EMBO/sd-character-level-ner huggingface dataset:
Reduced Expression of Bcl6 by Bob1-deficient Tfh cells.Figure 6(A) Expression of Bcl6 in CXCR5hiICOS+Tfh cells in Pou2af1-/- and Pou2af1+/-mice seven days after immunization with SRBC. Left: Representative histogram showing Bcl6 expression in Pou2af1-/- (red) and Pou2af1+/- (blue) CD4+CXCR5hiICOS+Tfh cells; filled line: Bcl6 isotype control. Right: Summarized results illustrating the proportion of CXCR5hiICOS+Tfh cells that express Bcl6 in Pou2af1-/- and Pou2af1+/-mice. (n=5, bars indicate means). (B) Bcl6 expression by Tfh cells in Pou2af1+/+:Pou2af1-/- mixed bone marrow chimeras seven days after immunization with SRBC. The graphs illustrate the proportion of Bcl6hi CXCR5hiICOS+ Tfh cells within the population of Pou2af1+/+ and Pou2af1-/- CD4+ T cells from bone marrow chimeras (n=7). (C) Expression level of Bcl6 in Pou2af1-/- and Pou2af1+/+ CXCR5hiICOS+ Tfh cells vs. non-Tfh cells from mixed bone marrow chimeras and a comparison of the mean fluorescence intensity (MFI) for Bcl6 (n=9). (D) Frequency of CXCR5hiBcl6hi GC Tfh cells among CD4 cells in Pou2af1-/-:Pou2af1+/+ bone marrow chimeras (n=9).
Figure 7(A) Comparison of RAG2 and BbeRAG2L structures. Red color indicates the amino acid 333-342 RAG2 loop and the equivalent loop of BbeRAG2. The four amino acid deletion, (∆L in (D)), is indicated with green brackets. Due to areas of low resolution in the BbeRAG2L structure, some regions that are likely to be beta strands cannot be modeled as such. (B) Sequence alignment of RAG2 and RAG2-like proteins in the vicinity of mouse RAG2 loop 333-342. Green shading, four amino acids deleted at tip of loop. Red letters in blue box, relatively well-conserved residues; white letters on red background, highly conserved residues. (C) Cryo-EM map (transparent) and cartoon model of RAG2 showing contact between the extended loop (red) and target site DNA (yellow; stick model). (D) Results of in vivo plasmid-to-plasmid transposition assay using WT or R848M RAG1 and the indicated forms of RAG2 containing or lacking (∆L) the four amino acids at the tip of the 333-342 loop, performed as described in (Zhang et al., 2019); (****, p<0.0001). Three biological replicates for each condition except in the last two columns where data from 11 biological replicates are presented. (E) Results of in vivo plasmid-to-genome transposition assay using WT or R848M RAG1 and different forms of RAG2 as indicated. As previously demonstrated (Zhang et al., 2019), R848 (present in WT RAG1) and RAG2 acidic hinge residues 362-383 each potently suppress transposition. Substantial transposition is observed with the combination of RAG1 R848M and RAG2 1-361, which is further increased by ∆L; (***, p=0.001). Four biological replicates for each condition. (F) In vivo recombination activity in HEK293T cells of mouse wild-type RAG1 or RAG1 R848M with RAG2 1-361 or 1-361 ∆L. (p=0.42 for wild-type RAG1 group, p=0.41 for RAG1 R848M group). Three biological replicates for each condition.
