EMBO/sd-character-level-ner huggingface dataset:
(B) Gene correlation plot of average beta scores for lipidation dependent autophagy (tf-NDP52, from (Shoemaker et al, 2019)) and lipidation independent autophagy (average of tf-NBR1 from ATG7KO, ATG10 KO and ATG3 KO cells Highlighted in red are genes with a beta score > 0.5 across all three lipidation-deficient cell lines. In black are lipidation components. Dashed lines, top 1% of beta scores.
(A) Simultaneous imaging of Gal4 binding kinetics in vivo using single-molecule tracking and RNA imaging of the GAL10 target gene. Gal4 is tagged with a HALO-tag, which covalently binds to the dye JF646. Transcription of the target gene GAL10 is visualized by PP7-loops.
G Superoxide levels in response to Hcy treatment (20 μM, 4 h) in MARS-expressing and control cells were determined relative to the untreated NE4C cells.
(E) Internalization of soluble Fab or F(ab')2 fragments in Ramos cells targeted with indicated sgRNAs. *p=0.0124 **p=0.0092 ***p=0.0004 using 2-way ANOVA. Data show mean ± SEM, from 6 biological replicates across 2 experiments.
H Quantification of the mean cell velocity of shIP3R(1,3) C (green)‐silenced immature DCs migrating in micro‐channels in the presence of 2 mM BAPTA. shScramble‐infected DCs were used as a control (gray) (n > 70 cells from two independent experiments). P‐values were calculated using a Kruskal-Wallis test.
(E,F) PTPRN2 (E) and PLCB1 (F) levels were analyzed in human breast cancers (stages I-IV) and normal breast tissue from TissueScan qPCR Array Breast Cancer Panels II and III (Origene, N = 97). Expression levels were normalized to levels in normal tissue for each gene.
EFS and tumor volume in non-CNS subcutaneous PDX models which demonstrated objective response in 1 or more treatments.
B C, Wild-type CNNM4-dependent magnesium efflux is inhibited by co-transfection by PRL3 but the efflux by the CNNM4 D485A mutant, which is unable to bind PRL3, is resistant to inhibition. HEK293 cells transfected with the indicated constructs were loaded with Magnesium Green and Mg2+ removed from the medium at the indicated time point (arrowhead). The means of relative fluorescence intensities of 10 cells are graphed as a function of time in panel B and the mean and s.e.m. at 5 min plotted in panel C. P-values were calculated using one way ANOVA followed by Bonferroni's multiple comparison test. n.s., not significant.
C Immunoprecipitation using an anti‐secretagogin antibody in Ca2+‐free and Ca2+‐loaded conditions was subtractively used to decipher the Ca2+‐dependent protein interactions. Silver‐staining gel is shown.C1 Ontology classification of the 99 protein hits based on primary function assignments. Unbiased MALDI‐TOF proteomics was used to identify interacting proteins. The most abundant hits (Supplementary Table S3 is referred to for details on individual proteins) are proteins implicated in vesicle fusion, trafficking, transport and formation and the regulation of vesicle exocytosis. "Other" refers to a group of proteins without known function.
Kinetics of p53 degradation (a) and induction of GFP-LC3 puncta (b) in WT HCT116 cells cultured in complete medium in the presence of rapamycin (Rapa), tunicamycin (Tn), lithium (Li) or in nutrient-free medium (NF).
C) Bacterial uptake by MoDCs was determined by CFSE fluorescent intensity (* p<0.001, # p<0.01).
Activation of WOX5 and SCR by DEX treatment of 35S::GVG-WOX5 UBI::mCHERRY-GR-SCR in Col and hag1-6.
Representative MS2 spectra of the CO1A1 peptide in ECM generated with unlabeled ([12C] Gln, left) and fully labeled glutamine ([U-13C] Gln, right). Peaks representing fully labeled (m+5, m+10) proline are highlighted in red. Peptides were measured by LC-MS.
F. NCI-H69-CRISPR/Cas9 cells presented in E. were subcutaneously engrafted with either NCI-H69- CRIPSR/cas9-NTN3 with three different guide RNAs (sG1 p=0.017; sG2 p=0.028; and sG3 p=0.001) or control cells (sGc) n=6/group. Positive tumor engraftment was report when tumors reached 20 mm3 (Mantel Cox).
