EMBO/sd-character-level-ner huggingface dataset:
(D) Quantification of Parkin recruitment to mitochondria (YFPParkin signal overlapping MtDsRed signal - intensity-adjusted and normalised to t=0) in WT and Miro1KO neurons following valinomycin treatment (n=15 cells all conditions per genotype over 3 neuronal preparations; two-way ANOVA). Error bars represent s.e.m. Significance: *p<0.05, **p<0.01 and ***p<0.001
(B) Treatment of SARS-CoV-2 (106 PFU) infected human kidney organoids with hrsACE2 (200µg/ml) and/or remdesivir (Remd.4µM)). Viral RNA was determined by qRT-PCR 72 hours after the inoculation of 106 PFU of SARS-CoV-2.
(e) Overexpression of EndoB1T145A attenuates α-synucleinA53T mutant-induced autophagy in neurons. Data are means ± s.e.m.; n=7. Uncropped images of blots are shown in Supplementary Fig. S8.
Comparison of binding affinities of Nxph1 binding to different Nrxn1-3 LNS2 isoforms.
Fate of Purkinje cells in mice with Purkinje-cell-specific knockout of Ccp1. Ccp1flox/flox L7-cre mice show a massive but incomplete degeneration of Purkinje cells (calbindin-stained; red) at 4 months, while all Purkinje cells are degenerated at 18 months. Scale bar: 500 µm.
Open field (OF) test. (C) Total distance traveled during the OF test. No significant difference among groups. F(2, 25) = 2.23, P = 0.13, ANOVA. (D) Velocity of travel during the OF test. F(2, 25) = 1.60, P = 0.22, ANOVA. N.S. (not significant). (E) Fraction of time spent in the periphery. F(2, 25) = 10.71, P = 0.00044, ANOVA followed by Tukey Kramer, ** P < 0.01. (F) Fraction of traveled distance in the periphery, F(2, 25) = 3.84, P = 0.035, ANOVA followed by Tukey Kramer, * P < 0.05 (n: WT = 10, WT-old =7, Lmnb1-cKO = 11 for the OF test).
Mean C16:0 ceramide levels in MEFs pre-treated for 3 h with vehicle (n=7), SH-BC-893 (5 μM, n=7), myriocin (myr, 10 μM, n=5), or fumonisin B1 (FB1, 30 μM, n=3) then treated with BSA or palmitate (250 μM) for 3 h. Cells treated with C16:0 CER (100 μM, n=2) for 3 h shown as a positive control. Error bars, SEM.
B. Representative time-lapse sequences of mitotic progression in cells in (A). NEBD corresponds to t=0. White arrowheads indicate cells undergoing division. Scale bar, 10 μm.
Above: Representative funduscopic pictures from living healthy Cx3cr1CreERT2:Rosa26-tdTomato mice on d0. Funduscopy and red fluorescence visualize the fundus and regular distribution of tomato+ microglia before the laser-induced lesion formation. Below: Corresponding immunofluorescence pictures. Non-lesioned retina show a regular pattern of Iba1+ (green) tomato+ (red) retinal microglia while macrophages are absent on the retinal pigment epithelium (RPE) under native conditions. Pictures are representative for six mice analyzed in one experiment. Scale bars represents 50 µm. c) Above: In vivo funduscopy on d7 post lesion. Funduscopic and red fluorescence image depict the lesions (encircled with dashed white lines) and accumulation of tomato+ microglia in Cx3cr1CreERT2:Rosa26-tdTomato mice. Intraperitoneal fluorescein application was performed to label retinal vessels and areas of choroidal neovascularization. Below: Representative immunofluorescence for Iba1 (green) in Cx3cr1CreERT2:Rosa26-tdTomato mice. Resident retinal microglia are Iba1+tomato+ (asterisks) whereas blood-derived myeloid cells are Iba1+tomato- (arrows) and accumulate at sites of laser-induced CNV. Overlay is shown left. Typical pictures from six mice obtained from one independent experiment are shown. Scale bars represents 50 µm.
