EMBO/sd-character-level-ner huggingface dataset:
Figure 6A) Live imaging movie of GFP-Sgo2 expressing oocytes (from GV onwards) undergoing meiosis I. The green channel to visualize GFP-Sgo2 localized to chromosomes is shown. Timepoints were taken every 10min, shown is a montage of selected frames such as indicated (hours: time after GVBD) at the metaphase-to-anaphase transition of meiosis I and in metaphase II. Each timepoint is comprised of an overlay of 11 z-sections of 3μm for GFP. The arrow heads indicate bivalents. In (A-C) n indicates the number of control non-injected and GFP-Sgo2-expressing oocytes analyzed B) Representative images of a chromosome spread showing Rec8 localization on a bivalent chromosome in metaphase II of a GFP-Sgo2 expressing oocyte. Spreads were stained with Rec8 antibody (pink), CREST serum (green), Hoechst for DNA (blue), GFP-Sgo2 was visualized by GFP fluorescence (grey). The scheme on the right shows a bivalent with individualized kinetochores and some Rec8 staining on arms. In (A-C) n indicates the number of control non-injected and GFP-Sgo2-expressing oocytes analyzed C) Metaphase I (4hrs after GVBD) and II spreads after GFP-Sgo2 expression (from GV onwards). Spreads were stained with CREST serum (red), Hoechst for DNA (grey), GFP-Sgo2 was visualized by GFP fluorescence (green). Arrowheads mark separated sister kinetochores. On the right, graph showing percentage of metaphase II spreads containing bivalent chromosomes. In (A-C) n indicates the number of control non-injected and GFPD) Chromosome spreads of oocytes expressing GFP-Sgo2 (from GV onwards) in metaphase II, and upon activation. Spreads were labeled with Rec8 antibody (shown in red, far-red secondary antibody was used), CREST serum (pseudo-colored in green), and Hoechst (blue). GFP direct fluorescence was imaged for GFP-Sgo2 detection. Below the corresponding chromosome figures are represented in the schemes.
Figure 1(A) Heat map showing a subset of mRNAs of genes immunoprecipitated by anti-EZH2 antibody in LNCaP and C4-2 prostate cancer cell lines, which was generated based on the distinct RIP-seq reads on specific gene exons. The units of the heatmap values were reads per kilobase million (RPKM). (B) Screen shots from the UCSC genome browser showing signal profiles of p53 mRNA immunoprecipitated by anti-EZH2 antibody in LNCaP and C4-2 cells. (C) C4-2 cells were subjected to UV-RIP assay. RT-qPCR measurement of p53 mRNA immunoprecipitated by IgG or anti-EZH2 antibody. Data shown as means ± SD (n=3). Statistical significance was determined by two-tailed Student's t-test. ** P<0.01. (D) Schematic diagram of four GST-EZH2 recombinant proteins (EZ1 to EZ4). EBD, EED binding domain; SANT1(/2) L, "Swi3, Ada2, N-Cor, TFIIIB"1(/2) like; MCSS, motif connecting SANT1L and SANT2L; CXC, cysteine-rich domain; SET, catalytic domain. (E) Top, RT-qPCR analysis of p53 mRNA in C4-2 cell lysate pulled down by GST or GST-EZH2 recombinant proteins EZ1-EZ4. Bottom, western blotting analysis of GST or GST-EZH2 proteins used for GST pull-down assay. Asterisks indicate the protein bands at expected molecular weight. Data shown as means ± SD (n=3). Statistical significance was determined by two-tailed Student's t-test.** P<0.01. (F) C4-2 cells were transfected with Myc-tagged EZH2-WT or EZH2-∆336-554 for 24 h and cells were harvested for RIP with IgG or anti-Myc tag antibody. Transfected proteins and pull-down p53 mRNAs were analyzed by western blot and RT-qPCR, respectively. Data shown are means ± SD (n=3). * P<0.01. ERK2, a loading control. (G) GST pull-down assay using in vitro transcribed different fragments of p53 mRNA and indicated GST proteins followed by RT-qPCR analysis of pull-down p53 mRNA. FL, full length; ORF, open reading frame; UTR, untranslated region. (H, I) RNA EMSA evaluation of EZh2 binding of p53 mRNA. GST-EZH2 recombinant proteins (EZ1 to EZ4) were incubated with biotin-labeled in vitro transcribed p53 5'UTR (biotin-labeled probe) in the presence or absence of 100-fold unlabeled p53 5'UTR (unlabeled probe), followed by PAGE and immune blotting with HPR-conjugated streptavidin.
