EMBO/sd-character-level-ner huggingface dataset:
Figure 6pi4kβ1 pi4kβ2 double mutant phenotype were recorded. Immunostaining of meristem cells of five-day-old plants using anti-tubulin (green) counterstained with DAPI (false colour red). Asterisks, multiple nuclei in one cell; arrowheads, multiple phragmoplasts in one cell. Dashed lines indicate the outlines of exemplary multinucleated cells. Scale bars, 10 µm. pi4kβ1 pi4kβ2 double mutant phenotype were recorded. Whole-mount immunostaining of root tips of five-day-old seedlings. Green, anti-KNOLLE; red, anti-tubulin; blue, DAPI. (І), (ІІ), magnifications of areas highlighted in dashed boxes. Arrowheads, solid phragmoplasts with thicker cell plates. Scale bars, 20 µm. pi4kβ1 pi4kβ2 double mutant phenotype were recorded. Quantification of the incidences of multinucleated cells, multiple phragmoplasts and solid phragmoplasts during late cytokinesis (multinucleated cells: wild type, n = 316 cells, 29 roots; pi4kβ1 pi4kβ2, n = 673 cells, 44 roots; multiple phragmoplasts: wild type, n = 64 cells, 29 roots; pi4kβ1 pi4kβ2, n = 188 cells, 44 roots; solid phragmoplast in late cytokinesis: wild type, n = 41 cells, 29 roots; pi4kβ1 pi4kβ2, n = 89 cells, 44 roots). Data are mean ± SD from three independent experiments. n.d. not detected.
Figure 5A, Maximum intensity projection of a confocal micrograph of a cryostat section from a Nrl-/- recipient retina 21 days after transplantation with Nrl::GFP donor photoreceptors, showing protrusions (pink arrows) connecting donor and acceptor photoreceptors. , 3D reconstruction of confocal microscopy images of whole-mounted retinas from Nrl-/- (B) recipients 21 days after transplantation with Nrl::GFP donor photoreceptors (top layer) shows that donor and GFP+ acceptor photoreceptors appear to be attached via protrusions. C, 3D reconstruction of confocal microscopy images of whole-mounted retinas from C57BL6/J (C) recipients 21 days after transplantation with Nrl::GFP donor photoreceptors (top layer) shows that donor and GFP+ acceptor photoreceptors appear to be attached via protrusions. Higher magnification 3D reconstruction (C) showing protrusions (pink arrows) and GFP labelling in acceptor cells. D, E, Left: 3D reconstruction of two-photon (D) and light sheet microscopy images of intact cleared ROSAmT/mG eye showing that GFP+-donor and -acceptor photoreceptors are in the same area. D, Right: A high magnification image showing GFP+-donor and -acceptor photoreceptors connected by protrusions (pink arrows). E, Right: In detail, donor and acceptor photoreceptors also appear connected through cell protrusions (pink arrow). F, Percentage of GFP+-acceptor photoreceptors connected via a donor cell protrusion across different imaging modalities (n=3 animals per imaging technique).
Grin2aS/S mice are viable and show no signs of neurodegeneration or altered brain structure.Fig. 3aGrin2aS/S mice had a reduced body weight (by 18% in adults), significantly lower score in nesting and in overnight burrowing activity (filled circles: 30-week-old mice, white dots: 18-week-old mice). b Grin2aS/S mice showed the clasping reflex. c During the 12 h dark phase on the Lafayette running wheel the running distance is reduced in Grin2aS/S mice. d Nissl staining revealed unaltered cell density throughout the brain (top) and layering of hippocampal subfields, including CA1 (bottom), in Grin2aS/S mice compared to Grin2a+/+. Error bars represent mean ± SEM. e In the TUNEL assay no apoptotic cells could be detected in the hippocampus of Grin2aS/S mice, compared to the staining of the nuclease-treated positive control slice. f No aberrant mossy fiber sprouting in either genotype in the dentate gyrus inner (IML) and outer molecular layer (OML) can be found by Timm staining of coronal sections. g The distribution of NeuN-positive neurons and of GFAP-positive glial cells are indistinguishable between Grin2aS/S and Grin2a+/+ mice. Mossy fiber projections visualized by anti-calbindin (CB) staining, as well as numbers of parvalbumin (PV)-positive interneurons are similar between both genotypes. Anti-GluA1 immunosignal in all hippocampal layers is comparable between brain sections of Grin2aS/S mice and control littermates. CA1 cornu ammonis region 1, CA3 cornu ammonis region 3, DG dentate gyrus, mol stratum moleculare, ori stratum oriens, rad, stratum radiatum. Scale bars in d-g are in mm. The number of animals is given below the bars. For the Nissl stain, Tunnel test and Timm stain 3 mice were used per genotype. For the immunohistological analysis of glial fibrillary acidic protein (GFAP), neuronal nuclear antigen (NeuN), Calbindin (CB), and Parvalbumin (PV) five mice and for the GluA1 immunofluorescence stain, three mice were used (for statistics: Supplementary Statistics to Fig. 3).