(F) Plasmids expressing HP1c CSD domain and Su(H) BTD domain were transfected into S2 cells. Immunoprecipitation and western blotting were performed with the indicated antibodies.
Venn diagram depicting the common interacting proteins between different tsRNAs in RA treated cells.
E. Reduced levels of fasting circulating insulin in 6-10 month old diabetic Rb∆K7 mice compared with control wild type littermates (0.13 vs 0.2 ng/ml; n=8 for each group). P value was calculated by one-tailed unpaired student's t-test. Bars represent mean ±SE. Average levels of blood glucose are indicated (17 mM in Rb∆K7 compared with 3.2 in control mice; P=0.0001 by two-tailed unpaired student's t-test).
B, C HeLa cells were treated with the indicated siRNAs, synchronized by double thymidine block, released for 12 hours and analysed by immunofluorescence microscopy. Arrowheads indicate blebbed regions of nuclei. The percentage of interphasic cells with Nup granules was quantified (C), 900 cells were analysed (mean ±SD, ***P < 0.001; N = 3).
b-g, Ten days after TAC. c, Echocardiography. Scale bars, 0.2 s and 5 mm. Echocardiographic parameters (d) (n = 7-13 per group). LVIDd and LVIDs, end-diastolic and end-systolic left ventricular internal dimension, respectively; LVFS, left venticular fractional shortening;
(B) Specific ligand binding assay in COS cells with overload of cold ligand and increasing concentrations of radioactive labeled DHT. Values are normalized to protein expression.
C SETDB1 cellular amount is mostly maintained in Atf7ip KO mESCs. Endogenous SETDB1-ATF7IP interaction is validated by anti-ATF7IP antibody co-IP experiment with anti-ATF7IP antibody.
(m) L. monocytogenes in spleen and liver of L2A-cKO mice and their control littermates (n = 3 per genotype) immunized with L. monocytogenes and then reinfected with a high dose of L. monocytogenes. CFU, colony-forming units. *P = 0.008 and **P = 0.001 (t-test).
Representative FACS plots showing of red blood cell maturation as determined by CD71/Ter119 staining in single dox-treated E10.5 control (left) and chimeric (right) embryos.
Colonic lesions were scored from H&amp;amp;E by identification of disorganised epithelium. Staining for β-catenin and SOX9 confirmed activation of Wnt signalling in the observed lesions. Scale bar, 100 μm.
A structure of Ubc1 was superimposed onto the model Key residues were highlighted as sticks for the binding interfaces (E) between the UBC domain and UBA domain. Additionally, amino acids whose side chains contribute to the UBC/UBA binding interface were determined and classified into "hot spot" or "contributing" residues using the algorithm SpotOn
Proliferation assay of CrizR1, CrizR4, CrizR5, CeritR isogenic drug-resistant cell lines, treated with the indicated concentrations of alvocidib(G) for 72h. P >0.05 was calculated for IC50 shift, as indicated in the methods.
(a) AMPK wild-type or DKO MEFs were starved of glucose (4 h) as indicated. Total cell lysates were probed for Ulk1 protein and phosphorylation.
B. Quantification of the number of CETN3 foci per spermatocyte (n = 30) at each meiotic prophase substage.
Distributions (quantified by kernel density estimation) of p-values for differential expression (shERα vs. shCtrl BM67 cells) using data from polysome-associated mRNA (orange), cytosolic mRNA (purple) or from analysis of changes in translational efficiency (i.e. changes in polysome-associated mRNA not paralleled by changes in cytosolic mRNA) leading to altered protein levels (red; a higher density of low p-values indicates a higher frequency of changes in gene expression).
(B and C) Mutation of Tau40 at acetyltransferase activity region abolishes its acetylation activity on β-cat. eGFP-Tau40, eGFP-Tau-2CA and eGFP-Tau-k18(-) were purified from HEK293 cells (using anti-GFP beads) and GST-β-cat was from E. coli (B), or both Tau40 and β-cat were from E. coli (C) (n = 3 from three independent experiments).
B. A heatmap showing changes in indicated cytokines in response to 500 nM treatment of PBMCs with 35 "tool" kinase inhibitors as the training set. Cytokines released in the media were measured by Luminex and normalized to DMSO control.
