EMBO/sd-character-level-ner huggingface dataset:
GATA6 and its regulon are expressed in CDK7i-tolerant melanoma cellsA.Figure 3A. qRT-PCR analysis showing average TBP-normalized expression of GATA6 in the indicated cells. Data information: data are presented as mean values + SD for six replicates (n=6). The p-value (Student's t-test) is indicated, **<0.01, ***<0.005 and ns=non-significant B. Protein lysates from the indicated cells were immuno-blotted for the indicated proteins. Molecular sizes of the proteins are indicated in kDa. The numbers below the gel lanes represent relative protein level, which was determined from the band intensity using ImageJ software normalized relative to each relative ACTIN control. C. UMAP (Uniform Manifold Approximation and Projection) dimension reduction representative of GATA6 regulon expression in MM011, MM029, MM047, MM074 and MM099 from Wouters dataset UMAP is colored according to the continuous GATA6 AUCell values (from 0 to 0.2). D. qRT-PCR analysis showing average TBP-normalized expression of AMIGO2, SERPINE1 and ABCG2 in the indicated cells. Data information: data are presented as mean values + SD for six replicates (n=6). The p-value (Student's t-test) is indicated, **<0.01, ***<0.005 and ns=non-significant E-H. qRT-PCR analysis showing average TBP-normalized expression of GATA6 (E), AMIGO2 (F), SERPINE1 (G) and ABCG2 (H) in the indicated cells treated with either siCTL or siGATA6 for 72h. Data information: data are presented as mean values + SD for six replicates (n=6). The p-value (Student's t-test) is indicated, **<0.01, ***<0.005 and ns=non-significant I. MM011 and MM029 were treated with either siCTL or siGATA6 for 72h. Cell proliferation was analysed using CellTrace staining and flow cytometry in the indicated cell lines and the % of slow proliferating cells is shown for each condition. Data information: data are presented as mean values + SD for six replicates (n=6). The p-value (Student's t-test) is indicated, **<0.01, ***<0.005 and ns=non-significant J. MM029 (left) and MM099 (right) were pre-treated with either siCTL or siGATA6 for 48h and treated with increasing concentrations of THZ1 for 72h. Mean growth is shown relative to vehicle (DMSO)-treated cells. Data information: data are presented as mean values + SD for six replicates (n=6). The p-value (Student's t-test) is indicated, **<0.01, ***<0.005 and ns=non-significant
Figure 3Volcano plot representation of label-free quantitative proteomic analysis of P. falciparum proteins present in intron- (left) and promoter-targeted (right) dCas9 immunoprecipitations. Each dot represents a protein, and its size corresponds to the sum of peptides from both conditions used to quantify the ratio of enrichment. For the main volcano plot, x-axis = log2(fold-change), y-axis = -log10(p-value), horizontal yellow line indicates adjusted p-value = 0.05, and vertical yellow lines indicate absolute fold-change = 1.5. Side panels indicate proteins uniquely identified in either sample (y-axis = number of peptides per 100 amino acids). dCas9 protein is highlighted in green for reference. Two replicates were used for the promoter-targeted dCas9 and four replicates were used for the intron-targeted dCas9. Fold enrichment and adjusted p-values for each protein highlighted in red (promoter) and blue (intron) can be found in Tables EV5 and EV6, respectively. Correlation plot of label-free quantitative proteomic analysis of P. falciparum proteins enriched in promoter-targeted (top right quadrant in red) versus non-targeted control (top left quadrant in grey) or intron- (bottom right quadrant in blue) dCas9 immunoprecipitations. Each dot represents a protein, and its size corresponds to the sum of peptides from both conditions used to quantify the ratio of enrichment. The x-axis = log2[fold-change(promoter/non-targeted control)] and y-axis = log2[fold-change(promoter/intron)]. dCas9 protein is highlighted in green for reference. Two replicates were used for the promoter-targeted dCas9 and four replicates were used for the intron-targeted and non-targeted dCas9. Fold enrichment and adjusted p-values can be found for each protein highlighted in red (promoter) in Tables EV5 (intron comparison) and EV7 (control comparison) and for each protein highlighted in blue (intron) in Tables EV6 (promoter comparison) and EV8 (control comparison). Venn diagram comparing proteins enriched in the ISWI IP LC-MS/MS (red) and the promoter-targeted dCas9 IP LC-MS/MS compared to the intron-targeted dCas9 (blue) or the non-targeted dCas9 (yellow). Schematic of proteins shared between the ISWI IP LC-MS/MS and the promoter-targeted dCas9 IP LC-MS/MS (overlapping regions shown in Fig 3C): ISWI (PF3D7_0624600), MORC (PF3D7_1468100), ApiAP2 (PF3D7_1107800), MCM2-4,6,7 (PF3D7_1417800, PF3D7_0527000, PF3D7_1317100, PF3D7_1355100, PF3D7_0705400), and acetyl-CoA synthetase (ACS, PF3D7_0627800).