Figure 4A. Amino acid sequence of human Fis1. The location of the six α-helices, the tetratricopeptide repeat (TPR) motifs, and transmembrane domain (TM) are indicated. B. Interaction of different hFis1 mutants with Mfns and OPA1. hFis1-/- 293T cells were transfected with the full-length and the truncated hFis1 mutants as indicated, followed by in vivo crosslinking with 1% FA. Cell lysates were used for co-IP with anti-Myc or anti-GFP agarose beads and the immunoprecipitates were analyzed by Western blotting with indicated antibodies. C. Summary of hFis1 mutated constructs used in this experiment.
Variation and phylogenetics of NYC COVID-19 Cases.Figure 4:The phylogenetic placement of these COVID samples is shown on the tree (left) and the global map of known COVID genomes (right). Genetic variants called from the RNA-seq data show a range of variants that are distinct from the Wuhan reference strain, and the samples from this study, highlighted in blue, show enrichment for European and Asian alleles. (b) Proportion of the L (green) and S strain (yellow) are shown for the NYC viruses. Phylogeny of samples from this study on the left and total GISAID samples on the right, with a map of variants in this study's samples in the middle, colored by event type and sized by number of nucleotides impacted. Annotation track on the bottom shows frequency of alternate alleles in this study and in the GISAID database. (c) The 9-bp deletion in ORRF1b (NSP1) that was detected in samples from two patients was confirmed in the GISAID database (below). (d) Comparison of the ORF1b of the SARS-CoV-2 and SARS-CoV genomes with the 9-bp deletion region shown enlarged.
Figure 1A. Representative plots of splenocytes from CD45.1 mice adoptively transferred with CD45.2 OT-II WT or CCR5-/- lymph node cell suspensions, five weeks after infection with rVACV-OVA virus. The gating strategy used to identify the memory CD4+ T cell subtypes is shown (n = 5). B. Absolute number of OT-II cells recovered in spleens of mice as in A (n = 5). C, Percentage of CD4+ TEM (C) in the OT-II WT and CCR5-/- populations (n = 5). D. Percentage of TCM (D) in the OT-II WT and CCR5-/- populations (n = 5). E. IFNγ-producing OT-II WT and CCR5-/- memory cells isolated from mice as in A and restimulated ex vivo with OVA323-339 (1 μM) (n = 4). F. Immunization scheme for NIP-OVA and NIP-KLH in WT and CCR5-/- mice. G-I. Representative plots (G) and quantification of the frequency (H) and absolute number (I) of Tfh cells (CD4+CD44+PD-1+CXCR5+) in the spleen after primary immunization (day 7) with NIP-OVA (n = 7). J, K. ELISA analysis of high- (J) and low-affinity (K) isotype-specific anti-NIP antibodies in sera from OVA/OVA- and OVA/KLH-immunized mice (day 15 post-challenge; n = 5 mice/group). Data representative of one experiment of two.