Interactions between nCoV-RBD and ACE2.Figure 3Interactions between nCoV-RBD and ACE2.(A) The protease domain (PD) of ACE2 mainly engages the α1 helix in the recognition of the RBD. The α2 helix and the linker between β3 and β4 also contribute to the interaction. Only one RBD-ACE2 is shown. Detailed analysis of the interface between nCoV-RBD and ACE2. Polar interactions are indicated by red, dashed lines.
Figure 3Western blot analysis for CSTB in the CSF from 2 different E14 mouse embryos (A), in the CM from E14 cortical cells in culture for 4 days Western blot analysis for CSTB in the CM from hCOs after 4 days in culture (C). The CM was collected from hCOs at different time points in culture (d19, d38, d60, d80). Protein extract (PE) from 60 days hCOs was used as a positive control. GAPDH immunostaining was used as a negative control on hCOs CM. Ponceau red staining is shown to indicate the presence of proteins at comparable levels in the CM samples Micrograph of coronal sections of E17 mouse cerebral cortices electroporated at E14 co-electroporated with mCherry expressing vector and HA-empty vector or -Cstb, and analyzed 3 dpe. Immunostaining with RFP to identify electroporated cells and GFP to identify migrating interneurons in the GAD65-GFP transgenic mouse line. Ventricle (V) and cortical plate (CP) are indicated. The dashed lines represent the apical surface of the ventricles. Distribution of GFP+ interneurons in the 5 equal Bins of the mouse developing cortex-Bin1 corresponded to apical side and Bin5 to pial side. Ventricle (V) is indicated. The dashed lines represent the apical surface of the ventricles. Micrograph of coronal sections of E17 mouse cerebral cortices electroporated at E14 co-electroporated with mCherry and HA-Cstb expressing vector, analyzed 3 dpe. Immunostaining with RFP to identify electroporated cells and GFP to identify migrating interneurons in the GAD67-GFP transgenic mouse line. Ventricle (V) and cortical plate (CP) are indicated. The dashed lines represent the apical surface of the ventricles. Distribution of GFP+ interneurons in the 5 equal Bins of the mouse developing cortex comparing electroporated sides with the contralateral ones in the same section to avoid different distribution of interneurons rostro-caudally.
