EMBO/sd-character-level-ner huggingface dataset:
| The EV vsiRNAs of NoV induce homology-dependent virus resistance in AG6 mice.Figure 5A, Schematic overview of antiviral function of EVs. Briefly, EVs from naïve or NoVΔB2-infected BALB/c suckling mice were extracted 3 days post infection, and then injected into AG6 suckling mice. After 12 hours post immunization, AG6 suckling mice were challenged by WT NoV and then sacrificed at 1dpi. B, NoV replication was determined by RT-qPCR from hindlimb muscle of AG6 suckling mice challenged with NoVΔB2 as control or EVs from naïve or NoVΔB2-infected BALB/c suckling mice at 3 dpi. Normalization was done by β-actin mRNA levels. * indicates p<0.05 by student's t-test. n=3~4 per group. Data are shown as mean±SD. C, Relative viral RNA1 accumulation levels at 1dpi in NoV WT-infected AG6 mice 12 hours after injection with EVs from NoVΔB2 or mock-inoculated BALB/c mice were measured by RT-qPCR. Normalization was done by β-actin mRNA levels. **** indicates p<0.0001 by student's t-test. n=5~6 per group. Data are shown as mean±SD. D, Diagram of recombinant SINVNoV (top) and SINVGFP (bottom). The gene of B2 protein was inserted at downstream of the duplicated subgenomic promotor sequence as shown. E, AG6 suckling mice were challenged with EVs from naïve or NoVΔB2 -infected BALB/c suckling mice and then inoculated by SINVNoV and SINVGFP respectively. Viral titer (PFU/ml) in the hindlimb muscle of infected mice was measured by a standard plaque assay and normalized by tissue mass, and the viral genomic RNA accumulation was determined by RT-qPCR amplification of the viral nsP1 coding region. Normalization was done by β-actin mRNA levels. *** indicates p<0.001, ns indicates no significant difference by student's t-test, n=5 per group. Data are shown as mean±SD.
FTLD patient-derived TDP-43 aggregates seed neoaggregates in cells with subtype-specific potency A) Seeding efficiency was compared between control non-neurodegenerative and FTLD-TDP patient samples.Figure 1A) Seeding efficiency was compared between control non-neurodegenerative and FTLD-TDP patient samples. Cells were transfected using Lipofectamine after mitotic arrest with AraC and were fixed 3 to 6 days post-transfection with SarkoSpin fractions (Laferrière et al, 2019). Surface masks generated with Imaris show DAPI (blue), nuclear TDP-43 (NuclTDP-43, cyan) and neoaggregates (NeoAgg, orange). All images are presented at the same scale, 20 μm as indicated in AraC panel Day 6. B) Higher magnification imaging of cells seeded for 6 days revealed presence of large cytoplasmic TDP-43-HA aggregates in cells seeded with FTLD-TDP-A and multiple cytoplasmic TDP-43-HA aggregates in cells seeded with FTLD-TDP-C. Surface representation generated with Imaris showing DAPI , TDP-43-HA (green), phosphorylated TDP-43 (pTDP-43, magenta) and neoaggregates (NeoAgg, orange). All images are presented at the same scale, 5 μm as indicated in FTLD-TDP-C top panel. C) Quantification of the number of neoaggregates over time (n=3 independent experiments, with multiple biological replicates: non-neurodegenerative control (n=5), FTLD-TDP-A (n=7) and FTLD-TDP-C (n=6) ). Whiskers in the box plot represent median and minimum/maximum values. 2-way ANOVA repeated measure, with post-hoc Fisher's LSD, F(3,15)=1.3, p=0.3110, NS difference for Day 3 and 4. Day 5: Non-neurodegenerative control vs FTLD-TDP-A p=0.0344, non-neurodegenerative control control vs FTLD-TDP-C p=0.4606. Day 6: non-neurodegenerative control vs FTLD-TDP-A p=0.1028, non-neurodegenerative control vs FTLD-TDP-C p=0.2919. D) Linear regression model of neoaggregate formation over time. Best-fit values: non-neurodegenerative control control (y=0.06902x-0.1281, R2=0.3655, p=0.0048), FTLD-TDP-A (y=0.1810x-0.4836, R2=0.3532, p=0.0009), FTLD-TDP-C (y=0.1902x-0.5896, R2=0.2581, p=0.0112). Null-hypothesis slope=0.06902: non-neurodegenerative control vs FTLD-TDP-A p=0.0278, non-neurodegenerative control vs FTLD-TDP-C p=0.0918. E) Comparison of pTDP-43 score to neoaggregate formation per patient showed positive correlation for FTLD-TDP-A and no correlation for FTLD-TDP-C. Trendline is shown per subtype. Best-fit value of linear regression model: FTLD-TDP-A (y=1.286x+1.123, R2=0.6099, p=0.0667), FTLD-TDP-C (y=-0.2655x+2.805, R2=0.03133, p=0.7758).