G. Cell survival assays of WT and PRIMPOL KO cells with or without MCM3 downregulation in the presence of increasing amounts of MMC or TMP (followed by 90 sec UVA irradiation) for 72 h. The same control curves without MCM3 downregulation are also shown as part of Figure EV6D. Each point in the curve represents the percentage of surviving cells (average and SD of three assays for MMC and two assays for TMP). Statistical analysis was conducted with two-way ANOVA followed by Bonferroni post-test. p-values of comparisons between KO and KO-siMCM3 are indicated.
B Electron micrograph of a cell expressing GPD promoter-driven GFP-Pho8. Arrows indicate the edges of ER cisternae that are part of a bent stack in a vacuole membrane invagination. Black arrowheads point to vacuole membrane that encases the bent stack and the whorl.
(f) Hold out analysis shows the robustness of associations to holding out a dataset. The values show the number of held-out datasets that result in loss of association between a given covariate (rows) and ACE2, TMPRSS2, or CTSL expression in a given cell type (columns). Robust trends are determined by significant effects that are robust to holding out any dataset (0 values).
F Co-IP analysis indicating RNAP2 binding in H3K9cr enriched regions in NSPCs. H3 was used as a loading control for the input nuclear proteins.
C. Sufu protein levels in DAOY cells treated as in A. Representative image of three independent experiments is shown.
F GNL2 and SEC13 contribute to the negative NMD feedback loop, regulating the levels of transcripts encoding NMD factors. RT-qPCR analysis of total cellular RNA from HeLa cells depleted of GNL2 (in green) and SEC13 (in purple) is shown. The graph shows distribution of relative fold-change from eight independent experiments relative to mock-depleted cells (control). Statistical analysis was performed using Student's t-test. *P < 0.05.
E. Chromatin immunoprecipitation showing that H2AX binding to the common fragile site FRA3B is reduced in HUWE1-knockout 8988T cells. Cells were treated with 600nM aphidicolin for 24h. Binding was quantified relative to input material. Shown is the average of 4 experiments -/+ SD. P-value is 4.4x10-6.
C Histogram of amino acid frequencies at the linchpin position of all 42 yeast RING domains. The linchpin position was defined as the residue at the n+1 position after the final Zn2+ ligand coordinating residues. "-" refers to the atypical RING of Pib1, which lacks the loop containing the linchpin. The SP-RING domains of the two yeast SUMO E3 ligases Siz1 and Siz2 as well as the RING1 domains of the two yeast RBR E3 ligases Hel1 and Itt1 were excluded from this analysis.
The loss of SP1 expression is associated with a decrease in SWI/SNF occupancy of the SP1 promoter and an increase in H3K27me3. Data represents ChIP-qPCR analysis following 8-hours or 18-hours of 100nM mithramycin exposure and immunoprecipitation of SMARCC1 (8-hour, P= 0.0006; 18-hour P=0.0001) . ChIP quantitation is percent input (ng of immunoprecipitated DNA/input DNA *100) determined by absolute quantitation of sheared chromatin relative to a standard curve. Data represents mean with standard deviation derived from three independent experiments. P-values were determined by one-way ANOVA using Dunnet test for multiple comparisons.
B) Representative immunoblots showing endogenous LC3 lipidation in the presence of bafilomycin A1 (BafA1). (C-E) Transgenic C57BL/6 mice expressing a GFP-LC3 fusion protein were injected with resveratrol and spermidine at the indicated concentrations. 3 h later, mice were killed, and tissues were processed for immunofluorescence microscopy determinations of GFP-LC3vac.
(A) Immunofluorescence analysis of HeLa cells transfected with either a non-target dsRNA (NT) or one of the three siRNAs targeting LETM1 (siR1, siR2 or siR3) and labeled with anti-TOM20 antibody. In siR1 treated cells, the mitochondria formed a 'honeycomb' of swollen distinct organelles; siR3 resulted in giant organelles with a central region distinguished by reduced TOM20; siR2, produced relatively little swelling and the mitochondrial network was generally well preserved. The pronounced swelling induced by siR1 significantly increased circularity (p = 2.32E-58) (ImageJ analysis), siR1 circularity = 0.683 ±0.012 compared with 0.414 ±0.008 for the NT; where 1 = a perfect circle. Over 150 mitochondria were quantified in each cell type; data are means ± SEM. Scale bar represents 12 µm in the main images and 4 µm in offset magnification
(i) Precipitated proteins in A10 cell lysates with m7GTP-agarose beads were identified by Western blot analysis. eIF4E and tubulin were shown as positive and negative controls, respectively. Total lysates were used as input. (ii) A10 cell lysates were incubated with GTP-agarose beads, and none of the proteins tested were precipitated with the beads. Total lysates were used as input control
(B) Poly(A) tail analysis of codon tag reporter mRNAs at 2 and 6 hpf. The developmental stages are shown above. The lane labeled A0 shows the 3´ UTR fragment without a poly(A) tail. (C) Quantification of the PAT assay at 6hpf in (B).