miR‐290 cluster enhances glycolysis by upregulating Pkm2 and Ldha in ESCs A–C Screening for miRNA clusters that rescue the cellular metabolic phenotype of Dgcr8−/− ESCs.Figure 2A-C Screening for miRNA clusters that rescue the cellular metabolic phenotype of Dgcr8−/− ESCs. Mimics of control oligo (CTR), miR‐290 cluster (290), miR‐302 cluster (302), miR‐106b cluster (106b), and miR‐17 cluster (17) were introduced in Dgcr8−/− ESCs, and cellular glucose uptake (A), lactate production (B), and oxygen consumption rates (C) were measured using specified assay kit. D qRT-PCR analysis of metabolic enzymes in Dgcr8−/− ESCs upon introduction of miR‐290 cluster. E Western blotting analysis of metabolic enzymes in Dgcr8−/− ESCs transfected with mimics of miR‐290 cluster. F Cellular glucose uptake, lactate production, and oxygen consumption rates were measured in WT and miR‐290 cluster−/−ESCs. G qRT-PCR and Western blotting analysis of Hk2, Pkm2, and Ldha in WT and miR‐290 cluster−/−ESCs. H Cellular glucose uptake and lactate production were measured in Dgcr8−/− ESCs expressing Pkm2 shRNAs and further infected with miR‐290 cluster. I Cellular glucose uptake and lactate production in medium were measured in Dgcr8−/− ESCs expressing Ldha shRNAs and further infected with miR‐290 cluster.
Figure 2(A) The classic model of HPA axis dynamics without gland mass dynamics produces elevated levels of stress hormones during prolonged stress. However, it does not produce blunted ACTH responses in the CRH test, and, after cessation of the stressor, all hormones return to baseline within hours. Data information: In all panels, simulations are of a CRH test (Methods), where 'case' (black) is after stress and 'control' (gray) is after basal HPA axis activation (B) To account for the control of corticotroph growth by CRH and adrenal cortex growth by ACTH, we added to classic HPA model two equations that represent the dynamics of the functional mass of corticotrophs (C) and the adrenal cortex (A). (Inset) Such dynamics explain, for example, the enlarged adrenals of stressed rats (inset, right) compared with control (inset, left), adapted from (Selye, 1952). The model shows the three distinct phases of HPA axis dysregulation observed in experiments. At the end of a prolonged stress period, the adrenal mass is enlarged and the corticotroph mass is slightly enlarged, which results in hypercortisolemia and blunted ACTH responses in the CRH test. After a few weeks, corticotroph mass drops below baseline, while adrenal mass is slightly enlarged, causing normal cortisol dynamics with blunted ACTH responses to the CRH test. Finally, after a few months both tissue masses return to normal, leading to normalization of both cortisol and ACTH dynamics. Data information: In all panels, simulations are of a CRH test (Methods), where 'case' (black) is after stress and 'control' (gray) is after basal HPA axis activation
Figure 6Representative negative stain images of endospores of NVH 0075-95 mutants lacking ena1A, ena1B, or ena1C, as well as endospores of strains complemented with the respective ena subunit expressed from plasmid (i.e. pA, pAB and pC). The ena1B mutant was complemented with a plasmid carrying both ena1A and 1B (pAB) due to repeated failure to transform with a plasmid holding ena1B only. Inset are 2D class averages of Enas observed on the respective mutants. Knockout of ena1A, ena1B, or ena1C results in the loss of S-Ena, a phenotype that is restored by plasmid-based complementation. Number (top) and length (bottom) of Enas found on WT, mutant (∆ena1A, ∆ena1B, or ∆ena1C) and plasmid complemented (∆ena1A:pena1A, ∆ena1B:pena1AB, or ∆ena1C:pena1C) NVH 0075-95 endospores. Statistics: pair-wise Mann-Whitney U tests against WT (n: ≥18 spores; n: ≥50 Enas; ns: not significant, * p<0.05, ** p<0.01, *** p<0.001 and **** p<0.0001. ---: mean ± s.d.)