BMP activity caudalizes ectoderm to spinal cord fates.Figure 4One-cell stage embryos were injected animally with increasing concentrations of bmp4 encoding mRNA (5-50 pg). AC explants were removed from control and injected embryos at blastula stage 9, and explants were cultured to neurula stage 16. Total RNA was isolated from five control embryos (lane 2), and from eighteen ACs from each group (lanes 3-7). Neural, epidermal and mesodermal markers were examined by sqRT-PCR: hindbrain (krox20), spinal cord (hoxb9, hoxc10/d10, cdx1-2-4), epidermal cytokeratin (E13), lateral mesoderm (osr1-2) and dorsal-lateral mesoderm muscle actin (MA). ODC is the control for quantitating the RNA levels. -RT-PCR was performed on total RNA isolated from control embryos (lane 1). One-cell stage embryos were injected animally with mRNAs encoding the bmp4 (50/150 pg) and/or meis3 (0.7 ng) proteins. AC explants were removed from control and injected embryos at blastula stage 9, and explants were cultured to neurula stage 16. Total RNA was isolated from five control embryos (lane 2), and from eighteen ACs from each group (lanes 3-8). Neural and mesodermal markers were examined by sqRT-PCR: hindbrain (krox20), spinal cord (hoxb9, hoxc10, cdx2), and lateral mesoderm (osr1). Ef1α is the control for quantitating the RNA levels. -RT-PCR was performed on total RNA isolated from control embryos (lane 1). One-cell stage embryos were injected animally with increasing concentrations of bmp4 encoding mRNA (75-200 pg). AC explants were removed from control and injected embryos at blastula stage 9, and explants were cultured to gastrula stage 11-11.5. Total RNA was isolated from five control embryos (lane 2), and from eighteen ACs from each group (lanes 3-6). Marker expression was determined by sqRT-PCR. Xenopus brachyury (xbra) expression was induced by BMP4 in a dose-dependent manner. Marker expression was determined by sqRT-PCR. Neither dorsal (chd, goosecoid-gsc) or ventral-lateral (osr1) mesoderm markers were induced by BMP4. Sox2/3 neural markers are expressed in the ACs and are unchanged by BMP expression. EF1α is the control for quantitating RNA levels. -RT-PCR was performed on total RNA isolated from control embryos (lane 1).