A. ChIP-seq analysis was performed in control or TRIM46-overexpressing MCF-7 cells with antibodies against HDAC1. Density distributions (read count per million of mapped reads) of HDAC1 peaks were analyzed by deepTools.
B. BMDMs were isolated from HDAC6+/+ and HDAC6-/- mice and were transfected with 5ˊppp-dsRNA. Whole cell lysates were prepared after 8 h of transfection and co-immunoprecipitation analysis of RIG-I and HDAC6 was performed.
A SARS-CoV-2 genome annotation, which predicts 29 virus gene products. The 16 non-structure proteins (NSPs) are cleaved products of the large polyprotein open reading frame (ORF) 1ab or ORF1a. These polyproteins are cleaved into small function fragments or NSPs after translation.
G-I, Immunostaining for HIF-1α (red) and DAPI (blue) in WT cortices at E10.5 (G-G''), E11.5 (H-H'') and E12.5 (I-I''). Panels G',G'',H',H'',I' and I'' are magnifications of the boxed areas in their respective original panels. Arrows point to the border of lateral and dorsal cortex.
(A) ROS accumulation in Gmlmm1-1 and Gmlmm1-2 mutant plants. Soybean leaves (aged 3 weeks) were subjected to ROS accumulation examination by DAB staining. Scale bar, 1 cm. Data information: The experiments were performed three times as biological replicates, with similar results. The exact number (n) and p values are shown in the source data.
(C) The average number of insulin granules in β-cells from the mouse pancreatic islets (A) in microscopic sections. Data information: A total of 15-23 images per group were analyzed. Data from each group are presented as the mean ± SD. One-way ANOVA test was used for statistical analysis of differences between groups, and P (*) < 0.05; P (**) < 0.01 and P (***) < 0.001 are considered statistically significant.
K, Boxplots showing the proportion of G/C nucleotides in all genes and in genes depleted/enriched in dsRNA-seq compared to total RNA-seq (n=3). Shown are the P-values in the non-parametric Mann-Whitney U test (*** P < 0.001). Boxes extend from the 25th to 75th percentiles (inter-quartile range (IQR)), horizontal lines represent the median, whiskers indicate the lowest and highest datum within 1.5*IQR from the lower and upper quartiles, respectively.
a, Sections of the thymus and muscle of fed and starved GFP-LC3 mice were stained with anti-GFP Alexa 488 antibody (green) and 4,6-diamidino-2-phenylindole (DAPI; blue). Images are representative of n = 6.
D. same as A using GST-tagged C-terminally truncated Elp1 in combination with truncated versions of Elp4.
Human neural cultures transduced by Auto-hLAG3 were treated by α-synuclein PFFs 4 days post LAG3 expression induction by DOX and kept in culture for 2 or 4 weeks. Both transgenic (hLAG3 D2G40-positive in (A) and 17B4-positive in (B)) and non-transduced, wild-type neurons (selected neurons in zoomed-in insets) propagated α-synuclein and developed characteristic pS129-positive (81A-positive in (A) and EP1536Y-positive in (B)) α-synuclein aggregates. Scale bars 25 µm. Trained ilastik algorithms were used to segment pixels of 81A, EP1536Y and MAP2 stainings imaged by high-content wide-field microscope, which were used to quantify the signal of 81A-positive (C) and EP1536Y-positive (D) α-synuclein aggregates expressed as % of MAP2-positive area. Almost the entire wells (182 fields per well) were imaged for every condition and replicate and each datapoint in the plot represents the entire well. Error bars indicate mean ± SD of biological replicates (duplicates). For few conditions, unicates were used, shown as one dot. One-way ANOVA followed by Tukey's multiple comparison test demonstrated that neurons that did not express LAG3 (DOX OFF) showed no difference in α-synuclein propagation when compared to LAG3-expressing neurons (DOX ON) as demonstrated by two different pS129 α-synuclein antibodies (p=0.9998 for 81A at 2 weeks and p=0.2522 at 4 weeks; p=0.9986 for EP1536Y at 2 weeks and p=0.3042 at 4 weeks).