Cortactin is recruited to protein aggregates and required for efficient autophagosome–lysosome fusion in homeostatic autophagy.Figure 6(A) Wild‐type and HDAC6 KO MEFs were treated with MG132 and immunostained with antibodies against cortactin (red), ubiquitin (green), and phalloidin for F‐actin (blue) as indicated. The arrows indicated ubiquitin‐positive aggregates that were colocalized with F‐actin and cortactin. (B) Wild‐type MEFs were transfected with control or cortactin siRNA, treated with MG132, and stained with antibodies for Lap‐1 (red, to label lysosome), or ubiquitin (green) and phalloidin for actin (blue). Note that F‐actin staining at protein aggregates was lost, but lysosomes remained concentrated in cortactin knockdown cells (arrow). (C) Wild‐type MEFs were transfected with control or cortactin siRNA, treated MG132 2.5 μM for 18 h, and subjected to filter‐trap assay using a ubiquitin antibody. The knockdown level of endogenous cortactin was confirmed by immunoblotting using an antibody against cortactin and GAPDH in the right panel. (D) U2OS cells were transfected with control siRNA and cortactin siRNA. Autophagosome-lysosome fusion was analysed with or without starvation (6 h) using the mCherry‐GFP‐LC3 reporter as described in Figure 3A. (E) The mCherry‐GFP‐LC3 plasmid was cotransfected with wild type, 9KQ (acetylation‐mimic), or 9KR (deacetylation‐mimic) cortactin‐expressing plasmids into wild‐type MEFs. Autophagosome-lysosome fusion was analysed as described in Figure 3A.
Figure 3Different expression of proteins in samples collected at the first timepoint. The heatmap (left panel) indicated expression patterns of proteins overrepresented (fold change > 2) in healthy control (1,656 proteins), mild (1,547 proteins) and severe (2,362 proteins) patient groups. The top categories enriched for clusters are shown. Values for each protein in each sample (columns) are color-coded based on expression level, low (blue) and high (red) z-scored FOT. The line plots (middle panel) indicated selected gene sets with up-up (upregulation in both mild and severe patients compared with healthy controls) or up-down or down-down regulation trend during disease progression. The bar plot (right panel) indicated Gene Ontology annotations with up-up or up-down or down-downregulation trend during disease progression (Fisher-exact test, p <0.05). The Weighted Gene Co-Expression Network Analysis (WGCNA) of 31 COVID-19 samples shows modules that are highly correlated with clinical features (left heatmap). Among the 33 modules (ME0-ME32), six of them showed significant correlation with clinical parameters (asterisk in the heatmap). Gene Ontology (GO) enrichment analysis were performed on these six modules and the enriched pathways were presented on the right panel (Fisher-exact test, p <0.05). Boxplots showing differences of the APTT time (left) and ME1-enriched protein expressions (right) between mild (n=13) and severe (n=18) COVID-19 patients. Differences between groups were estimated using Mann-Whitney-Wilcoxon test. * p <0.05, ** p <0.01, *** p <0.001. The horizontal box lines in the boxplots represent the first quartile, the median, and the third quartile. Whiskers denote the range of points within the first quartile − 1.5× the interquartile range and the third quartile + 1.5× the interquartile range. Boxplots showing differences of IL-6 level, IL-10 level (left) and ME15-enriched protein expressions (right) between mild (n=13) and severe (n=18) COVID-19 patients. Differences between groups were estimated using Mann-Whitney-Wilcoxon test. * p <0.05, ** p <0.01, *** p <0.001. The horizontal box lines in the boxplots represent the first quartile, the median, and the third quartile. Whiskers denote the range of points within the first quartile − 1.5× the interquartile range and the third quartile + 1.5× the interquartile range.
Representation of reactivation status of ex vivo cultures of CD8+-depleted PBMCs isolated from HIV+ patients.Figure 6Figure 6: Representation of reactivation status of ex vivo cultures of CD8+-depleted PBMCs isolated from HIV+ patients.A-C From data of ex vivo cultures of CD8+-depleted PBMCs isolated from HIV+ patients (Table 2), the extracellular HIV-1 genomic RNA levels for each LRA treatment are represented from all patient cell cultures (A), from patient cell cultures presenting no viral reactivation in mock condition (C), and from patient cell cultures exhibiting reactivation in mock condition (B). One night after cell purification, cells were mock-treated or treated with 5-AzadC. Three days post-treatment, 1/3 of medium was replaced and HDACIs were added in the cultures. Six days after 5-AzadC treatment, the concentration of viral RNA in culture supernatants was determined (in copies/ml). The results were reported as the actual HIV RNA copy numbers/ml or as an estimated value calculated as 50% of the smallest value when HIV RNA was not detected in order to assign a log value. Means are represented.