Expression patterns of Hoxa5 and Hoxc8 mRNAs, as revealed by in situ hybridization, aligned with corresponding sections showing Hoxa5 and Hoxc8 immunostainings in MNs (demarcated with pink/white dashed lines) along the cervical spinal cord (cervical C5 to C7) in mouse embryos at E12.5. Pink scale bar, 50 μm. White and pink dash lines demarcate the postmitotic MN region.
(f) HEK293 cells were transfected with ULK1 and Beclin-1 in conjunction with either ATG14L WT, or one of two mutants (ΔBATS,ΔN) that are defective in phagophore localization. Samples were handled as in e. Uncropped images of blots are shown in Supplementary Fig. S4. Lysates were resolved by SDS-PAGE and blotted with the indicated antibodies.
(A, B) In vitro degradation of model proteins by purified S. cerevisiae proteasome. The model proteins (UbL-DHFR-95) consisted of an N-terminal UbL domain, followed by an E. coli DHFR domain and a 95 aminoacid tail derived from S. cerevisiae cytochrome b2. UbL-DHFR-95 proteins were degraded by proteasome (red solid circle) and can be stabilized by removing the UbL domain (no UbL, black triangles), by removing the 95-amino-acid tail (no tail, blue diamonds) or by proteasome inhibitor (MG132, green squares). (A) Degradation of model proteins containing the UbL domain of S. cerevisiae Rad23.
Time-lapse micrographs of agarose-embedded cells were taken at RT with fresh PM Glc+ura medium supply throughout one budding cycle. Representative images of characteristically reoccurring fluorescence localization stages are displayed. In all combinations, fluorescence foci localize to areas of membrane outgrowth in the bud. Often, another prominent fluorescent spot is found in the mother cell on the side of the bud neck. Scale bar 5 µm. Kymographs of all time-lapse datasets shown in this Figure are presented in Fig EV3A.I. β'-COPmGFPfluorescence foci in Dsl1p-depleted GAL-DSL1 cells after incubation in glucose-containing medium (YEPD) for 4 h at 30 °C.
a, Skeletal muscle. Muscle extracts from patients 1-6 and 10 and a control were blotted and labelled with antibodies against LAMP-2 (top panel) and β-actin (bottom panel). LAMP-2 is deficient in all muscles examined except the sample from patient 1 that showed a trace amount of protein, which is probably the LAMP-2a isoform. In support of this idea, LAMP-2b is the principal LAMP-2 species in skeletal muscle6. Numbers denote patients as in Table 1; C, control. b, Heart. Protein was extracted from cardiac muscle of patient 4 and a control, blotted onto a nitrocellulose membrane, and labelled with the antibodies against LAMP-2 (too panel) and β-actin (bottom panel). LAMP-2 is deficient also in cardiac muscle. C, control; 4, patient 4.
(B) mRNA-tRNA translocation monitored in a time-resolved Pmn assay upon mixing 80S 2C with Pmn in the absence (brown triangles) or presence of eEF2 and eEF3 (green, 0.13±0.01 min-1), eEF2 (red, 0.030±0.005 min-1) or eEF3 (blue, 0.10±0.01 min-1). Data are normalized to Met-Phe-Pmn formation in presence of eEF2 with the maximum value set to 1. Data presented as mean ± SEM of n = 3.
(C) Average fEPSP amplitude values at individual acquisition points between min 75 and 90 of the recording (vertical dashed lines in B) across experimental groups. Individual symbols represent the averaged fEPSP amplitude measured in one animal (2 slices), with bars representing average ± SEM across all slices.
Plasma insulin levels after a 5-hour fast, before injecting insulin (n = 11-13 animals per group).
D) N-specific antibody ELISA was used to compare antibody responses in sera of WT and KO mice.
(F) FAK−/− PEMs were incubated with DQ-BSA-red for 1 hour before infection with S. typhimurium strain ΔinvG for 5 h. Cells were co-stained for LC3 and DAPI was used to visualize nuclei and bacteria. White boxes show enlarged regions in inset panel.
C. Enhanced antibody production following immunisation with cGAMP-VLPs relies on STING signalling. WT or Sting-/- mice were immunised with PBS, cGAMP-VLPs, Empty-VLPs or Empty-VLPs + poly(I:C). IgG2b antibodies recognising VLP proteins were assessed by ELISA. The optical density at increasing serum dilutions is shown in the first two graphs from the left and the EC50 is on the right.