Figure 3Percentage of body weight that is taken up by the inguinal fat pad (iWAT, A), the mesenteric fat pad (mWAT, B) and the perirenal fat pad (pWAT, C) 24h post-surgery in sham (with or without starvation) and CLP mice. P-values were calculated with a 1-way ANOVA test. Combined data of 2 independent experiments, n = 6/group. Visual representation of (D) iWAT and (E) pWAT sizes in sham, sham starved and CLP mice 24h post-surgery. Data are representative of 3 independent experiments. Mice (n=6-10/group) underwent a sham (with or without starvation) or CLP operation and blood was collected 6h and 24h post-surgery. Plasma was isolated and (F) FFA concentration and (G) glycerol concentration was determined as described in the method section. P-values were calculated with 2-way ANOVA tests. Combined data of 2 independent experiments. Mice (n=6-10/group) underwent a sham (with or without starvation) or CLP operation and blood was collected 6h and 24h post-surgery. Normalized abundances of (H) palmitic acid and (I) palmitoyl-carnitine determined via liquid chromotography-mass spectrometric lipidomics. Values were normalized to IQ values , P-values were calculated with 2-way ANOVA tests. n = 8/group.
Down-regulation of BACE1 protein in aged Mgat3−/− mouse brain.Figure 6Immunostaining of 12-month-old brain sections (cerebral cortex) with anti-BACE1. Scale bar, 100 μm. Proteins from the membrane fractions of 12-month-old mouse brains (n = 4-5) were immunoblotted for BACE1, and BACE1 intensity was quantified. *P = 0.044. Lysates from MEFs were immunoblotted for APP, BACE1, or GAPDH (loading control), and BACE1 intensity was quantified (n = 4). Amount of Aβ40 in the culture medium of immortalized MEFs (n = 3). *P = 0.045. MEFs were transfected with control siRNA or GGA3-siRNA and then immunoblotted for BACE1, GGA3, or GAPDH (loading control) (n = 6).