Figure 1Silver staining of protein components in the Ago3 complex immunoisolated from BmN4 cells. P150 and P130 co-immunoprecipitated with Ago3. n.i.: non-immune IgG. Western blots showing the presence of Vret-L (P150) and Vret-S (P130) in the Ago3 complex in A. n.i.: non-immune IgG. Top: western blot showing Vret-L and Vret-S immunoprecipitated from BmN4 cells upon UV irradiation. Bottom: CLIP signals showing that Vret-L, but not Vret-S, binds RNAs in vivo. Five upper panels: Northern blots showing the levels of RT3-piRNA, Bmmar6-piRNA, R2Bm-piRNA, and R1Bm-piRNA in normal (Control), Vret-L/S-depleted (Vret KD), Vret-L-depleted (Vret-L KD), and Vret-S-depleted (Vret-S KD) cells. Bantam (miRNA) was detected as a loading control. Two lower panels: western blots showing the efficiency of the Vret knockdown. β-Tub was detected as a loading control. Two upper panels: western blots showing the abundances of Flag-Vret-L and Ago3 in both samples. Note that Flag-Vret-L is present only in the Vret-L complex but not with Ago3 isolated from total cell lysates (Fig EV1F). Bottom panel: Northern blot showing the level of PiggyBac-piRNA associated with Ago3 in total lysate (Ago3-IP) and in the Vret-L complex (Flag-Vret-IP). The ratio of the piRNA signal intensity in the two samples is indicated at the bottom. Two upper panels: western blots showing the abundances of Flag-Vret-L and Siwi in both samples. Bottom panel: Northern blot showing the level of RT3-piRNA associated with Siwi in the total lysate (Siwi-IP) and in the Vret-L complex (Flag-Vret-IP). Boxplot showing log2 fold-change of Vret-KD (RPM)/Control (RPM) values of piRNAs in increased, unchanged and decreased groups defined by Ago3-KD (RPM)/Control (RPM) values shown in Fig EV1I. Boxplot represents median, first and third quartile, and maximum and minimum values of the log2 fold-change of piRNAs (N = 2). Outliers are represented as open circles. Flag-EGFP and -Vret-L WT and mutants were immunoprecipitated using the anti-Flag antibody and then western blotting was performed using anti-Flag, anti-Siwi, anti-Ago3 and anti-Vret antibodies. The domain structures of Tud1mut (Y753A/Y757A/N759A) and Tud2mut (Y973A/ Y976A/N978A) are shown.
Figure 4 A-B SH-SY5Y cells were immunostained for ATP5a (green) and 20S proteasome core (red) after IU1 treatment (6h-24h). Scale bar 10 μm. Colour mapping of the co-localisation of HSP60 and 20S proteasome from the selected regions were generated by image j and represented in the bottom panel. Intensity of the co-localisation is represented in the colour map scale at the bottom of the panel. (B) 20S proteasome subunit and mitochondria co-localisation was quantified by measuring Mander's coefficient. Bar graphs represent mean fold change (mean ± SEM) from 3 different experiments. At least 30 cells were evaluated for the calculations. ANOVA followed by Dunnet's test  C Electron microscopy images showing subcellular localization of immuno- gold labelled 20S subunit of proteasome complex after 12h or 24h of IU1 treatment. Experiments were repeated twice in two biological replicates with similar results. The contrast in the magnified region is enhanced from the original image to highlight the immunogold labelled signals  D-E Protein proximity ligation assay for Prohibitin2 (PHB2) and LC3 after 24h IU1 treatment. Images showing signal are merged with bright field images to count the signal dots (cyan false colour) as depicted in the figure. In the inset only fluorescence signal is shown, without merging with the bright field from the selected regions. Scale bar 10 μm. (E) Bar graphs represent mean ±SEM of the number of signal dots per cell from 3 different experiments. Student's t test; *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001