Figure 4(A) Number of proteins identified after analysis by mass spectrometry in the six fractions of EVs from control and HIV-infected cells. The majority of proteins were quantified in both samples. (B) Proteomic profiles of different proteins showing the relative abundance distribution across the six subfractions. Each profile consists of two independent data triplets (F1- F2- F3A and F1- F2- F3B). Abundance profiles of the same groups of proteins showed the same clustering behaviour, although in this second dataset, the proportion of proteins recovered in the F2 was more variable. These differences, however, did not affect the results of the NNP or movement analysis. (C) Unbiased identification of significant translocation events triggered by HIV-1 infection. Each protein is scored for magnitude of translocation (M score, x-axis) and reproducibility of translocation direction (R score, y-axis) across the two replicates. MR plot analysis reveals significant translocations in the top right quadrant. Proteins with M>1 and R>0.9 are candidate hits for changing localization (estimated FDR = 8%). (D) Movement of the 26 proteins identified as top hits for significant and reproducible translocation upon HIV-1 infection Superimposition of two PCA plots with top hits in black and key hits (MOV10, SPN, SERINC3) highlighted with different colours. The position of each protein is shown both in non-infected (circle) and HIV-1 infected (red triangle) conditions. Arrows indicate movements of candidate proteins upon infection.
a) Filter trap assay analysis of HTT-polyQ aggregation in nine R6/2 mouse brains injected with three different combinations of rAAV.figf5(a) Filter trap assay analysis of HTT-polyQaggregation in nine R6/2 mouse brains injected with three different combinations of rAAV. (b) Quantification of the SDS-insoluble material detected by filter trap assay (*, P = 0.037; **, P = 0.0083; ***, P = 0.0015). (c) Western blot analysis of HTT aggregation (gel top, EM48 antibody), soluble HTT (1C2 antibody), and higher molecular polyQ complexes by agarose gel electrophoresis for resolving aggregates (AGERA). Full-length western blots are presented in Supplementary Figure 13. (d) Quantification of the gel top-detected aggregation (*, P = 0.093; **, P = 0.0027; ***, P = 0.0045). (e) Quantification of soluble HTT-polyQ (*, P = 0.034; **, P = 0.00003; ***, P = 0.0001). (f) Brain sections stained with anti-RFP, EM48 and anti-ubiquitin antibodies (bar, 40 μm). (g) Numbers of ubiquitin-positive inclusions in the transduced areas (*, P = 0.0026; **, P = 5.535 × 10−8; ***, P = 2.009 × 10−8). Bars in b, d, e and g represent the relative mean values ± s.e.m. from three brains injected with different rAAV in each striatum. R in R/Q and R/HQ, and Q in Q/HQ brains represent the control value of 1.
Figure 8A. HEK 293T cells were transfected with pCS2 vector (black), pCS2-Endouc (red), and pCS2-EndoucK242A (green) for 24 hr and subjected to either DMSO (control) or thapsigargin (TH; stress) treatment for 1 hr. The lysates of transfected cells were applied to polysome profile analysis (PPA). The sucrose gradient from top to bottom was shown from left to right, respectively. The sedimentation of 40S, 60S, 80S, and polysomes was indicated. Lysate from control, Endouc-expressing, and EndoucK242A-expressing cells were applied to PPA. The resulting fractions were TCA-precipitated, analysed by SDS-PAGE, and subjected to Western blot using antibodies against Flag and S6. B, The lysates from control (pCS2), Endouc-expressing (Endouc), and EndoucK242A-expressing (EndoucK242A) cells were applied to PPA. The resulting fractions from 1-10 collected from sucrose gradient were subsequently subjected to RT-qPCR assay to quantify (B) exogenous huORFchop-tagged luc (luc mRNA) The distribution of relative abundance of luc and CHOP transcripts contained in each fraction was determined. C. The lysates from control (pCS2), Endouc-expressing (Endouc), and EndoucK242A-expressing (EndoucK242A) cells were applied to PPA. The resulting fractions from 1-10 collected from sucrose gradient were subsequently subjected to RT-qPCR assay to quantify (C) endogenous CHOP transcripts (CHOP mRNA). The distribution of relative abundance of luc and CHOP transcripts contained in each fraction was determined. D, HEK 293T cells were co-transfected with pCS2-Endouc and phuORFchop-luc for 24 hr and then subjected to DMSO or TH treatment for 2 hr. The lysates of transfected cells were then applied to PPA. Total RNAs were isolated from heavy polysome fractions followed by performing 5`RACE to characterize the nucleotide sequences of truncated (D) exogenous huORFchop-luc The first sequence at N-terminus of RNA samples from the truncated forms derived from exogenous huORFchop-luc (n=14) and endogenous huORFchop-CHOP transcripts (n=11) was determined; the percentages of positive colonies containing various 5'UTR lengths (truncated forms) of luc and CHOP transcripts among total colonies examined were calculated. E. HEK 293T cells were co-transfected with pCS2-Endouc and phuORFchop-luc for 24 hr and then subjected to DMSO or TH treatment for 2 hr. The lysates of transfected cells were then applied to PPA. Total RNAs were isolated from heavy polysome fractions followed by performing 5`RACE to characterize the nucleotide sequences of truncated (E) endogenous huORFchop-CHOP transcripts. The first sequence at N-terminus of RNA samples from the truncated forms derived from exogenous huORFchop-luc (n=14) and endogenous huORFchop-CHOP transcripts (n=11) was determined; the percentages of positive colonies containing various 5'UTR lengths (truncated forms) of luc and CHOP transcripts among total colonies examined were calculated.