a, b) Bif-1+/+ and Bif-1−/− MEFs or Bax+/+Bak+/+ and Bax−/−Bak−/− MEFs were infected with Beclin 1 shRNA or control shRNA (SCR shRNA) lentiviruses.figf2(a, b) Bif-1+/+ and Bif-1−/− MEFs or Bax+/+Bak+/+ and Bax−/−Bak−/− MEFs were infected with Beclin 1 shRNA or control shRNA (SCR shRNA) lentiviruses. Twenty-four hours after infection, the medium was replaced with fresh culture medium and the cells were allowed to recover for an additional 48 h. The cells were then cultured in EBSS for 0, 6 or 12 h, and the percentage of dead cells was determined by trypan blue exclusion assay (mean ± s.d.; n = 3). Knockdown of Beclin 1 expression was confirmed by immunoblot analysis (insets). (c, d) Bif-1+/+ and Bif-1−/− MEFs were pre-treated with 10 mM 3-methyladenine (3-MA), 0.2 μM wortmannin (WM) or control DMSO for 30 min followed by culture in EBSS for the indicated times. The percentage of dead cells was determined by trypan blue exclusion assay (mean ± s.d.; n = 3). (c, d) Bif-1+/+ and Bif-1−/− MEFs were pre-treated with 10 mM 3-methyladenine (3-MA), 0.2 μM wortmannin (WM) or control DMSO for 30 min followed by culture in EBSS for the indicated times. The percentage of dead cells was determined by trypan blue exclusion assay (mean ± s.d.; n = 3). Immunoblot analyses were performed to detect LC3 modification and caspase-3 activation. (e) Atg5+/+ and Atg5−/− MEFs were cultured in EBSS in the presence of 50 μM of z-VAD-fmk or control DMSO for 0, 6 or 12 h. The percentage of dead cells was determined by trypan blue exclusion assay (mean ± s.d.; n = 3). (f) Total cell lysates (TCL) were prepared from Atg5+/+ and Atg5−/− MEFs cultured in EBSS for the indicated times and subjected to immunoblot analyses. Uncropped images of the immunoblots in a, b, d and f are shown in the Supplementary Information, Fig. S5.
Expression of HMGB1 or the C106S mutant, but not C23S and C45S mutants, restore autophagic flux in Hmgb1−/− MEFs.Figure 7.Expression of HMGB1 or the C106S mutant, but not C23S and C45S mutants, restore autophagic flux in Hmgb1−/− MEFs. (A) HMGB1−/− MEFs were transfected with wild-type (WT) or the cysteine mutant HMGB1-GFP plasmids as indicated, starved (HBSS) for 3 h in the presence or absence of 100 nM bafilomycin A1, and then immunostained with LAMP2-specific antibody/Alexa Fluor 594 secondary antibody (shown in red), LC3-specific antibody/Alexa Fluor 647 secondary antibody (shown in green), and Hoechst 33342 (shown in blue). Images were acquired digitally from a randomly selected pool of 10-15 fields under each condition. (B) Quantitative analysis of the percentage of LAMP2/LC3 colocalization was detected by Image-Pro Plus 5.1 software. Data are means ± SEM.
Fig.Figure 4TSS enrichment score of primary samples (PRI) and PDXs Reads number per peak in primary samples (x-axis) plotted against the reads number in corresponding PDX model; PCC=Pearson's correlation coefficient) Fraction of peaks shared between PDX and corresponding primary leukemia for ATAC peaks with a certain height (x-axis; average number of reads that map into an accessible region called by Macs2) Unsupervised learning by principal component analysis (PCA), circle - primary sample (PRI), triangle - PDX sample; each color corresponds to 1 patient MA plot of log2 fold changes between primary and PDX samples plotted against the mean of normalized read counts per peak, red - differentially accessible regions (p<0.05) as analyzed by DESeq GO term enrichment of the predicted cis-regulatory regions downregulated in PDX in comparison to primary samples performed by Genomic Regions Enrichment of Annotations Tool (GREAT)
Figure 1Figure 1. MLL-ENL leukemic stem cells derived from LSK or GMP populations have contrasting functional requirements of β-catenin for their initiation and maintenance of disease. (A) Schematic overview of the experimental procedures. Keys and colour codes in the legend box indicate the cells-of-origin and the -catenin status of MLL-ENL transduced cells in the following experiments (B-K). (B) Colony numbers in serial replating assay of the different MLL-ENL transduced cells. Data are represented as mean ± SD (C, G) PCR validation of Ctnnb1 deletion on genomic DNA isolated from the indicated MLL-ENL transduced GMP and LSK cells (C) L, 100bp ladder; W, wild type control, F, Ctnnb1 floxed allele, D, Ctnnb1 deleted