Comparison of several methods of drug candidate identification.Figure 2 Five methods of transcriptome analysis / drug candidate identification were compared in their ability to successfully produce drug targets in at least one prior study showing a survival benefit from sepsis. 1) Pathprint-PDN: Comparison of pathways by Pathprint and drug candidate analysis by pathway-drug network (PDN); 2) DEGs-PDN: Comparison of differentially expressed genes (DEGs) by standard methods and drug candidate analysis by PDN; 3) Random: Drugs chosen at random from the CMap database; 4) DEGs-LINCS: Comparison of DEGs generated by standard methods and drug candidate analysis using LINCS database and 5) BarCode-LINCS: Comparison of DEGs generated by BarCode method and drug candidate analysis using LINCS database. The three gene-level methods were found to be no better at generating positive drugs than picking drugs at random. All methods produced significantly lower percent positive rates than the Pathprint-PDN method (p<0.02). Prism software (GraphPad) was used to compare the frequency of prior studies showing benefit for drug leads Fisher"s exact tests
Figure 5(A) Schematic representation of ARSA activity in the brain of NHP S1.1 and S2.2 along the rostro-caudal axis (slices 2-9). Data are expressed as fold increase to ARSA activity detected in matched blocks of UT and LV.GFP-injected NHPs (physiological levels). Grading of colors ranges from light yellow (<1.2 fold) to dark green (≥ 2 fold). (B-C) Enzymatic activity measured in single brain slices of NHPs of study group 1 (B) and study group 2 (C) (filled grey bars, injected hemisphere; striped grey bars, contralateral hemisphere), and in UT and LV.GFP-injected NHPs (white bars; physiological levels). Arrows on x axis indicate the injection sites. n=2-12 tissue blocks/slice. Number of tissue blocks analyzed per animal: 51 (UT/LV.GFP), 98-122 (S1.1 and S2.2), 12-22 (S1.2, S1.3, S2.2, S2.3). Two-way ANOVA followed by Bonferroni's multiple comparison tests, *p<0.05; **p<0.01; ***p<0.001 vs matched slices of UT/LV.GFP. (D) Summary of ARSA activity in the injected and contralateral hemispheres of NHPs in study group 1 and study group 2. Number of tissue blocks analysed: 51 (UT/LV.GFP), 105 (study group 1, injected hemisphere), 52 (study group 1, contralateral hemisphere); 103 (study group 2, injected hemisphere), 72 (study group 2, contralateral hemisphere). One-way ANOVA and Tukey's multiple comparison tests, *** p< 0.001 versus UT/LV.GFP, §p<0.05 versus study group 1. (E) ARSA activity in the spinal cord of individual LV.GFP- (n=4 blocks/animal, 2-3 replicates) and LV.hARSA-injected NHPs (n=2-4 blocks/animal; n=10 and n=11 total blocks for study group 1 and study group 2, respectively). Graph on the right represent the summary of ARSA activity in each treatment group. One-way ANOVA and Dunnet's multiple comparison test. * p<0.05 versus pilot group. (F) ARSA activity in the sciatic nerve of LV.GFP- (n=2) and LV.hARSA-injected NHPs (n=3). In B-F data are expressed as floating bars (line at mean). (G) ARSA activity in the cerebrospinal fluid (CSF) of UT/LV.GFP (n=3) and LV.hARSA-injected NHP (n=5). Data are expressed as mean±SEM. Student t-test, *p=0.035.
Figure 8SATB2 binds to the MAR region of its target gene FOXM1 to remodel local chromatin by recruting histone acetyltransferase CBP, which in turn promotes FOXM1 transcription, leading to GSC proliferation and GBM tumor growth. Inhibition of SATB2/CBP transcriptional activity by the CBP inhibitor C646 suppressed GSC proliferation and tumor growth. Reprinted with permission, Cleveland Clinic Center for Medical Art & Photography © 2020. All Rights Reserved.
Fig.Figure 1Quantitative RT-PCR shows that Lpin1 expression is depleted in the skeletal muscles Gastrocnemius (GC) and Tibialis Anterior (TA), but not in White Adipose Tissue (WAT) from 3-month-old HSACre/+/Lpin1fEx2-3/fEx2-3 mice (HSA-Cre) compared to control mice. Expression levels are corrected for expression of the control gene Pinin and presented in the graph as fold change relative to control mice. PAP1 and PAP2 activities were respectively substantially decreased and increased in TA, of HSA-Cre mice as compared to control mice, respectively (n = 6; *P<0.05). Measurements of muscle/body weight ratio of GC and TA of 3-month-old HSA-Cre and control mice. Hematoxylin and Eosin (HE) staining of representative transverse sections of GC muscles from 3-month-old HSA-Cre and control mice. The black arrows indicate fibers with centrally located nuclei and yellow arrows indicate mononuclear cell infiltration. Scale bars: 40 μm Relative force measurements after normalization of absolute tetanic force to GC muscle weight from 3-month-old HSA-Cre and control male mice.