Figure 5A, B, C. mRNA levels of indicated genes in ISEMFs from wild-type (n=4) and Angptl2-/- (n=3) mice based qRT-PCR analysis. D. Representative staining for P-Smad1/5 in crypts from wild-type and Angptl2-/- mice following DSS treatment for 6 days and then untreated water for 6 more days (DSS) or untreated water for 12 days (Untreated). Dashed lines indicate half of the crypt and arrowheads show P-Smad1/5-positive cells in lower halves of crypts. Scale bar = 50 μm.E. The number of P-Smad1/5-positive cells in the crypt base (n=60) from wild-type and Angptl2-/- mice following DSS treatment for 6 days and then untreated water for 6 more days (DSS) or untreated water for 12 days (Untreated). Multiple areas in 3 mice per genotype were quantified. F. Western blotting analysis of isolated colon crypts from wild-type and Angptl2-/- mice. HSC70 serves as an internal control. Numbers below panels represent normalized protein expression.
Figure 7-F Representative kymographs depicting EB1 comet movement patterns in the proximal dendrites of ddaC neurons at 96 h AEL. Horizontal arrow indicates the direction towards the somas, and vertical arrow indicates the time. (B) RNAi knockdown of klp10A in msps810/mspsP18 mutants restored the retrograde movement pattern of EB1 comets. MT depolymerization or severing induced by Colchicine treatment (D) or overexpression of Kat-60 (F) led to mixed MT orientation in ddaC neurons. G-J Quantitative analysis of the percentages of anterograde EB1 comets, the track length, number (per 30 µm) and the velocity of EB1 comets in each neuron imaged.
p300/ CBP can succinylate H3K122, SIRT5 and SIRT7 desuccinylate it.Figure 3A H3K122succ levels (detected by immunoblot), relative to the scramble siRNA control, upon depletion of indicated HAT enzyme(s) (GCN5, pCAF, p300 and/or CBP) from MCF7 cells by siRNA. The bar graph shows a representative experiment B In vitro succinyltransferase assay on recombinant histone octamers as substrate. After incubation with p300 and suc-CoA, the H3K122succ levels were assessed by immunoblot. Ponceau staining of membrane is shown as a loading control. C In vitro succinyltransferase assay on unmodified peptides spanning H3K122. Samples were incubated with increasing amounts of p300 (0 to 160 ng) in the presence of radiolabeled suc-CoA. The radioactivity incorporated was quantified by liquid scintillation counting. The plot displays average counts per minute (CPMs), ± SD and individual datapoints of two independent replicates. In vitro desuccinylation assay on H3K122succ peptides with recombinant SIRT7 (D) Unmodified H3K122 peptide was used as control (left dot). Peptides were probed with H3K122succ AB #2 after incubation with sirtuins. In vitro desuccinylation assay on H3K122succ peptides with recombinant SIRT5 (E). Unmodified H3K122 peptide was used as control (left dot). Peptides were probed with H3K122succ AB #2 after incubation with sirtuins. F H3K122succ levels in Sirt5-KO cells. Histones were acid extracted from wild type (WT) and Sirt5-KO MEFs and the levels of H3K122succ determined by immunoblot. The level of H3K122succ was normalized to histones extracted from WT MEFs.