a) Schematic view of human Parkin structure (UBL, ubiquitin-like domain; R0, RING0/unique Parkin domain; R1/2, RING finger domains; IBR, in-between-ring domain).figf1(b) Parkin is recruited selectively to depolarized mitochondria and directs mitophagy. HeLa cells transfected with HA-Parkin were treated with CCCP for the indicated times. Mitochondria were stained by anti-TOM20 (pseudo coloured; blue) and a ΔΨm dependent MitoTracker (red). Parkin was stained with anti-HA (green). Without treatment, mitochondria are intact and stained by both mitochondrial markers, whereas Parkin is equally distributed in the cytoplasm. After 2 h of CCCP treatment, mitochondria are depolarized as shown by the loss of MitoTracker staining. Parkin completely translocates to mitochondria clustering at perinuclear regions. After 24 h of CCCP treatment, massive loss of mitochondria is observed as shown by the disappearance of the mitochondrial marker. Only Parkin-positive cells show mitochondrial clustering and clearance, in contrast to adjacent untransfected cells. Scale bars, 10 μm. (c) Pathogenic Parkin mutations abrogate mitochondrial translocation and/or clearance. HeLa cells transfected with Flag-Parkin wild-type or pathogenic mutants were treated with CCCP and stained with anti-Flag (green), anti-TOM20 (red) and Hoechst33342 (blue). Representative merged images are shown. Scale bars, 10 μm. Immunostaining of cells transfected with non-functional cysteine mutants and functional Parkin variants is shown in Supplementary Information, Fig. S1a and S1b, respectively. (d, e) Based on the mitochondrial effects, Parkin mutants can be classified into functional and non-functional classes. (d) Quantification of Parkin translocation to damaged mitochondria after 2 h of CCCP treatment, as defined by complete colocalization of Parkin with the mitochondrial marker (n = 3). (e) Quantification of cells with uncleared mitochondria after 24 h of CCCP treatment, as shown by complete loss of mitochondrial staining (n = 3). Data represent the mean ± s.d., *P ≤ 0.05, **P ≤ 0.005, ***P ≤ 0.0005 compared with wild-type. WT, wild-type.
Figure 7A. Hierarchical clustering of proteins significantly differentially abundant between NASH (n=20), cirrhosis (n=10) and control groups (n=15). Significance was calculated by ANCOVA, followed by Benjamini-Hochberg correction for multiple hypothesis testing (FDR < 0.05). Two major clusters of proteins were identified with Cluster 1 mainly upregulated in cirrhosis compared to NASH and controls and Cluster 2 downregulated. No additional replications of the experiment was done in laboratory. B. Ten representative highly enriched GOBP terms in proteins in Cluster 1 of Panel (A). Significance was calculated by Fisher's Exact test corrected for multiple hypothesis testing with FDR < 0.05. C. Ten representative highly enriched GOBP terms in proteins in Cluster 2 of Panel (A). Significance was calculated by Fisher's Exact test corrected for multiple hypothesis testing with FDR < 0.05. D. Box‐and‐whisker plot showing the distribution of log2‐intensity values of statistical significantly regulated proteins across three groups. Number of independent biological replicates is n=15, 20 and 10 for the control, NASH and cirrhosis group, respectively. The black line in the middle of the box is the median, the top and the bottom of the box represent the upper and lower quartile values of the data and the whiskers represent the upper and lower limits for considering outliers (Q3+1.5*IQR, Q1‐1.5*IQR) where IQR is the interquartile range (Q3-Q1). Significance was defined by ANCOVA followed by Benjamini-Hochberg correction for multiple hypothesis testing (FDR < 0.05) with a significance level of ***P < 0.001. E. Heatmap of CYP 450 family members that are statistically significantly abundant between three groups. Data was presented as mean protein abundance followed by Z-score normalization across. Number of replicates is same as Panel (C). F. Extracellular matrix (ECM) remodeling in liver cirrhosis. Upper and lower panel shows the abundance rank of proteins involved in ECM organization in the control (upper) and cirrhosis (lower) groups, highlighting top shifted ECM components in the cirrhosis group as compared with the control group.
Acetylation of FXR at K217 in obesity promotes hepatic inflammation A, B FXR‐WT or K217 mutants were expressed in livers of lean (ND) (A) or obese (HFD) (B) mice as indicated, and Illumina microarray was performed followed by gene ontology analysis.Figure 2A, B FXR‐WT or K217 mutants were expressed in livers of lean (ND) (A) or obese (HFD) (B) mice as indicated, and Illumina microarray was performed followed by gene ontology analysis. C, D The mRNA levels of selected inflammatory genes were confirmed by qRT-PCR for lean (ND) (C) and obese (HFD) (D) mice (n = 4). E Hepatocytes were isolated from FXR‐KO mice and infected with Ad‐FXR‐WT or the Ad‐K217Q mutant, and the mRNA levels of inflammatory genes were measured by qRT-PCR. F, G Histology of the liver of lean (ND) (F) and obese (HFD) (G) mice stained for F4/80 by IHC to detect macrophages and by HE, or with Oil Red‐O to detect liver fat. Infiltrated macrophages are indicated by arrows. H, I Cytokine and TG levels in liver extracts of lean (ND) (H) and obese (HFD) (I) mice (n = 4). J, K Glucose tolerance test (GTT): Serum glucose levels of lean (ND) (J) and obese (HFD) (K) mice were measured at the indicated times after injection i.p. of glucose (n = 4). L Ad‐FXR‐WT and K217 mutants were expressed in hepatocytes isolated from lean mice and treated with insulin for 30 min, and phospho‐ and total AKT levels were measured by IB (n = 3).