M2-induced human monocyte-derived macrophages were treated with 100 nM of the indicated drug either continuously for 48h, or initially for 2h in the presence or absence of FA-glucosamine (competition) followed by 46h in the absence of drug (2+46h), as described in Fig 1. mRNA levels of profibrotic markers, Arg1 (A), CD206 (B) and CD163 (C) Changes in both sets of cytokines were inhibited by blockade of unoccupied folate receptors with excess FA-glucosamine (2+46h, competition).
(B) Isolation of GFP-positive cells by FACS: schematic diagram showing the selection of the GFP-positive population (green).
A FAK-WT, interface mutant FAK or FAK -/- SCC cells were resuspended in methylcellulose solution in growth medium on a layer of agarose and observed for 3D sphere formation. Images were taken from 10 random fields after nine days (representative images are shown) and the relative colony size (top graph) as well as the number of colonies (bottom graph) were assessed from three independent experiments. Scale bars, 200 μm. Student's t-test was carried out to calculate the statistical significance. Error bars s.d. * = p < 0.01, # = p < 0.05.
J, qRT-PCR analysis of HIF-1 target gene expression in VZ cells from the cortex of control and Gpr124KO E13.5 embryos (mean±SEM; N=4; *** p<0.001). SVZ, subventricular zone; VZ, ventricular zone.
H No ternary complex is observed when CaM and caldendrin are mixed together with excess myoVa peptide in native MS. The individual proteins bind individually to myoVa peptide indicating competition for the same or closely located epitope. Fraction of caldendrin degrades into a 28kDa fragment, which has lost it affinity towards myoVa peptide almost completely.
The diffusion times of Scc1-GFP at S phase (B) were measured in strains yIO664 (wild-type) and yEB005 (WPL1∆), and in auxin-depleted yEB002 (PDS5-AID), yAM085 (SCC3-AID) and yAM946 (ECO1-AID) cells. Data are shown as mean ± s.e.m. from at least three independent experiments. *: p< 10-5, **: p< 10-13.
(c) Autophagic flux was analysed by immunoblotting with anti-LC3B and anti-p62 antibodies.
Differential interaction heatmap for chromosome 3L (40k resolution), showing increased (red) and decreased (blue) interacting bins in log2 fold change upon Piwi-KD.
(A) Western blotting of AXL and OPCML protein levels in AXL-expressing, OPCML-null SKOV3 and PEA1 cell lines transduced with OPCML "O" or control Empty vector "E"
Agarose gel electrophoresis of LOR, FLG, and LCE1B promoter fragments amplified by end-point PCR after ACTL6A and BRM (left), or BRG1 (right) chromatin IP in proliferating keratinocytes.
(D) Fluorescence micrographs of A549 cells of the indicated genotypes. Cells were either left untransduced (mock) or transduced with PIDD1-V5 lentiviral vectors carrying the indicated point mutations, and stained with the indicated antibodies. Blow-ups without Hoechst 33342 are magnified 2.5X. Scale bar: 5 μm.
C. Silver staining of protein components in the Vret complex The lane α-Vret (left) contained only the anti-Vret antibodies used in these experiments.
(d-i) CftrF508del mice treated with cystamine or PBS in the presence or absence of 3-MA (e-i) (n = 7 per group). (f) Immunoblot of TG2. Uncropped images of blots are shown in Supplementary Information, Fig. S10.
Hoxa5 expression upon miR-27 knockdown, with exposure to RAhi for 72 h, or with exposure to RAhi for 48 h, and then switching to RAlow plus RAR inhibitor (1 µM) for a further 24 h, or with exposure to RAmed for 72 h.
H A working model of the miR‐290‐Mbd2‐Myc axis in regulating metabolism and reprogramming. miR‐290/371 cluster posttranscriptionally represses Mbd2, leading to the downregulation of MBD2 protein and reactivation of Mbd2 target gene Myc. Subsequently, Myc activates glycolysis through directly stimulating the transcription of glycolytic enzymes Pkm2 and Ldha. This regulatory circuit orchestrated by miRNAs facilitates metabolic switch in reprogramming and enhances glycolysis in ESCs.