The action of bicarbonate and Ca2+ on longitudinal rolling of human sperm.Figure 3Experimental setup to subject optically trapped sperm to different conditions in a three-channel microfluidic capillary. Rotation frequency of a trapped sperm cell before and after dragging across the barrier stream. The green trace indicates the fluorescence of fluorescein included in the barrier stream. Error bars indicate the full width at half prominence of the frequency peaks determined by the fast Fourier analysis. Paired plot of rotation frequencies of individual sperm cells before and after dragging across the barrier stream. Rotation frequency of a trapped sperm cell before and after dragging from the control stream containing 0 mM bicarbonate into the stimulus stream containing 25 mM bicarbonate. Error bars indicate the full width at half prominence of the frequency peaks determined by the fast Fourier analysis. Paired plot of rotation frequencies of individual sperm at 0 and 25 mM bicarbonate. Rotation frequency of a trapped sperm cell in the presence and absence of extracellular Ca2+. Error bars indicate the full width at half prominence of the frequency peaks determined by the fast Fourier analysis. Paired plot of rotation frequencies in the presence and absence of extracellular Ca2+. Fraction of motile sperm (mean ± SD) in a sperm population incubated in the presence (black) and absence (at t = 0) of extracellular Ca2+ (red) (n ≥ 5). Distribution of rotation frequencies in populations of freely swimming sperm in the presence (black, n = 335) and absence (red, n = 224) of extracellular Ca2+ (red).
Figure 7(A-H) Graphs plotting predicted versus measured phenotypes are shown for the best-performing whole-transcriptome and metabolome models for each phenotype, based on the pooled R2 scores in Table 2. (A) Transcriptome e-net model for leaf 16 blade length, (B) metabolome e-net model for leaf 16 blade length, (C) transcriptome e-net model for leaf 16 blade width, (D) metabolome e-net model for leaf 16 blade width, (E) transcriptome e-net model for husk leaf length, (F) metabolome random forest model for husk leaf length, (G) transcriptome e-net model for ear length, (H) metabolome random forest model for ear length. The dot colors represent different outer cross-validation folds. Perfect predictions are located on the diagonal line in each panel.
Figure 4A. The vgrG genetic modules of the tested A. tumefaciens strains. Genes are color coded according to the predicted function or results of functional assays [24]. The vgrG-associated genes are v1-v6 for 12D1, v1-v9 for 1D1108, v1-v7 for 15955. B. Secretion assay of wild type and various mutants of 12D1, 1D1108 and 15955. C. Antibacterial activity assay of wild type and mutants of 12D1, 1D1108 and 15955 was carried out in a ratio of 30:1 against E. coli harboring the plasmid pRL662. The target E. coli cells were serially-diluted and grown overnight on gentamicin-containing LB agar prior to photographing. Each competition was done at least four times and reproduced in three independent experiments. D. Secretion assay of wild type 12D1, ΔtssL, Δv1, Δv3-4 containing pRL662 empty vector (V) or its derivatives with v1 or v3-4 were analyzed for secretion. E. Secretion assay of wild type 1D1108, ΔtssL, Δv4, Δv6 containing pRL662 empty vector (V) or its derivatives with v4 or v6.