RAB21 interacts and colocalizes with the WASH and retromer complexes.Figure 3Strumpellin and VPS35 are in close proximity to RAB5 and RAB21. (A) Anti-biotin immunoprecipitation and immunoblot of endogenous Strumpellin and VPS35. Lysates correspond to 1% of input, n=3 independent experiments. RAB21 can be seen interacting with various WASH and retromer complexes subunits. GFP trap IP of WT, Q78L and T33N RAB21 variants in HeLa cells followed by GFP immunoblot and endogenous (B) VPS26 immunoblot Lysates correspond to 5% of input, n≥ 3 independent experiments. GFP trap IP of WT, Q78L and T33N RAB21 variants in HeLa cells followed by GFP immunoblot and (C) FAM21, Strumpellin and VPS35 immunoblots. Lysates correspond to 5% of input, n≥ 3 independent experiments. RAB21 actively pull-downs WASH and retromer complex subunits (D) Bacterially-purified and GTP-loaded GST:RAB21 pull-down of HeLa cell lysates followed by Strumpellin, FAM21, CAPZα and VPS35 immunoblots. Ponceau staining reveals the purity of the GST and GST:RAB21 used for the pull-downs. n=3 independent experiments. (E) RAB21 colocalizes with the WASH and retromer complexes. Transiently expressing GFP:RAB21 HeLa cells were fixed and stained for endogenous EEA1, FAM21, WASH and VPS26. Boxed region is magnified and single channels are depicted. Scale bars: 10µm or 5µm in enlarged view. (F) Per cell Pearson Correlation between RAB21 and the various markers. Error bars are SEM, n=2 independent experiments.
Figure 7The IgGsc-molecule was applied in escalating doses, with a starting dose of 20µg (patient 1) or 40µg (patients 2 and 3) to a maximum of approx. 2.6mg, as indicated. PSA values were monitored and T-cell activation was assessed by flow cytometry.
Fig.Figure 2Cytokine secretion of macrophages incubated with the indicated stimuli for 24 h. RBCL, iRBCL, XO were added to macrophages for 24 h. Each symbol represents the value obtained for cells from an independent donor in an independent experiment. Insets show paired samples by donor in two different conditions. Blue asterisks indicate significance when values were compared with RBCL (a) n=7 One-way ANOVA with Tukey test for multiple comparisons was performed to determine statistical significance (*p<0.05, **p<0.01 and ***p<0.001). Cytokine secretion of macrophages incubated with the indicated stimuli for 24 h. , XO, Hypoxanthine (Hyx) were added to macrophages for 24 h. Each symbol represents the value obtained for cells from an independent donor in an independent experiment. Insets show paired samples by donor in two different conditions. Blue asterisks indicate significance when values were compared with XO Grey asterisks mark comparison with Control n=3. One-way ANOVA with Tukey test for multiple comparisons was performed to determine statistical significance (*p<0.05, **p<0.01 and ***p<0.001). Cytokine secretion of macrophages incubated with the indicated stimuli for 24 h. XO added to macrophages for 24 h. Febuxostat (Feb) was pre-incubated for 30 minutes with XO before addition to macrophages. Each symbol represents the value obtained for cells from an independent donor in an independent experiment. Insets show paired samples by donor in two different conditions. Grey asterisks mark comparison with Control n=3. One-way ANOVA with Tukey test for multiple comparisons was performed to determine statistical significance (*p<0.05, **p<0.01 and ***p<0.001). Cytokine secretion of macrophages incubated with the indicated stimuli for 24 h. , XO the anti-oxidants N-acetyl-L-cysteine (NAC) and 1-Thioglycerol (1-TG), and heat inactivated (HI) XO were added to macrophages for 24 h. Each symbol represents the value obtained for cells from an independent donor in an independent experiment. Insets show paired samples by donor in two different conditions. Blue asterisks indicate significance when values were compared with XO Grey asterisks mark comparison with Control n=3. One-way ANOVA with Tukey test for multiple comparisons was performed to determine statistical significance (*p<0.05, **p<0.01 and ***p<0.001). Cytokine secretion of PBMC (e) incubated with the indicated stimuli for 24 h. RBC, iRBC, RBCL, iRBCL, XO Each symbol represents the value obtained for cells from an independent donor in an independent experiment. Insets show paired samples by donor in two different conditions. Blue asterisks indicate significance when values were compared with RBCL n=3. One-way ANOVA with Tukey test for multiple comparisons was performed to determine statistical significance (*p<0.05, **p<0.01 and ***p<0.001).
