, twenty-fifth and seventy-fifth percentiles and range (all other). Dots indicating individual values. *p<0.05, **p<0.01, paired t-test/ Wilcoxon matched-pairs signed rank test, as appropriate.
(O, P) Immunoblotting analysis of PR82 levels in WT, NDST3 KO, and Noco-treated NDST3 KO RPE1 cells from (M, N) after the drug treatment. Quantification of the levels of PR82 relative to β-tubulin is shown in (P) (n = 4 independent experiments, *P = 0.0141, **P = 0.0045). Data information: Error bars represent ± standard deviation.
(G) Unscheduled DNA synthesis in non-cycling C5RO cells measured by the incorporation of EdU (20 µM) after UV irradiation (16J/m2) visualized by ATTO594 labelling using Click-chemistry. Levels in siControl cells were set as 1. N=2, >200 cells analyzed, error bars represent SEM
B) In vitro RB ubiquitination by MDM2 wild type and the phosphomimetic mutant MDM2(S395D).
F-I ChIP assays with antibodies against H3K9me3, H4K20me3, H3K4me3 and H3K9Acetyl were performed as in (D). Only 'C' position signals were showed and normalized to Input.
Overview of mutations found in organoid cultures and matching tumor tissue obtained by targeted sequencing of 121 HGSOC related genes confirms almost identical profile between parental tissue and in vitro long-term organoid culture. Color code indicates type of mutations which were detected.
(D) Representative IHC images of CDC50A expression at synapses. Scale bar, 5 µm. Arrows indicate VGluT2+ RGC inputs. (E) Relative CDC50A expression at mCherry+ synapses in Cdc50a RNAi mice and controls. N = 3 for biological replicates, mean ± SD, * p < 0.05 by Student's t-test.
Co-immunoprecipitation of exogenous HA-USP28 and FLAG-ΔNp63 in HEK293 cells. Either HA-USP28 or FLAG-ΔNp63 were precipitated and blotted against FLAG-ΔNp63 or HA-USP28. The input corresponds to 10% of the total protein amount used for the IP (ACTIN as loading control).
C, D Cr-release assay showing % specific lysis of MCF7 cells by survivin-specific T cells at different ratios upon CCR9 knockdown (C) or overexpression (D). MCF7 cells were transfected with either CCR9 siRNA s1 (Δ), pooled siRNA sequences (○), positive control PD-L1 (□), and non-specific control siRNA (▪) (C) or with pCMV6-AC-His control vector (▪) and pCMV6-AC-His-CCR9 expression construct (○) (D) 72 h prior to the assay.
Figure 5d e. Examples of genes whose transcription processivity and expression is dependent on the CDK12 kinase activity. Nuclear RNA-seq data on the respective strand and RNAPII, P-Ser2, P-Ser5 and SPT6 ChIP-seq data for UBE3C genes from cells either treated (red) or not (blue, CTRL) with 3-MB-PP1 were visualized with Gviz. Read counts were normalized to the total number of mapped reads per sample and averaged between replicates. Blue and red boxes below the RNA-seq data indicate the 90% distance (see Fig. 7d and 7e and corresponding text) in control and CDK12 inhibited samples, respectively.
Swelling‐induced efflux of 3[H]‐taurine from WT and LRRC8D −/− HEK cells (A, B) and partial rescue by transient transfection of LRRC8D (B). Rescue is incomplete due to low transfection/expression efficiency of LRRC8D (Voss et al, 2014).
D, E. Cell lysates of HEK293 cells transfected with empty vector control, Myc-C9orf72S or Myc-C9orf72L were subjected to immunoprecipitation with anti-Myc antibodies Immune pellets were probed for Myc-C9orf72 and endogenous FIP200, ULK1, and ATG13. There are multiple alternatively spliced forms of ATG13 (Jung et al., 2009; Alers et al., 2011); ATG13* is most likely a smaller alternative spliced form of ATG13 that is enriched by interaction with C9orf72.
B. Circular representation of lysine-lysine inter-subunit cross-links identified by mass spectrometry (XL-MS) in buffer containing 250 mM NaCl. For simplicity, cross-links between proteins (or domains) are grouped, and the thickness of the lines indicate the total number of crosslinks of this particular type. Smc5 and Smc6 proteins are divided into N- and C-terminal head domains (HD) and coiled-coil arms (CC) as well as the central hinge domain. For a full representation of individual inter-subunit as well as intra-subunit cross-links
A. HEK293 cells treated for 4 h with 100 μM H2O2. Phospho-histone H2A.X (γH2A.X) was assessed by immunofluorescence analysis. Nuclei (blue) were stained with Hoechst 33342. scale bar: 10μM. 4X digital magnification is shown at the bottom left of each image.
(B) Metagene profile of ChIP data (Data ref: Ha et al, 2019). The genomic intervals cover the first 500 bp of the coding gene (scaled to 100 bins), the first 1 kb of the DNC transcript (scaled to 200 bins), and the variable length gap between the coding TSS and the DNC TSS (scaled to 50 bins). The vertical lines represent transcription start site (TSS) of protein-coding gene (left) and the DNC transcript (0 bin). The H3ac ChIP-seq signal was normalized by the H3 signal in the same samples.