a,b) The occupancy of dUTX on the promoter regions of apoptosis and Atg genes was detected by chromatin immunoprecipitation (ChIP).f5(a,b) The occupancy of dUTX on the promoter regions of apoptosis and Atg genes was detected by chromatin immunoprecipitation (ChIP). SL2 cells overexpressing HA-dUTX were chromatin-immunoprecipitated with anti-HA and anti-GFP (IgG control) antibodies following ecdysone treatment. qPCR was used to assess the binding to the promoters of dark, dronc, drice and rpr (a) (a,b) The occupancy of dUTX on the promoter regions of apoptosis and Atg genes was detected by chromatin immunoprecipitation (ChIP). SL2 cells overexpressing HA-dUTX were chromatin-immunoprecipitated with anti-HA and anti-GFP (IgG control) antibodies following ecdysone treatment. qPCR was used to assess the binding to the promoters of Atg1, Atg2, Atg3, Atg4, Atg5, Atg6, Atg7, Atg8a, Atg9, Atg12 and Atg18 (b), expressed relative to the internal control gene Gapdh. Results are shown as the fold enrichment of % input compared with IgG control. (c) The abundance of H3K27me3 on the promoters of apoptosis and autophagy genes detected by ChIP. SL2 cells knocked down for dUTX were chromatin-immunoprecipitated with anti-H3K27me3 and anti-GFP (IgG control) antibodies following ecdysone treatment. Results are shown as the fold enrichment of % input compared with RNAi control. (d) The occupancy of EcR on the promoter regions of dark, dronc and Atg1 by ChIP expressed relative to the internal control gene rp49. Chromatin was immunoprecipitated with anti-EcR-B1 and anti-GFP (IgG control) antibodies from SL2 cell following ecdysone treatment. Results are shown as the fold enrichment of % input compared with IgG control. Data in all panels are expressed as means from three independent experiments, with error bars representing s.e.m. *P0.05 (Student's t-test).
Figure 7Diagram of the pre-rRNA processing showing the pre-rRNA processing intermediate products and the location of probes (5'-ETS, ITS1 and ITS2) used in Northern blot analysis (left) as well as the major (black) and minor (grey) processing pathways (right). (B). USP36 is required for rRNA processing. HeLa cells transfected with scr or USP36 siRNA (left panels) or infected with scr or USP36 shRNA lentiviruses (right panels) were assayed for rRNA processing by Northern blot using 5'- ETS, ITS1 and ITS2 probes as indicated. Knockdown of USP36 inhibits translation. HeLa cells infected with scr or USP36 shRNA lentiviruses were incubated with O-propargyl-puromycin (OPP) followed by Click-iT OPP protein synthesis assays. Representative images (C) and quantification from three independent experiments (D). Data were presented as mean ± SD, n=3 biological replicates. ***P<0.001, compared to scr control (Student's t-test). Scale bar = 50 μM.