a–f, Histological analyses of Atg7flox/+; nestin-Cre (left) and Atg7flox/flox; nestin-Cre (right) cerebral cortex at P56.figf2a-f, Histological analyses of Atg7flox/+; nestin-Cre (left) and Atg7flox/flox; nestin-Cre (right) cerebral cortex at P56. Cryosections were stained with HE (a-d) or immunostained for the glial marker GFAP (e, f). Boxed areas in a and b are magnified in c and d, respectively. Arrows in c point to large pyramidal neurons in the cerebral cortex. g-j, Histological analysis of Atg7flox/+; nestin-Cre (left) and Atg7flox/flox; nestin-Cre (right) cerebellum at P56. Cryosections were stained with HE (g, h) or immunostained for the Purkinje marker calbindin (i, j). Arrows in g and h indicate cerebellar Purkinje cells. k-p, TUNEL staining of the cerebral cortex (k, l) and cerebellum (n, o) at P56 in Atg7flox/+; nestin-Cre (k, n) and Atg7flox/flox; nestin-Cre (l, o) sections. TUNEL-positive cells are indicated with arrows and shown as higher magnification images beneath panels l, n and o. Histograms show the average number (± s.d.) of TUNEL-positive cells in ten sections for three animals of each genotype (m, p). *, P 0.05 (t-test). Scale bars, 1 mm (a, b), 100 µm (c-j), 250 µm (k, l, n, o). We observed no sex difference in brain morphology or neuronal loss in Atg7flox/flox; nestin-Cre mice.
Figure 1Schematic representation of the experimental method used to identify lincRNAs upregulated in metastatic subpopulations isolated from HCT116 colon cancer cells. Gene ontology (GO) analysis of upregulated genes in metastatic lesions. Cell migration assays were performed using Transwell migration chambers. HCT116 cells were transfected with siRNAs targeting 11 candidate lincRNAs and cell migration activities were evaluated by Transwell migration assays (n = 3). CALIC is the lincRNA targeted by si-LINC00920. Migration of HCT116 cells transfected with the indicated siRNAs or negative control. (n = 4) GO analysis of differentially expressed genes in CALIC knockdown HCT116 cells. Migration of colon cancer cells treated with an siRNA targeting CALIC. (n = 3) (Top) Schematic representation of the chromosomal location of the CALIC gene locus. Arrows indicate the orientation of transcription. (Bottom) ENCODE ChIP-seq data for H3K4Me3 is displayed in the UCSC browser illustrations. Gray boxes mark locations of exons (Ex1-2) of CALIC. Subcellular localization analysis of CALIC. RNAs were isolated from the nuclear and cytoplasmic fractions of HCT116 cells and quantified by qRT-PCR (n = 3). Nuclear controls: U6, MALAT1; Cytoplasmic control: GAPDH. CALIC expression levels in cancer tissues. N, normal mucosa; T, tumor. Expression of CALIC in primary skin tumors (P) and their metastases (M). Expression of CALIC in primary skin tumors of patients without (w/o) or with (w) metastases. qRT-PCR analysis of CALIC expression in early (I/II) and late (III/IV) stage colon cancers. [Normal (N) n = 8, stage I/II n = 14, stage III/IV n = 26] Kaplan-Meier curves for disease-free survival of colon cancer patients whose primary tumors expressed low (blue, n = 113) or high (red, n = 250) levels of CALIC.