Representative pictures of BAT and inguinal WAT (iWAT) hematoxylin and eosin staining from mice in (E). Scale bar: 20μm for BAT, 50μm for iWAT.
G-J Effects of TAT-2031 treatment on TBS-induced LTP (G) or LFS-induced LTD (I) in CA1. H, J Means ± SEM of fEPSPs measured 40 min after TBS (H, n = 6 slices from 6 mice) or LFS (J, n = 3-4 slices from 3-4 mice) in different groups. Data information: Slopes of fEPSPs were normalized to the average values recorded during the first 10 min baseline Statistical significance was assessed by Student's t-test *P < 0.05, **P < 0.01, ***P < 0.001 compared with shSc, Vehicle, or TAT Insets show representative traces of evoked fEPSPs before and 40 min after TBS/LFS. Scale bar 0.5 mV/10 ms.
d, ISX transcriptionally activates luciferase activity driven by Snail1 (d) promoter regions in A549 cells. Data are presented as mean ± SD in bar graph (p < 0.001 , Student's t‐test) of 3 independent experiments, each performed in triplicate.
(G) Human primary macrophages differentiated from PBMCs from two healthy donors were infected with H5N1 Wt and ΔF2 for 5 h at MOI 10. Levels of IL-1β were measured by ELISA in technical replicates. The mean of two independent experiments is shown. Statistical analysis was performed by one-way ANOVA, p-values are indicated.
O ISRE reporter activity in co-transfection of HA-MAVS (200 ng) together with Myc-SIRT5 (200ng) or Myc-SIRT5-H158Y(200ng) respectively in HEK293T cells for 24 h. Data information: Graphs represent fold-induction relative to the luciferase activity in the control cells. UI, uninfected. All data are presented as the mean values based on three independent experiments, and error bars indicate S.E.M. (unpaired two-tailed Student's t test).
(A) Model of Rag GTPase-dependent and Rag GTPase-independent pathways that signal to mTORC1. Alanine (Ala), arginine (Arg), histidine (His), leucine (Leu), methionine (Met), serine (Ser), threonine (Thr), and valine (Val) promote mTORC1 lysosomal localization and activation through the Rag GTPase pathway. Glutamine (Gln) and asparagine (Asn) promote mTORC1 lysosomal localization and activation independently of the Rag GTPases, and require Arf1 through an unknown mechanism. Both pathways require growth factor signaling resulting in Rheb GTP-bound allosterically activating mTORC1.
(D) Quantification of the telomeric overhang signal at d11 after doxycyclin addition to SMCHD1 wild-type and SMCHD1 knockout HeLa shTRF2 inducible cells. The bar graph represents the average overhang signal intensity from two biological replicates as percentage of the signal in the cells untreated with doxycycline with error bars representing the standard deviation (SD) of the sample.
SNX18 tubules are positive for TfR. HeLa GFP-LC3 cells were transfected with myc-SNX18 or a control myc plasmid and then immunostained for myc and TfR (A) or Rab11 (B). Bars: (A) 10 µm; (B) 5 µm. Insets show enlarged views of the boxed regions.
G. Images of HeLa cells stably expressing PRC1-GFP treated with control (left image) and siRNA targeting PRC1 (right image) immunostained for PRC1 (magenta). Graph showing the length of the immunostained PRC1 signal, LPRC1 (p=0.5167), signal intensity, I (p=0.0003) and Icross (p=0.0001) in siRNA targeting PRC1 in comparison to control HeLa cells expressing PRC1-GFP and immunostained for PRC1.H. Images of HeLa cells stably expressing PRC1-GFP (green) treated with control (left image) and siRNA targeting PRC1 (right image) immunostained for PRC1. Graph showing the length of the PRC1-GFP signal, LPRC1 (p=0.3407), signal intensity, I (p=0.0001), and Icross (p=0.0001) in siRNA targeting PRC1 in comparison to control HeLa cells expressing PRC1-GFP and immunostained for PRC1.
Coupling efficiency, measured as the ratio of ATP-linked/Basal respiration x 100. Error bars represent the mean±SEM of 4-6 separate experiments
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).
(C) Co-immunoprecipitation experiment upon co-expression of FLS2-HA, BAK1-HA and RGI3-GFP in N. benthamiana. Protein extraction was performed 30 min after mock, 1 μM flg22 or 1 μM GLV2 treatment. IP was performed using GFP-TRAP agarose beads. Western blots for protein detection were probed with α-GFP or α-HA antibodies. Data information: Similar results were obtained in 3 independent experiments.
E Gating strategy for the identification of U-2932 cells residing in spleen and bone marrow, among all leukocytes in these organs. Cells were stained for human and mouse CD45. Very few hCD45+ U-2932 cells were detectable in the spleen, whereas their frequencies were much higher in the bone marrow (BM). In BM, >90% of U-2932 cells were positive for ZsGreen. The green curve shows U-2932 cells recovered from BM, overlayed with unlabelled U-2932 cells grown in vitro (grey curve).
