Journal: PLoS Pathogens

Article Title: Norovirus infection results in eIF2α independent host translation shut-off and remodels the G3BP1 interactome evading stress granule formation

doi: 10.1371/journal.ppat.1008250

Figure Lengend Snippet: G3BP1 interactome differs in MNV-infected cells and arsenite treated cells as addressed by immunofluorescence (A-C). BV2 GFP-G3BP1 cells were either infected with MNV (MOI 10) for 9h p.i. or treated with arsenite 0.1mM for 1h and subcellular colocalisation of candidate proteins identified by mass spectrometry as interacting with GFP-G3BP1 was addressed by immunostaining (magenta) in GFP-G3BP1 positive cells (cyan). MNV-infected cells were detected by immunostaining against dsRNA (gold), nuclei were stained with DAPI. Scale bars, 10μm, red arrows indicate example of colocalisation between GFP-G3BP1 and a candidate protein. Representative view (n = 3) of confocal analysis using staining against (A) YTHDF3, candidate for arsenite-induced SG only, (B) FMR1, candidate for both arsenite- and MNV-induced G3BP1 granules, (C) Rab7, candidate for MNV-induced G3BP1 granules only.

Article Snippet: Primary antibodies dilutions: rabbit anti-NS3 (1:600), mouse anti-dsRNA (1:1,000), mouse anti-puromycin (1:5, http://dshb.biology.uiowa.edu/PMY-2A4 ), mouse anti-G3BP1 (1:400, Invitrogen), goat anti-eIF3B (1:400, Santa Cruz), rabbit YTHDF3 (1:600, Proteintech), rabbit anti-RAB7A (1:600, Proteintech), rabbit anti-FMR1 (1:600, Novusbio).

Techniques: Infection, Immunofluorescence, Mass Spectrometry, Immunostaining, Staining

Journal: BMC Cancer

Article Title: N6-methyladenosine RNA modification (m6A) is of prognostic value in HPV-dependent vulvar squamous cell carcinoma

doi: 10.1186/s12885-022-10010-x

Figure Lengend Snippet: Summary of the analyzed m6A proteins as indicated and their correlation with overall survival (indicated as %alive) for the entire study cohort, HPV-independent, and HPV-dependent VSCC. The HPV-status was not available for 24 patients. Samples were grouped according to high and low expression based on the staining intensities. p -values for the group comparisons are based on log-rank tests (significance threshold p < 0.5). q -values are based on multiple hypotheses testing using the method of Benjamini and Hochberg with a significance threshold of q < 0.1

Article Snippet: Immunostaining of METTL3, METTL4, METTL14, WTAP, KIAA1429, FTO, ALKBH5, HNRNPA2B1, HNRNPC, YTHDC1, YTHDF1,YTHDF2, and YTHDF3 was performed on the TMAs using an automated staining system (BenchMark ULTRA; Ventana Medical Systems) which performed deparaffinization, pretreatment with cell conditioning buffer (CC1 buffer, pH8), and incubation with primary antibodies (FTO (1:50; Atlas Antibodies #HPA041086), ALKBH5 (1:200; Novus #NBP1-82,188), METTL3 (1:1000; Biorbyt #orb374082), METTL4 (1:40; Atlas Antibodies #HPA040061), METTL14 (1:100; Atlas Antibodies #HPA038002), WTAP (1:100; Atlas Antibodies #HPA010550), KIAA1429 (1:25; Atlas Antibodies #HPA031530), HNRNPC (1:25; Atlas Antibodies #HPA051075), HNRNPA2B1 (1:100; Atlas Antibodies #HPA001666), YTHDC1 (1:25; Atlas Antibodies #HPA036462), YTHDF1 (1:10; Biorbyt #orb179018), YTHDF2 (1:200; Biorbyt #orb39199), YTHDF3 (1:200; Biorbyt #orb374095) at 4 °C overnight.

Techniques: Expressing, Staining

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Multi-Omics Analysis Reveals Translational Landscapes and Regulations in Mouse and Human Oocyte Aging.

doi: 10.1002/advs.202301538

Figure Lengend Snippet: Figure 4. YTHDF3 deficiency inhibits RNA TE in mouse oocytes. a) The in vitro PB1 emission rates of mouse oocytes from the control groups and the m6A-related gene depletion groups. Each dot represents a single biological replicate. p-Values were calculated with Student’s t-test for paired samples. b) Immunofluorescence verifying the depletion of YTHDF3 by Trim-Away. Scale bar, 50 μm. The right panel shows the quantification of YTHDF3 protein levels. The average intensity of the control group oocytes was set as 1.0. Each dot represents a single oocyte analyzed. p-Value was calculated with two-tailed Mann–Whitney test. c) Immunofluorescence verifying the expression of YTHDF3 in young and aged mouse GV oocytes. Scale bar, 50 μm. The right panel shows the quantification of YTHDF3 protein levels. The average intensity of young mouse oocytes was set as 1.0. Each dot represents a single

Article Snippet: Antibodies used in this study are listed as follows: YTHDF3 (Novus Biologicals, 94636), HELLS (Proteintech, 11955-1-AP), Electroporation: Denuded mouse GV oocytes were collected as described above.

Techniques: In Vitro, Control, Two Tailed Test, MANN-WHITNEY, Expressing

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Multi-Omics Analysis Reveals Translational Landscapes and Regulations in Mouse and Human Oocyte Aging.

doi: 10.1002/advs.202301538

Figure Lengend Snippet: Figure 5. YTHDF3 modulates RNA translation efficiency in an m6A-dependent manner. a) Gene set enrichment analysis demonstrating that the TE of m6A-enriched RNA was significantly decreased upon YTHDF3 depletion. b) Bar plots showing the numbers of up- (FC>1.5) and down-regulated (FC<0.67) genes for m6A-enriched genes or genes not enriched by m6A, respectively. Pink denotes m6A-enriched genes. Blue denotes genes not enriched by m6A. c) Venn diagram portraying the overlap of YTHDF3 target genes among three independent RIP-seq biological replicates. d) Motif identified by HOMER within YTHDF3 RIP-seq peaks in HEK293T cells. e) Gene set enrichment analysis showing the TE alterations of YTHDF3-binding RNA upon

Article Snippet: Antibodies used in this study are listed as follows: YTHDF3 (Novus Biologicals, 94636), HELLS (Proteintech, 11955-1-AP), Electroporation: Denuded mouse GV oocytes were collected as described above.

Techniques: Binding Assay