Figure 1A PLX4032 dose-response curves of parental and resistant melanoma cells. Relative cell viability, compared to DMSO control, was measured by CCK8 assay after PLX4032 treatment for 72 hr. B Sequencing results confirming BRAF V600E mutation (c.1799T>A) in resistant cells. C Immunofluorescence images showing BrdU incorporation in parental and resistant WM3248 cells after PLX4032 (2 &amp;amp;#x3BC;M) or DMSO treatment for 24 hr. D Quantification of the experiment presented in (C). Both SKMEL28 and WM3248 cells were analyzed. E qRT-PCR analysis comparing SOX10 and MITF expression levels in parental and resistant cells. F Immunoblotting for the indicated proteins. Cells were plated and incubated without drugs for 24 hr, and then treated with PLX4032 (2 &amp;amp;#x3BC;M) or DMSO for 24 hr.
Figure 2A, B MICAL2PV is down-regulated in H1299 (A) and A549 (B) cells. H1299 and A549 cells stably expressing shRNAs against MICAL2PV (shM2) or a control shRNA (shCtr) were examined by Western blotting using an anti-MICAL2PV antibody. MICAL2PV mRNA levels were detected by quantitative RT-PCR. Data information: data were analyzed by one-way ANOVA with Bonferroni post-test and shown as mean ±SEM; *** P < 0.001. Three independent experiments were performed C, D TNT formation (marked by black triangles and white arrowheads) was imaged in control and shM2 H1299 (C) and shM2 A549 (D) cells by confocal microscopy. Scale bar: 60 μm. Boxed areas are enlarged and shown at the bottom of the corresponding images. E, F Quantification of the percentage of cells forming TNTs in control and shM2 H1299 (E) and shM2 A549 (F) cells. 13 images per group (E) and 10 per group (F) were captured for quantification. Three independent experiments were conducted. G Data information: data were analyzed by one-way ANOVA with Bonferroni post-test and shown as mean ±SEM; *** P < 0.001. Three independent experiments were performed G H1299 cells were transfected with a GFP-tagged MICAL2PVb vector (GFP-M2) or control vector (Ctr) followed by Western blotting with an anti-GFP antibody. The mRNA levels of MICAL2PV were measured by quantitative RT-PCR. Three independent experiments were conducted. H,I H1299 cells stably expressing the Ctr or GFP-M2 were cultured in the presence of 20 μM 2-DG for 24 h to induce TNT formation. TNTs were imaged by confocal microscopy (A) and the percentage of cells forming TNTs was quantified (B). n=21, 20 and 19 images were captured from Ctr, GFP-M2#1 and GFP-M2#2, respectively. Three independent experiments were performed. Scale bar: 60 μm. Data information: data were analyzed by one-way ANOVA with Bonferroni post-test and shown as mean ±SEM; *** P < 0.001. Three independent experiments were performed
Chemical inhibition of CPAP-Tubulin interaction causes interphase centrosomes to recruit enhanced levels of PCM.Figure 5(A) Interphase centrosomes of CCB02-treated two-centrosome containing MCF10A cells display an enhanced level of PCM proteins. CPAP (green) intensity remains same. Cells are stained for CPAP (green), PCM proteins (magenta) Cep152 (i), PCNT (ii) and CDK5RAP2 (iii) and microtubules (α-tubulin, red). Scale bars, 2µm for left panels and 0.2µm for right panels. Heat map images show intensity saturation (blue) for CPAP, Cep152, PCNT and CDK5RAP2 upon CCB02 treatment. Line graphs at right to each panels indicate the heat map intensity of single centrosomes of cells (given in boxes at heat map panels) treated with vehicle (blue line) or CCB02 (red line). Images were recorded with low laser excitation and high zoom factor (B) Bar graph quantifies relative intensity of CPAP-interacting proteins in interphase centrosomes compared to vehicle treatment. At least 150 centrosomes (for vehicle); 130 centrosomes (for CCB02) were analyzed in each condition. Error bars, mean ± SEM. (N)=3. Ordinary Two-way ANOVA test ***P < 0.0001 (C) Immunopurification of CPAP complexes from CCB02-treated cell extracts. (i) CCB02 specifically prevents CPAP-Tubulin interaction. This allows CPAP to bind an enhanced amount of Cep152 and γ-tubulin. Control IP experiment is shown below. (ii) Plot showing PCM protein intensities co-immunoprecipitated with CPAP at various concentrations of CCB02 treatment. CPAP amount does not change with CCB02 treatment. Data represent mean ± SEM. (N)=3.