Flow cytometric analysis of splenic GC B cells in Prdm1ihCd2/+ (red) and wild-type (gray) mice (at the age of 2 months) at day 14 after NP-KLH immunization. Bar graphs display absolute numbers of GC B cells detected in the spleen of both genotype The absence of Bcl6 expression in non-GC B cells (black) is shown as reference
C B6 mice were orally gavaged with 109 CFU erythromycin‐resistant NCK56 or NCK2187. Fecal pellets were collected daily and tested for the presence of erythromycin‐resistant strains. n = 3 mice/group. Data are representative of five independent experiments and are shown as mean ± SEM.
(C) BT-474 cells were injected subcutaneously into athymic Foxn1nu nude mice. Mice were treated with either vehicle (8 mice) or MK-2206 (100mg/kg) (10 mice) or 14h (25mg/kg) (6 mice) or both, MK-2206 and 14h (10 mice) for 24 days. The tumour volume was measured twice a week. Tumourgrowth was represented as the fold change mean ± SEM.
O, P SQST‐1::GFP aggregates are partially restored in daf‐16(mu86); rpl‐43(bp399); daf‐2(RNAi) animals.
Volcano plot of all RP genes in splenic WT and miR-132-/- CD4+ T cells from d28 L. donovani infected mice. RPL genes are shown as circles, RPS genes as triangles, and pseudogenes as squares. Red symbols indicate significant difference between WT and miR-132-/- cells (p<0.05) whereas black non-significant.
Histogram showing the distribution of the fluorescence peak time of individual cells expressed as a percent of the generation time (52 < n < 88). Different strains are color-coded. Statistical analysis by one-way ANOVA and Holm-Sidak's multiple comparisons test. Pulses are weaker and fewer in the strain devoid of positive regulation but are not displaced relative to the generation time.
Quantitation of the BODIPY signal shown as normalised LD size of lipid droplets for the genotypes The box end points are the upper (75%) and lower (25%) quartiles, the whiskers define the maximum 95th percentile and minimum 5th percentile values respectively, the central band is the median and the square is the mean. Data were analysed using the Kruskal Wallis test followed by Dunn's test for post hoc analysis for significance due to unequal sample sizes. ***p<0.001, **p<0.01.
A, Schematic representation of the genomic area of Dnmt3a and Dnmt3b in wildtype (WT) mice and conditional knock-out (KO) mice. WT mice were Nestin::Cre-ERT2(‑/‑)/ Dnmt3afl/fl/Dnmt3bfl/fl, while KO mice were Nestin::Cre-ERT2(‑/+)/Dnmt3afl/fl/Dnmt3bfl/fl.
Performance in the Rotarod test of ASMko and wt mice treated or not with Ca074Me for 1 month expressed as mean ± SEM time spent in the rotating rod on each of four trials (B)
Overexpression of MEF2C in the NDIME-knockout cells rescued neural differentiation defects at day 5 of neural differentiation, as shown by microscopy
Ratio of oxygen consumption rate (OCR) to extracellular acidification rate (ECAR) of preadipocytes and adipocytes after 4 days of in vitro differentiation. Data are shown as mean ± SEM of 3-7 cell lines per group.
A. Representative pictures of TUNEL staining together with CD31 and Lyve-1 in Peyer's Patches of Casp8WT and Casp8ECko mice. TUNEL+ LECs, but not BECs could already be seen in non-hemorrhagic Peyer's Patches (a'), and further accumulated in hemorrhagic Peyer's Patches (a''). White dotted areas outline location of the lymphatic plexus. White arrowheads point to TUNEL+ LECs (note that necroptotic LECs only weakly express Lyve-1). Scale bars: 50µm, insets: 10µm.
Representative images and analysis of Western blots for PAR, performed with lysates of HME control (vec) OAS single KO cells (KO1-3), treated with H2O2 for 15 or 40 min. β-actin is shown as a loading control. Signal intensity was measured using ImageJ software. Average lane densities, normalized to vector from three independent experiments, are plotted below as mean + SD, * p < 0.05 as assessed by Student's t-test.