D Wild type and Traf3ip3-/- HEK293T cells were transfected with pcDNA3-FLAG-MAVS-(Region III only). Twenty-four hours post-transfection, cells were infected with or without VSV. Twelve hours post-infection, cells were collected and subjected to immunoprecipitation assay and immunoblotting.
F, Protein blot of polyAla RAN proteins expressed from pure or interrupted constructs; n=3 transfections
C7-hypomorphic mice were treated with losartan for 7 weeks, and the forepaws of age-matched untreated, losartan-treated, and wild-type mice were subjected to immunofluorescence staining with antibodies to fibrosis markers (A, B) and to picrosirius red staining (C). C Picrosirius red staining and visualization of the collagen fibers under cross polarizing light. Under this light, thin fibers appear green and thick rigid collagen bundles orange-red. The staining revealed significantly reduced collagen fiber size in losartan-treated skin, indicating softer tissue similar to wild-type skin. Below, the bar graph shows quantification of picrosirius red-positive areas, n ≥ 19 areas quantified, values represent mean ± S.E.M. Unpaired t-test with Welch's correction, ***P-value wild-type vs. C7-hypomorph < 0.0001; ***P-value C7-hypomorph + losartan vs. C7-hypomorph = 0.0004; *P-value wild-type vs. C7-hypomorph + losartan = 0.0189. Images acquired with a 20× objective, scale bar = 50 μm.
d, MSNs were depolarized with potassium chloride (HiK, 55 mM) or treated with the D1-agonist SKF-38393 (10 µM) for 1h, 2h or 4h. Subsequently, qPCR was performed to assess immediate early gene mRNA levels. Mean + SEM; two-way ANOVA with Bonferroni post-hoc test; indicated p-values for induction over control for each treatment; p* < 0.05, p** < 0.01, p***<0.001; n = 4 independent cultures as biological replicates.
Representative images of β-galactosidase staining in TOPGAL epithelium (C) BALO cultures before (day 6, control) or 5 days after treatment with either 4 μM Scra or mo142-3p (day 11 of co-culture). BASC and rMC were isolated form the lung homogenate of TOPGAL mice. β-galactosidase+ rMC at day 11 of culture are indicated with arrows.
SARS-CoV-2 S1 RBD protein model.Basic amino acids that are solvent accessible on the surface are indicated (dark blue); these can be observed to form a continuous patch.
C. MLE-12 AECs incubated with MH-S EVs infected with PR/8 influenza. Replication was quantified by RT-qPCR at 12 h post infection. Data represent mean M gene transcripts normalized to β actin from 3 independent experiments.
RT-PCR analysis (top) and representative direct sequencing (bottom) of Ttn exon 326 transcripts from HL-1 cardiomyocytes transiently transfected with different 2OMePS AONs. Only 2OMePS-AON1 and 2OMePS-AON1 + 3 (*) lead to a correct excision of exon 326 as confirmed by direct sequencing.
A Alignment of the amino acid sequence of the yeastH subunit inhibitory loop region (shown in Fig 4C). The yeast (Sc) loop was aligned with those from insect (M. sexta), zebrafish (D. rerio), frog (X. trop), mouse (Mus), bovine (B. taur), chimpanzee (P. trog) and human (Hum1, Hum2), illustrating the lack of sequence conservation in this region. Pink coloring denotes predicted α helical secondary structure.
Representative images of WT and Mecp2 null neurons (DIV14) immunostained for MAP2 (white), Synapsin1/2 (green) and Shank2 (red). Mecp2 null cells were treated at DIV12 with tubacin (TUB, 1µM for 48 h) or Aurora A inhibitor TC-S 7010 (TC-S, 7 nM for 48 h). Scale bar 15 µm, and 5 µm in the enlarged image.