Figure 1(a) Phylogenetic tree of the 12-member synthetic ecology spanning the major phyla in the gut microbiome including Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria. Phylogenetic analysis was performed using a concatenated alignment of single-copy marker genes obtained via Phylosift(Darling et al, 2014). Maximum likelihood trees were generated using default options. The scale bar represents the number of substitutions per site in the alignment. (b) Schematic of the experimental design for this study. Species were combined using an approximately 1:1 or 19:1 initial proportion based on absorbance measurements at 600 nm (OD600) into microtiter plates using liquid-handling robotic manipulation. Approximately every 12 hr, samples were collected for multiplexed 16S rRNA gene sequencing (black circles). Relative abundance was measured using multiplexed 16S rRNA gene sequencing of the V3-V4 region using dual-indexed primers compatible with an Illumina platform (stacked bar plot, bottom right). Serial transfers were performed in approximately 24 intervals (purple bars, top) by transferring an aliquot of the communities into fresh media using a 1:20 dilution. In parallel, time-resolved OD600 measurements of monospecies and consortia were performed.
Fig.Figure 3A Experimental paradigm: Tamoxifen (TAM) was applied i.p. every 12 hours for five days to control and β-catex3 iDCX mice and animals were analyzed 3 and 13 days post Tamoxifen (dpT). B Representative reconstructions of control and β-catex3 iDCX neurons at 3dpT. Scale bar= 20µm. C Analysis of morphology showed increased dendritic length in β-catex3 iDCX neurons at 3dpT [p=0.0320]. The number of branch points was unaltered [p=0.0651]. Sholl analysis indicated increased growth of terminal dendrite branches in neurons of β-catex3 iDCX mice [p<0.0001] (control: n = 25 cells from 9 animals; β-catex3 iDCX: n = 25 cells from 8 animals). D Analysis of morphology showed no difference in dendritic length [p=0.1029] and branch point number [p=0.3726] between experimental groups at 13dpT. Sholl analysis indicated a decrease in length of terminal dendrites in neurons of β-catex3 iDCX mice [p<0.01] (control: n= 23 cells from 9 animals, β-catex3 iDCX: n= 26 cells from 8 animals). E Spine densities were comparable between experimental groups at 3dpT [iML p=0.1512, oML p=0.3841]. 13dpT β-catex3 iDCX neurons showed significantly increased spine density in the inner molecular layer [iML p<0.0001 , oML p=0.0402] (3dpT: control: n= 20 cells from 3 animals, β-catex3 iDCX: n= 20 cells from 4 animals; 13dpT control: n= 20 cells from 5 animals, β-catex3 iDCX: n= 20 cells from 7 animals). Representative images showing dendritic segments in the inner molecular layer of the DG at 13dpT. Scale bars= 1µm. F Representative images of recombined (GFP+) neurons co-expressing Prox1 (grey) as a marker for neuronal fate and the stage specific markers DCX (red) for immature neurons and Calbindin (red) for mature neurons. Arrows and arrowheads indicate marker negative and marker positive cells, respectively. Scale bar = 10µm. G Fraction of DCX expressing neurons is slightly reduced in β-catex3 iDCX at 3dpT [p=0.0338], while fraction of Calbindin expressing cells is slightly increased in β-catex3 iDCX at 3dpT [p=0.0345]. At 13dpT no difference is visible in stage specific marker expression [DCX p=0.8564, Calbindin p=0.0841] (3dpT: control: n= 12 animals, β-catex3 iDCX: n= 12 animals, 13dpT: control: n= 10 animals, β-catex3 iDCX: n= 14 animals).