HEK293 Tet-off experiments showing the degradation of REL-OPT, REL-OPT (+1 Frameshift) and REL-WT transcripts (E) post-doxycycline addition. data is representative of 3 independent experiments each with 3 replicates. The data represents the mean ± SD for 3 replicates. A two-way ANOVA with Holm-Sidak multiple comparisons was performed. P-values are denoted as follows: p < 0.05 (*), p<0.01 (**) and p<0.001 (***).
a, Sequence-structure alignment of S. flexneri IpaJ with members of C39-peptidase-like family identified by HHPred. IpaJ possesses invariant catalytic triad residues Cys 64, His 206 and Asp 218 (red). The conserved Gln 58, which helps form the oxyanion hole found in many proteases, is shown in green. Invariant residues and core hydrophobic residues found in the C39-family are shown in blue and magenta, respectively. Characterized C39-family member (GCN5-acyetyl transferase) and a C1-family member (Papain) are shown.
D Loss of autophagy activity suppresses the Muv phenotype in let‐60(n1046) mutants. The number of adult animals examined was: wild type (n=450), let‐60 (n=406), epg‐6; let‐60 (n=444), let‐60; atg‐2 (n=803), epg‐5; let‐60 (n=672), epg‐5; let‐60; ain‐1(ku322) (n=635). P values are determined by a two‐tailed χ2 test.
(C) Supernatants containing indicated proteins were diluted into PBS or pH 6.1 MES buffer and analyzed using SDS-PAGE and Western blot under reducing conditions. HRP-conjugated anti-FLAG antibody was used for detection of all proteins. Numbers on the side of blots indicate positions of molecular weight markers. Reduction-resistant dimers are indicated with an * and reduction sensitive protein with a +.
A. Scheme of chorion isolation where mesoderm is in green (mG), and the other cells are in grey (mT).
(C) Body temperature of AdRiKO and control mice infected with either AAV9-HKII or AAV9-empty housed at 22°C (n=8/group).
Knockdown efficiency of two shRNAs against GPR56 (shGPR56weak and shGPR56strong) versus shLuc as negative control measured on protein level by flow cytometry in CD34+ CB cells. Shown are representative FACS plots (left) and the percentage of GPR56+ cells of transduced GFP+ cells on day 5 (right panel). Biological N=4, unpaired t-test, bars and error bars represent mean and standard deviation. *** p<0.0005, ** p<0.005, * p<0.05.
G. Rotarod test of 6 month-old TH-Cre and TH-Cre;fl/fl mice. n=11 and 10, respectively (unpaired t-test, *p<0.05, ** p<0.01, mean +s.e.m.).
Precipitates were analyzed with the indicated antibodies. F, GST pull-down assay using E. coli extracts expressing GST-fused DND1 mixed with MBP-tagged LacZ, NANOS2, NANOS2 (C61A, C96A) or NANOS2 (60-114). Arrowheads indicate MBP-tagged NANOS2 (lane 2) or NANOS2 (60-114) (lane 4).
H) Data was extracted from Mauvoisin et al. (2014), where the proteome was quantified in WT and CKO mouse livers extracted under diurnal conditions. Absolute fold change was calculated from maximum/minimum protein abundances. Probability density distributions for fold-change are plotted. Kolmogorov-Smirnov test: ****p<0.0001.
D Loss of Cideb inhibits diet-induced SREBP activation. IB of hepatic SREBP processing and lipogenic enzyme levels (Fasn, Scd1) of WT and Cideb-/- mice under chow diet and HFLF diet.
(c) Co-localization between ATG9A and VAMP3-HA or GFP-LC3 and VAMP8-HA in control and CALM knockdown HeLa cells stably expressing VAMP3-HA or VAMP8-HA. Confocal pictures are shown with magnified areas showing co-localization between ATG9A and VAMP3 or GFP-LC3 and VAMP8 in control cells and no co-localization in CALM knockdown cells. Quantification of ATG9A-VAMP3 or GFP-LC3-VAMP8 co-localization is shown on the graph as a Pearson's coefficient (data are mean ±s.d.; *P0.05; two-tailed t-test). Scale bars, 5 μm.
PD-1 expression in T cells (n = 6 biological replicates, mean ± s.e.m., ns, not significant (P > 0.05), ***P < 0.0001, two-tailed unpaired Student's t-test) isolated from mock and OTUD1 overexpressing LLC tumors. MFI, mean fluorescence intensity.