B Time lapse images showing simulations of the coupled STEN-CEN model demonstrating CEN (top) and STEN (bottom) waves spreading in space. The threshold was lowered by increasing positive feedback of STEN (perturbation) at time 0. The white arrow (time 0, CEN) denotes a wave at a high threshold that disappears in the next image, while the red arrow (time 25, CEN) points to a lowered threshold wave that enlarges and spreads.
A. Schematic illustration of 2-DG uptake assay protocol for control and NICDiOE-EC mice.
D. SIRT7 is mono- and asymmetrically di-methylated at R388. Flag-tagged SIRT7 and its RK/RF mutants were expressed in HEK293T cells. Immunopurified SIRT7 was blotted with antibodies against mono-methylarginine (α-me1), asymmetric di-methylarginine (α-me2a), and symmetric di-methylarginine (α-me2s), respectively
Co-localization of Omp25 and Omp2b positive pixels with TRSE positive or negative pixels, respectively. Error bars correspond to s.d. from independent experiments. Differences were statistically analyzed by t-test. ***P < 0.001. nOmp25 = 279 bacteria, 85666 pixels (5 biological replicates). nOmp2b = 252 bacteria, 74630 pixels (3 biological replicates).
Dam-ID experiment to identify the binding of Pc on Octβ2R genomic locus. Primers used were shown in (I).
Morphological alterations and intracellular signalling in AT2 cells cultured with IMC-CM. On the left and middle, primary AT2 cells from 3-week-old BVE mice were cultured for 48 h and then incubated in serum-free IMC-CM (AT2 with IMC-CM) or DMEM (AT2 w/o IMC-CM, serving as a control) for another 48 h, followed by immunostaining for E-cadherin (red) and vimentin (green) and confocal laser scanning microscopy (CLSM) imaging. The 3× zoomed image in the middle right highlights dividing vimentin+ cells that show internalised E-cadherin (arrows). Scale bars, 50 μm. On the right, primary AT2 cells from 10-week-old BVE mice were cultured for 48 h, serum starved for 5 h and treated with or without serum-free IMC-CM for 30 min. Phosphorylation of AKT, SMAD3, MEK and ERK were analysed by immunoblotting.
E Measurement of serum urea levels of mice in their fifth week of life (Phb2fl/fl/het 44.00 ± 7.02 mg/dl versus Phb2pko 493.70 ± 45.34 mg/dl, n = 3 for both groups; ***P = 0.006).
D. Δ31-160 forms spontaneous aggregates with moderate chymotrypsin (Chy) resistance. Immunoblots probed with mAbs C16-S (upper panel) or 3F4 (lower panel) for comparing Chy-resistant fragments of Δ31-160 with that of Δ159. Lysates from N2a cells expressing Δ159 or Δ31-160 were digested with indicated concentrations of Chy for 30 minutes and then digested with PNGaseF. A sample from non-transfected N2a cells is shown in the lane 5. Asterisks indicate non-specific C16-S bands. Arrowhead denotes deglycoform of Δ159, bracket deglycoform of Δ31-160.
(A) NIH3T3 fibroblasts co-expressing DsRed-CA7 and EGFP-CA2 (n = 4 independent replicates).
A. Cysteine 7 is predicted to be palmitoylated and was changed into alanine in the concavinC7A-GFP mutant.
The fraction of RNA-seq intronic reads calculated for all genes (n=24912) in NRPB2WT +/+ nrpb2-2 -/- and NRPB2Y732F +/+ nrpb2-2 -/-. Two-sided Mann-Whitney U test: **** denotes p-value<2.2e-16. The solid horizontal lines and box limits represent medians, lower and upper quartiles of data values in each group. The upper and lower whiskers extend to the largest or smallest value, respectively, no further than 1.5 * IQR from the relevant quartile.
Relative quantification of COASY mRNA from human fibroblasts treated with pPanSH. n=3 for both genotypes.
ELISA of IL-1β secretion by WT or Spata2-KO BMDMs that were either untreated (-) or LPS-primed and then treated with the indicated NLRP3 inducers
B Confocal images of PHcrac (left) and PHcrac merged with LimE (right, PHcrac as red, LimE as green) in a giant cell after RacGEF1ΔN recruitment.