Figure 7RT-PCR validation of Bptf and Tbx3 alternative exon deletion, separately in hnRNPLL-KO H8 cells (upper) and simultaneously in hnRNPLL-KO H8 cells (lower). AP staining of H8-Bptf or H8-Tbx3 exon-KO ES cells after LIF withdrawal. hnRNPLL-H8, hnRNPLL-KO ES cells; H8-Bptf KO, alternative Bptf exon was deleted in hnRNPLL-KO H8 cells; H8-Tbx3 KO, alternative Tbx3 exon was deleted in hnRNPLL-KO H8 cells; H8-Bptf/Tbx3 KO, alternative Bptf or Tbx3 exons were simultaneously deleted in hnRNPLL-KO H8 cells. Scale bar, 200 μm. Colony formation assay of H8-Bptf or H8-Tbx3 exon-KO ES cells after LIF withdrawal. The data represent the means± SD (n=3, from biological repeats). Hematoxylin and eosin labeling of teratomas derived from WT and H8-Bptf or H8-Tbx3 exon-KO ES cells. The blue triangle represent ectoderm, the dark one represent mesoderm and the green one represent endoderm. Scale bar, 100 μm. Oct4 immunohistochemical staining of teratomas derived from WT and H8-Bptf or H8-Tbx3 exon-KO ES cells. Representative Oct4-positive regions are shown (left). Scale bar, 100 µm. The percentage Oct4-positive region (%) is shown (right). The data represent the means± SD (n=3 biological repeats). **P＜0.01; ns, not significant. T-test was used to determine the significance. Schematic representation of hnRNPLL regulating the AS of crucial genes that have an important role in mediating exit-from-pluripotency.
Figure 3A LC score does not correlate with smoking history. Box plot of the LC score detected in EBCs from control (Ctrl) and lung cancer patients (LC). Samples were grouped based on the smoking history into never smokers (NS), previous smokers (PS) and current smokers (CS). B LC score does not discriminate between the major LC subtypes. Box plot of the LC scores detected in EBCs from Ctrl, non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC) patients. C LC score can be used for detection of early staged LC. Box plot of the LC scores detected in EBCs from Ctrl and LC patients of stage I, II; III and IV. Patients were staged according to the TNM Classification (UICC, 7th edition).
Figure 7Heatmap showing log2 fold changes of mRNA expression in the cho2opi3 parent strain cultured in SD C-I- for 13 h vs. wild type, and in co S#3, #4, and #5 cultured to mid-log phase in C-I- vs. wild type and vs. parent. Transcripts changing more than 1.7-fold (p < 0.01) in at least one of the comparisons are depicted. Hierarchical clustering was by average linkage (cosine correlation); (functional) categories of enriched transcripts were assigned per cluster. Transcript profiles of the PC-depleted parent and evolved suppressors vs. wild type of UPR-induced genes (Kimata et al., 2006; Travers et al., 2000) changing more than 1.7 fold (p < 0.01) in the parent, and of ORE-genes from the cluster in Fig. 6A. Heatmaps show log2 fold changes according to the color scales; asterisk (*) indicates gene on chromosome XV. POX1 transcript levels after switching the strains indicated from SD C+I+ to the medium indicated at OD600 0.02 or 0.2 (co diploid in C-), and culture for 24 h at 30 oC. Data were normalized to ACT1 and expressed as means of 3 biological replicates relative to the corresponding strain cultured in C+I+, with the individual values indicated.
USP22 KO leads to increased TBZ-induced RIPK3 phosphorylation in HT‑29 cells A HT‑29 control and USP22 KO cells were stimulated with 20 µM zVAD.Figure 2A HT‑29 control and USP22 KO cells were stimulated with 20 µM zVAD.fmk, 0.5 µM BV6 and 1 ng/ml TNFα for the indicated time points. Detection of indicated proteins was carried out by Western blotting. GAPDH served as a loading control. B HT‑29 control and USP22 KO cells were stimulated with 20 µM zVAD.fmk, 0.5 µM BV6 and 1 ng/ml TNFα for 4 h. Detection of indicated proteins was carried out by Western blotting. β-Actin served as a loading control. C HT‑29 control and USP22 KO cells were incubated with 30 µM Nec1-s or 20 µM Dab for 18 h, as indicated. Cell were stimulated with 20 µM zVAD.fmk, 0.5 µM BV6 and 1 ng/ml TNFα for 5 h. 100 μg of each lysate were incubated with 400 U/μl λ-phosphatase for 30 min at 30 °C. Protein expression of RIPK3 was monitored by Western blotting. β-Actin was used as loading control. High molecular weight RIPK3 'smears' were quantified after λ-phosphatase treatment and normalized to total RIPK3 and β-Actin levels.