allele, N, negative control. (D, H) Cell lysates from indicated MLL-ENL transduced GMP and LSK cells after the fourth round of plating (D) were blotted with anti-β-catenin (top) and anti-actin (bottom) antibodies. (E, F) Kaplan-Meier survival curves of indicated MLL-ENL transduced cells transplanted into primary recipient (solid lines) and secondary recipient mice (dotted lines). N=10 mice per group were used in primary transplants (solid lines) in (E).For all secondary transplants (dotted lines) n=5 mice were used per group. (E, F) Kaplan-Meier survival curves of indicated MLL-ENL transduced cells transplanted into primary recipient (solid lines) and secondary recipient mice (dotted lines).N=15 mice were used for Ctnnb1fl/fl (blue solid line) and n=10 mice were used for Ctnnb1del/del (green solid line) in (F). For all secondary transplants (dotted lines) n=5 mice were used per group. (C, G) PCR validation of Ctnnb1 deletion on genomic DNA isolated from the indicated MLL-ENL transduced GMP and leukemic cells (G). L, 100bp ladder; W, wild type control, F, Ctnnb1 floxed allele, D, Ctnnb1 deleted allele, N, negative control. (D, H) Cell lysates from indicated MLL-ENL transduced GMP and from indicated leukemic mice (H) were blotted with anti-β-catenin (top) and anti-actin (bottom) antibodies. (I) Percentage of CD45.2+ donor cells in the bone marrow of recipient mice at the indicated time points post-transplantation of the indicated GMP-MLL-ENL and LSK-MLL-ENL transduced cells. Data are represented as mean ± SD. N=4, two-tailed t-test was performed. (J-K) Kaplan-Meier survival curves of secondary transplanted GMP-MLL-ENL (J) primary leukemia bone marrow cells treated with DMSO as floxed controls or tamoxifen for β-catenin deletion prior to secondary transplantation (n=3 per group). See also Appendix figure S1. (J-K) Kaplan-Meier survival curves of secondary transplanted LSK-MLL-ENL (K) primary leukemia bone marrow cells treated with DMSO as floxed controls or tamoxifen for β-catenin deletion prior to secondary transplantation (n=3 per group). See also Appendix figure S1
Figure 6Representative images of pADAM17-stained lung sections from normal (cancer-free) control and LAC patients. Scale bar, 100μm. Quantification of pADAM17-positive cells in (B) normal (cancer-free) control individuals (n = 8) and LAC patients (n = 18) Quantification of pADAM17-positive cells and pERK1/2 MAPK-positive cells in LAC patients either KRAS wild-type (WT, n = 13) or mutant (MUT, n = 5). Linear regression analyses of pADAM17 with pERK1/2 (D) and pp38 (E) MAPK staining in serial lung sections from LAC KRAS wild-type and mutant patients. Immunoblots of lysates from normal human bronchial epithelial cells (HBEC) and LAC cell lines A549, NIH-H23 and NIH-H2228 with the indicated antibodies. Shown are lysates from 2 separate passages per cell line. Pro, proADAM17; Mat, mature ADAM17. Densitometric semi-quantification of immunoblots showing the relative expression levels of pADAM17 (relative to Actin), pro and mature forms of ADAM17 (relative to Actin), and pERK1/2 and pp38 MAPK (relative to their total counterparts) protein levels. ELISA of sIL-6R protein levels in culture supernatants from matching cells in (F) cultured over 24 hours. Data are from 4 independent experiments. MTT viability assays of A549 cells transduced with nontargeted control sgRNA (Ctl) and ADAM17 sgRNA (knockout, KO). ATP proliferation (J) assays of A549 cells transduced with nontargeted control sgRNA (Ctl) and ADAM17 sgRNA (knockout, KO). ELISA of sIL-6R protein levels in culture supernatants from matching A549 cells (I and J) cultured over 24 hours. Data are from 3 independent experiments. Tumor volume (mm3) of KO and Ctl A549 xenografts assessed every 3 days starting at one week post injection of 106 cells into NSG mice (n = 5 per group). Weight (grams) (M) of tumor xenografts representative images (N) of tumor xenografts ELISA of human sIL-6R protein levels in serum of the indicated mice (n = 5 per group). Levels of sIL-6R are below detection in serum from KO xenograft mice. ND, not detected. Representative images of pADAM17 and pERK1/2 MAPK immunohistochemical staining on KO and Ctl A549 xenograft tumors. Scale bars, 100μm. Immunoblots of lysates from individual A549 KO and Ctl xenograft tumors with the indicated antibodies.