ChIP-qPCR detection of Pol II occupancy at the indicated promoters in Myoparr-depleted C2C12 cells. The data were normalized to input values. n = 3, mean ± SD. *p < 0.05. Myoparr KD decreased Pol II occupancy at the miR-133b promoter with a marginal trend toward significance (p = 0.052).
a. Immunofluorescent images of skin sections from PDGFRα-eGFP reporter mice with normal (WT, n=4), inflamed (InvEE, n=4), wounded (d14 post-wounding; wound, n=4), or tumour-associated (Papilloma, Pap d17 and d30 post-wounding, n=4 per condition) skin. Nuclei were stained with Dapi (4', 6-diamidino- 2- phenylindole; blue). Dotted line represents epidermal-dermal boundary. Scale bars: 50 μm. b. Quantification of the number of fibroblasts present in different skin conditions (n=4 mice per condition; ** p<0.01; *** p<0.001; **** p<0.0001; One-way ANOVA testing). Data represent means of multiple microscopic fields ± S.E.M.
(G) K+ efflux activity of the oocytes expressing the indicated proteins. The oocytes were injected with H2O, IAA, or BA. Data are means ± SE (n = 3, biological replicates; each replicate contains 6 oocytes).
Cell death analysis by Celigo of PI-positive HS-ES-2R Cells were treated with the indicated agents for 48 hrs (HS-ES-2R) DMSO (Unt), zVAD (10 µM), TNF (100 ng/ml) and SM (100 ng/ml) SM represents SM-164. Error bars represent SD. Displayed are representative results from n=3, and statistical analysis was performed with an unpaired t-test, *** P ≤ 0.001.
(G) iFRAP quantification of stably chromatin bound SMC3-LAP after siRNA treatment using cells synchronized in G2-phase. Error bars denote s.e.m., n > 11 cells per condition.
Global transcriptome analysis of Ctcflos KD impact on alternative splicing (KD day 1 of differentiation, analysis after 24h). (Y) MA-plot of differential alternative splicing analysis using SGSeq algorithm comparing Ctcflos tr1 (ASO 1) KD with controls. Each data point depicts a splice event. Splice events of differential abundance between Ctcflos tr1 (ASO 1) KD and controls (p-adj. <0.05) are colored in red.
A. Cartoon illustrating the single-molecule DNA unzipping experiment used for the detection of the interactions between dSaCas9 and DNA and the definition of the DNA sequence.
quantification (G) of β-TrCP associated with Ubch5, YAP or β-catenin in DU145 cells transfected with the indicated siRNA (control, siCTRL; UBTD1: siUBTD1pool).
Tracer distribution in AV+ types. Cells were allowed to endocytose tracer particles continuously for the designated periods of time. At each time, random sections were searched for AV+. The results are categorized according to AV types and plotted for the relative distribution of tracers. The total tracer counts are given on top of each bar. The number in each block represents the actual number of AV profiles, from which tracer counts are scored.
(g) Left, immunoblot for LC3-II in cells treated with 20 μM of the retinoid derivatives and protease inhibitors (PIs), as labeled. Actin is shown as loading control, and full-length blots are shown in Supplementary Figure 21. The amount of LC3-II in untreated cells, expressed as fold change relative to control (middle), and the increase in LC3-II after PI treatment (LC3-II flux, right), expressed as fold change, were calculated from the densitometric quantification of immunoblots. Values are mean ± s.e.m.; n = 3.
Groups of BALB/c female mice (n = 5) were inoculated twice (four weeks apart) i.m. with 10 µg/dose of the proteins indicated at the top of each panel, folded in their respective postfusion conformations. One week after the last dose, mice were sacrificed and their blood collected. A) Each mouse serum (identified by a number) was tested in ELISA for antibody binding to the postfusion F proteins indicated at the bottom. The results represented in the y axis for each individual mouse serum correspond to the inverse dilution that gave 50% of maximal binding, expressed in log10 units (EC50 (log10)). Each number represents an individual mouse. Short horizontal bars indicate mean values for each group. P values were calculated as indicated under Statistical Analysis. Only relevant P values are shown. *P = 0.00045. Differences were considered significant when P < 0.05.