Figure 6A. Map of SNPs positions observed in each sequenced genome after 48 hours adaptation (n=9) and 5 days adaptation with 1 passage per day (n=6). SNPs were mapped to the parent strain, sequenced after growth in minimal media in the absence of AEC. B. Categories of the SNPs found in Fig. 6A, with the categories from genes that are directly linked to the lysine pathway highlighted in green C. Circos plot of the SNPs found after 48 hours adaptation relative to the the lysine metabolism genes. The bar plots of each SNP represents the frequency across sequenced strains (n=9). Lysine metabolism genes are highlighted as black bars (50X zoom). The inner green plot represents the average sequencing coverage for each position across all sequenced genomes. D. Map of enriched mutations found for the same selective pressure (1,000 µM AEC) using our deep scanning mutagenesis library. The bar plots represent log2 enrichment scores
Figure 2 A Representative microCT images of trabecular bone from the femoral metaphysis of male Prx1-Cre;Usp34f/f and littermate control mice. Scale bar, 500 μm. B Von Kossa staining of undecalcified sections of femurs. Scale bar, 500 μm  C Histomorphometric analysis of trabecular bone from the femoral metaphysis. Results are shown as mean ± SEM; n=8; a: p<0.05, b: p<0.01 and c: p<0.001 by t test  D Serum levels of P1NP and CTX. Results are shown as mean ± SEM; n=8; *: p<0.05 by t test  E Quantitative RT-PCR analyses of the gene expression of femoral bone samples. Results are shown as mean ± SEM; n=6; *: p<0.05, **: p<0.01 and ***: p<0.001 by t test  F Representative images of ALP, ARS and Van Gieson's staining of MSCs obtained from Prx1-Cre;Usp34f/f and littermate control mice  G, H Quantitative analyses of the ALP activity and calcium mineralization of MSCs. Results are shown as mean ± SEM; n=6; *: p<0.05 by t test  I Quantitative RT-PCR analyses of the transcription of osteogenic markers. MSCs were differentiated in osteogenic medium for 5 days. Results are shown as mean ± SEM; n=3; *: p<0.05, **: p<0.01 and ***: p<0.001 by t test  J Representative images of ALP, ARS and Van Gieson's staining of primary calvarial osteoblasts  K, L Quantitative analyses of the ALP activity and calcium mineralization of primary calvarial osteoblasts. Results are shown as mean ± SEM; n=6; *: p<0.05 and **: p<0.01 by t test  M Quantitative RT-PCR analyses of the transcription of osteogenic markers. Calvarial osteoblasts were differentiated in osteogenic medium for 5 days. Results are shown as mean ± SEM; n=3; *: p<0.05, and **: p<0.01 by t test
Ago1x does not inhibit translation of target mRNAs (A) Schematic showing the principle of N-peptide and BoxB element-based assay to study translation repression ability of Ago proteins.Figure 7Schematic showing the principle of N-peptide and BoxB element-based assay to study translation repression ability of Ago proteins. N-peptide-tagged Ago1 and Ago2 proteins bind the BoxB elements in the 3′UTR of renilla luciferase (RLuc) and bring down their translation. Activity of RLuc relative to co-transfected FLuc in HeLa cells expressing N-HA-tagged or untagged Ago proteins as indicated. Bar graph (mean ± S.E.) is a representative of four independent experiments done in triplicates. Expression of HA-tagged Ago proteins was confirmed by Western blot (bottom). *, P = 0.018; #, P = 0.041; $, P = 0.013. Numbers on the bars represent % change in relative luciferase activity. Expression of RLuc protein in HeLa cells expressing N-HA-tagged or untagged Ago proteins as indicated. Densitometric analysis of three independent experiments is shown as bar graph (mean ± S.E.). RT-PCR analysis for RLuc mRNA level is shown (bottom). **, P = 0.001. Activity of RLuc relative to co-transfected FLuc in HeLa cells expressing N-HA-tagged Ago2 (N-HA-Ago2) or N-HA-tagged chimeric Ago2 appended with the ISR region of Ago1x (N-HA-Ago2-ISRAgo1) as indicated. Bar graph (mean ± S.E.) is a representative of three independent experiments done in triplicates. Expression of HA-tagged Ago proteins was confirmed by Western blot (bottom). ***, P < 0.0001; **, P = 0.0004.