Figure 7A) Enriched motifs defined by HOMER using a de novo motif search of the digital DNase I FPs identified in the pDHSs in TB. B) DNase I cleavage patterns in TB from the FPs determined by Wellington at the pDHSs centered on the motif named at the top and ordered according to increasing FP occupancy score. Left: Cuts are shown within a 200 bp window with positive (red) and negative (green) strand imbalances in DNase I cuts. Right: Average profiles of the actual DNase I cuts at footprinted motifs within the pDHSs, with upper strand DNA cuts shown in red and lower strand cuts in blue. C) Average profiles of the DNase I cuts at footprinted motifs within the pDHSs in TM determined as in (B). D) Example of FP patterns and motifs at the -3.7 and -35 kb pDHSs at the Ccl1 locus in TB. E) mRNA array values for Ccl1 expression. F) Luciferase reporter gene assays in stimulated Jurkat T cells performed as in Fig 1H of the IL3 promoter alone or in combination with the Ccl1 -3.7 or -35 kb DHSs, with SD. G) Footprinting of AP-1 sites in TB+. DNase I upper and lower strand cleavage patterns were calculated as in (B) (left) plus the average DNase I profiles (right) for all AP-1 motifs within the subset of 2882 defined pDHSs in TB cells before and after stimulation, ranked in order of decreasing FP probability score. H) Distribution of the FP probability scores for the data shown in (G).
Figure 8Initial stages of LSD pathology show direct lipid storage-induced neuronal degeneration, demyelination and myelin debris (shown in dark blue). These events activate resting microglia, which polarise into CD16/CD32 and Arg-1 expressing phenotypes. General microglia ablation causes myelin debris accumulation, which enhances neuronal death. CD16/CD32 expressing microglia promote increased levels of cytokines like TNFa, which could amplify neuronal death. Arg-1 expressing microglia phagocytose and remove myelin debris protecting neurons from their toxic effects (Protective Phase). Continued lipid overload of lysosomes, due to the genetic defects and myelin debris accumulation, causes lysosomal damage and activates lysosomal exocytosis in microglia. This process stimulates extracellular release of CathB (shown in red), which directly causes neuronal death (Neurotoxic Phase).
Figure 2A Diagram of the NB-3 induction (green) after SCI. B NB-3 expression in corticospinal neurons 14 and 28 days post-injury (dpi). FG (blue) was applied to retrogradely label corticospinal neurons.C Quantification of the fluorescence intensity of NB-3 in (B). **p < 0.01; one-way ANOVA followed by Bonferroni's post-test. The intensities of more than 300 corticospinal neurons from 3 mice in each group were quantified. D, E Western blot analysis (D) and quantification (E) of NB-3 expression in the sensorimotor cortex after SCI. NB-3 expression levels were normalized by b-tubulin. *p < 0.05, and **p < 0.01; one-way ANOVA followed by Bonferroni's post-test. Data were analyzed from 3 independent experiments each including 3 mice per group. F-K Induced NB-3 expression at the distal terminations of post-traumatic corticospinal axons and in reactive astrocytes at the lesion border 14 (H and I) and 28 days post-injury (dpi) (J and K). BDA (red) was applied to anterogradely label corticospinal axons. GFAP (blue) was used to label reactive astrocytes. (G) High-magnification image of the boxed area in (F). (I and K) High-magnification and x-y-z reconstruction images of boxed areas in (H and J), respectively, show co-localization of NB-3 and GFAP in astrocytes at the lesion border and co-localization of NB-3 in BDA-labeled axons and GFAP-positive astrocytes at the contact surface of corticospinal axons and astrocytes (arrowheads). The arrows in (H and J) indicate the lesion sites. The asterisks in (H and J) indicate the terminations of the BDA-labeled projecting axons.