Sublingual infection of WT, Stim1fl/flVav-iCre and Stim1fl/flStim2fl/flVav-iCre mice with 2x107 CFU of C. albicans. (D) Histological images of H&E (left) and PAS (right) stained tongue tissue at day 7 p.i. Clusters of polymorphonuclear (PMN) cells (neutrophils, white arrows) and Candida (black arrows) are indicated.
C Gene expression heatmaps of selected genes from the bound and activated and bound and repressed groups show activation of key hematopoietic target genes and repression of key cardiac target genes in Scl‐expressing mesoderm as compared to Scl‐deficient mesoderm. Control [Scl+/hCD4, divided into Scl‐expressing (Scl+) and non‐expressing cells (Scl−)] and SclKOEBs are shown.
(A) HH/SMO promotes slow-migrating forms of Smaug. Western blot analysis of Cl8 cells that transiently express HA-SmaugWT alone, or together with either SMOWT-GFP or SMOPKA-SD-GFP, in the presence or in absence of HH. GMAP serves as a loading control. Here and in the other panels, the antibodies used are indicated on the left. Here and in (B,C), the black arrows indicate the unphosphorylated form of HA-SmaugWT and the brackets indicate the slower migrating phosphorylated forms of HA-SmaugWT. NT: not transfected. See a phosphatase assay in Fig EV4A.
(M) Control shRNA and B56δ shRNA cells were transfected with vector, MYC-PPM1G and MYC-PPM1G ΔNLS in indicated combination and pS641 α-catenin levels were determined using western blotting.
The regional distribution of differential exon skipping events between WT and hnRNPLL-KO ES cells.
(B) IP of YFP-B56α from cells stably expressing the B56α WT, R222E, and 5A followed by immunoblotting of indicated proteins. Representative blots are shown (top). hSgo1 signals were normalized to YFP and plotted (bottom). Error bars represent SD (n=4).
Immunoblot analysis of total and phosphorylated ZAP70 and PLCγ1 expression in anti-CD3-stimulated thymocytes isolated from Ifih1-/- and Cd4cre A1fl/fl Ifih1-/- mice. The expression of GAPDH is shown as a reference. Representative images are shown (left panel). The band intensity of p-ZAP70 and p-PLCγ1 was normalized to that of total ZAP70 and PLCγ1, respectively and displayed as the mean ± SEM (n = 3; Mann-Whitney U-test, N.S.; not significant) (right panel) by setting the mean value of stimulated Ifih1-/- samples at 60 sec as 1.
a, BMDMs were infected with mCherry-expressing M. tuberculosis for 4 h and immunostained using linkage-specific ubiquitin antibodies. b, Quantification of K63 and K48 ubiquitin-positive M. tuberculosis from a.
Schematic of inferred topology of neuropeptidergic signalling axes between SCN neuronal sub-populations in circadian day. Top: clusters are annotated in the overview t-SNE for circadian day (A.) and night (B.), respectively. Below: the identified SCN subpopulations are depicted as nodes of each neuropeptide-receptor network, connected by edges that are weighted (shown as differential line thickness) based on the average count of the respective cellular gene expression for each ligand and receptor. The nodes of the inferred topology networks are colour-coded to match those of the overview t-SNE plots. Nodes further highlighted in yellow mark ligand-enriched source clusters. The edges are colour-coded for each neuropeptide-receptor signalling pair: Prok2/ProkR2 - purple, Vip/Vipr2 - green, Avp/Avpr1a - blue. The sum of edge weights within each day and night inferred signalling network is presented below each map to provide a quantification of the overall degree of connectivity.
Representative images (D) to visualize SA-β-gal activities in vehicle-, doxorubicin- or abemaciclib-treated mouse kidney sections at 15 dpt (arrows indicated positive area; scale bar, 1 mm; n=3 mice/group)
For each condition in (A), minor intron splicing (Figure 4A) was plotted against the percentage of cells showing a mitotic phenotype (Figure 2C). R2 and p-values are provided for the linear regression trendline (dotted line).
(G) Time course of mTOR inactivation during EBSS starvation in WT vs. STX16KO HeLa cells analyzed by Western blotting for mTOR substrates ULK1 and 4E-BP1.
F, G: Tissues were analyzed by label-free quantitative proteomics. C I-V, complex I-V; CAC, citric acid cycle enzymes; MICOS, contact site complex; mt TA, mitochondrial transaminases; NS, not significant; PDH pyruvate dehydrogenase; VDAC, voltage-dependent anion channel
E. Lysates from HeLa cells were incubated with in vitro synthesized biotin-labeled sense or antisense DNA probes against lncRNA-MIF for biotinpull-down assay, followed by real-time RT-PCR analysis to examine miR-586 levels. Data shown are mean ± SD (n = 3; *P < 0.05, **P < 0.01, two-tailed t-test).