Ago1x does not interact with GW182 (A) Result of co-immunoprecipitation experiment in HeLa cells shows that GW182 is co-immunoprecipitated with Ago1, but not with Ago1x.Figure 6Result of co-immunoprecipitation experiment in HeLa cells shows that GW182 is co-immunoprecipitated with Ago1, but not with Ago1x. Confocal immunofluoresence images of HeLa cells show co-localization of GW182 with Ago1, but not with Ago1x. White circles indicate zoomed areas. Confocal microscopy images of proximity ligation assay (PLA) in HeLa cells showing interaction of GW182 with Ago1, but not with Ago1x. Rabbit anti-GW182 and Mouse anti-FLAG antibodies were used to achieve proximity ligation. Interaction is indicated by red dots that results from proximity ligation. Quantification of dots per cell is shown (right). Each black circle represents a microscopic field. Mean ± S.E. are shown as horizontal lines (n=10 fields for vector control; n=12 fields for Ago1; n=11 fields for Ago1x). At least 100 cells were counted for dots in each category. ***, P < 0.0001 (Mann-Whitney test).
Figure 5A,B Suppression of nse3-ts4 and nse4-ts4 temperature sensitivity (A) and DNA damage sensitivity (B) by the NSE1 allele of UWOPS87-2421. Cultures of the indicated strains were diluted to an optical density at 600 nm of 0.1 and a series of ten-fold dilutions was spotted on agar plates and incubated for 2-3 days. UW = UWOPS87-2421. The plates shown in (B) were incubated at 30°C. C C A cartoon of the SMC5/6 complex. D An illustration of the various types of genetic interactions that have been observed between different alleles of NSE1 and NSE3. E E Recruitment of Smc6-FLAG by ChIP-qPCR at two known SMC5/6 binding sites (pericentromere of chromosome XIV and CEN3) and one negative control locus (arm of chromosome III) in G2/M arrested strains. Relative enrichment corresponds to the ratio of the signal after immunoprecipitation (FLAG) over beads alone, normalised to the ratio in wild-type cells at CEN3. Error bars: standard error of the mean of three independent experiments. Statistical significance was determined using an unpaired, two-tailed t-test (* = p < 0.05, ** = p < 0.005). E
Figure 1Schematic representation of the generation of trastuzumab resistant cell lines. Low density cultures of BT474 (5,000 cells/150 mm dish) were incubated with trastuzumab (50 nM, replenished weekly) for 12 months and resistant pools and clones selected. Cell counting experiments analyzing the response to trastuzumab (50 nM, 5 days) of naïve BT474 as well as the resistant BTRH pool and the BTRH#10 clone. Western blot analyses of the expression of HER2, pHER2, HER3 and pHER3 in BT474, BTRH and BTRH#10 cells. HER2 and HER3 were immunoprecipitated from 100 or 500 μg of cell extracts, respectively, and blots probed with anti-HER2, anti-HER3 or anti-phosphotyrosine. Calnexin was used as a loading control. Numbers below the blots show the quantitation of the signal of each band, referred to BT474 cells (intensity of band in BT474 cells=1). Analyses of cell surface levels of HER2 and HER3 in BT474 and BTRH cells by surface immunoprecipitation. Response of BT474, BTRH and BTRH#10 to EV20/MMAF. Cells treated with the indicated doses of EV20/MMAF for 5 days were counted. Action of the nude EV20 antibody or EV20/MMAF (10 nM each, 5 days) on BT474, BTRH and BTRH#10 cells. Comparison of the effect of MMAF and EV20/MMAF on the proliferation of BT474 and BTRH cells after five days of treatment with the drugs.
Figure 4Immunoblot analysis of STAT4 phosphorylation in CD4+ T cells pretreated with PBS or LY2784544 (1μM) for 12 hours followed by LIF for the indicated number of minutes. Immunoblot analysis of CD4+ T cells treated with LIF for the indicated number of minutes followed by immunoprecipitation with an anti-LIFR antibody. Coimmunoprecipitation and immunoblot analysis of LIF-treated HEK293T cells cotransfected with the indicated constructs. The red asterisks denote phosphorylated STAT bands. Sequence alignment showing that the C terminus of STAT4 and LIFR contains four SP repeats in both humans and mice. The SPXX motifs are displayed and underlined in red. The red asterisks point out the potential phosphorylated serine residues. The mass spectrometry analysis of purified STAT4 protein revealed the phosphorylation of STAT4 at serine residues in the C terminus. Immunoblot analysis of the indicated protein modifications in the total lysate of CD4+ T cells treated with LIF (20 ng/ml) at different time points. Coimmunoprecipitation and immunoblot analysis of LIF-treated HEK293T cells cotransfected with the indicated constructs. Coimmunoprecipitation and immunoblot analysis of LIF-treated HEK293T cells cotransfected with the indicated constructs. Coimmunoprecipitation and immunoblot analysis of LIF-treated HEK293T cells cotransfected with the indicated constructs. Immunoblot analysis of the distribution of the indicated proteins in the CD4+ T cell fraction. Cultured HeLa cells transfected with GFP-tagged STAT constructs were exposed to LIF and imaged 30 minutes later. Scale bar, 50μm.