Figure 2A. Immunoblotting of c-MYC in A375 cells treated with NT-siRNA or siASNS for 24 hr followed by cycloheximide treatment for indicated timepoints (left). Plot showing relative protein levels of c-MYC (y-axis) as a function of time lapsed (x-axis) in the left panel (right). Immunoblotting of indicated proteins in melanoma cells 72 hr after treatment with NT-siRNA (-) or siASNS (#1 or #2) (B and C) Immunoblotting of indicated proteins in melanoma cells 72 hr after treatment with siASNS, PLX-4032, or both (D). E. Immunoblotting of indicated proteins in A375 cells cultured in L-Asn and treated with L-A'ase, PLX-4032, or both for 72 hr. F. A375 cells were treated with siASNS, SCH772984, or both for 72 hr followed by immunoblotting of phosphorylated and total GSK3-β. G. A375 cells were treated with siASNS, HA-GSK3-β (S9A), or both for 72 hr followed by immunoblotting of indicated proteins. H. RT-qPCR analysis of indicated transcripts in melanoma cells 36 hr after treatment with siASNS, HA-GSK3-β (S9A), or both. I. A375 cells were treated as indicated for 72 hr followed by immunoblotting of c-MYC. J. Immunoblotting of c-MYC and HA proteins in A375 cells treated as indicated for 72 hr.
Figure 6(A) Schematic illustration of the LAP-AID-tag integrated into the bacterial artificial chromosome containing the mouse Cdca5 locus.(B) Immunofluorescence staining of cells expressing sor-LAP-AID and the F-box helper protein after treatment with auxin. Antibodies against GFP mark the fusion protein and Aurora B antibody stains cells in G2/M-phase. Scale bar, 10 µm.(C) Quantification of immunofluorescence after treating cells with auxin. Error bars denote s.e.m.; a.u., arbitrary units.(D) Time course quantification of GFP immunofluorescence signal decrease after auxin addition. (E) Analysis of chromosome spreads after auxin treatment. (F) Western blot showing cell extracts after treatment with auxin.
Figure 1a. Fold change in mRNA expression of members of the PI3K/AKT/Foxo pathway in IFN+ as compared to IFN- Tregs in the microarray analysis. b. mRNA expression of members of the PI3K/AKT/Foxo signaling pathway in sorted IFN+ and IFN- Tregs. c. Sorted CD4+CD25highCD127- Tregs were stimulated in the presence of IL-2 (white) or IL-2+IL-12 (black dots) and RNA was isolated every 24 hours for 4 days. mRNA expression of members of the PI3K/AKT/Foxo signaling pathway. d. Representative example of the protein expression of AKT1, AKT2, AKT3, PTEN and FOXO3 on sorted CD4+CD25highCD127- Tregs stimulated in the presence of IL-2 (grey line) or IL-2+IL-12 (black line) and day 2 for AKT1, AKT2, AKT3 and PTEN and day 3 for FOXO3. Grey histogram represents the isotype. e. Statistical analysis of AKT1, AKT2, AKT3, FOXO3 and PTEN gMFI of n=6 experiments performed. * p<0.05, ** p<0.005, *** p<0.0005.
Figure 4The mRNA levels of SAMHD1 and TRIM21 in different tissues of neonatal mice were detected by RT-qPCR with GAPDH as a control, and the expression of SAMHD1 and TRIM21 in the heart was set as 100% (n=3, mean±SD, * P < 0.05, paired t-test). The protein levels of SAMHD1 and TRIM21 in different tissues of mice were examined by Near Infrared Spectrum (NIR) western blot with tubulin as a control. Immunohistochemical (IHC) staining results of SAMHD1 in various tissues of mice. Bars, 100μm. The mRNA levels of EV71 virus and host factors in mock or EV71 infected mice were detected by RT-qPCR with GAPDH as a control. The expression of EV71 (D) were presented, and the levels of corresponding gene in mock infected mice were set as 100% The mRNA levels of EV71 virus and host factors in mock or EV71 infected mice were detected by RT-qPCR with GAPDH as a control. The expression of , IFNα and IFNβ (E) were presented, and the levels of corresponding gene in mock infected mice were set as 100% The mRNA levels of EV71 virus and host factors in mock or EV71 infected mice were detected by RT-qPCR with GAPDH as a control. The expression of SAMHD1 and TRIM21 (F) were presented, and the levels of corresponding gene in mock infected mice were set as 100%