Figure 2 A) Plasma levels of selected candidate biomarkers analyzed in healthy subjects (n=131), pancreatitis patients (n=30) and PDAC patients (n=131). Data are expressed as a scatter-plot, mean ± SEM, *p<0.001 (Wilcoxon Rank Sum Test)  B) Receiver operator characteristic (ROC) curves of the single biomarkers and of biomarker panels (indicated with All) for diagnosis of PDAC vs healthy controls and PDAC vs pancreatitis. Areas under curve (AUC) with 99% CI are presented
Figure 2(A) Representative 2D class average of 300 x 300 pixel boxes covering ex vivo S-Enas and (B) the corresponding power spectrum of the 2D class. The Bessel orders used to derive helical symmetry are indicated. Reconstituted cryoEM electron potential map of ex vivo S-Ena (3.2 Å resolution). Side and top view of a single helical turn of the de novo-built 3D model of S-Ena shown in ribbon representation and molecular surface. Adjacent Ena subunits are labelled i, i-1, i-2 to i-10 (colored red to blue), where subunit i and i-10 represent the upper and lower subunit in a single helical turn. Based on the orientation and low-resolution features of 2D classes obtained on spore-associated S-Ena (Fig 1), subunits i and i-10 would be oriented distal and proximal to the spore body, respectively. Ribbon representation and topology diagram of the S-Ena1B subunit (blue to red rainbow from N- to C-terminus). The eight β-strands making up the S-Ena core domain are labelled B to I from N- to C-terminus. The first 15 residues form an N-terminal connector (Ntc) that for a subunit i, is in interaction with subunits i-9 (sand) and i-10 (green) through disulfide crosslinking.
p62/SQSTM1 targets L.Figure 6(A) Representative images from 6 h infected BMDMs with JR32 or legA9 mutant bacteria. P62 is stained red with p62 antibody. White arrows show L. pneumophila colocalization with p62. (B) Percent colocalization of L. pneumophila with p62. Hundred bacteria were scored from each coverslips. Images were captured with the 60× objective and magnified 3×, scale bar = 10 μm. (C)BMDMs were infected with JR32 or the legA9 mutant for 6 h and the level of p62 protein was quantified in cell lysates by ELISA (performed in triplicate and the SD are too small to detect). BMDMs were nucleofected with scrambled control (sictrl) or siRNA against p62 (sip 62). Twenty‐four hours after nucleofection, murine macrophages were infected with JR32 or legA9 mutant with an MOI of 0.5 for 1 h (D and F) or 24 h (E and G). (H) Western blot analysis showing the downregulation of p62 at 24 and 48 h postnucleofection. (A-B and D-H) Data are shown as mean + SD of n = 3 and are representative of three independent experiments. (C) Data are representative of one experiment n = 3. Asterisks indicate significant differences (*p < 0.05, **p < 0.01; two−tailed t−test).
The 18-month-old Pgam5 WT and KO mice received either water as control (n=4), or 5 mg kg−1 L-DOPA and 12.f6The 18-month-old Pgam5 WT and KO mice received either water as control (n=4), or 5 mg kg−1 L-DOPA and 12.5 mg kg−1 Benserazide (n=5) by intraperitoneal injection. Twenty minutes later, mice were subjected to the open-field test and the centre area time in seconds (s) as well as the centre area distance in centimeters (cm) were measured as described in the Materials and Methods. Graphs show the mean and s.d. values and were analysed by the student t-test, NS, no significance; *P0.05; **P0.01.