Journal: International Journal of Biological Sciences

Article Title: Downregulation of m 6 A Reader YTHDC2 Promotes the Proliferation and Migration of Malignant Lung Cells via CYLD/NF-κB Pathway

doi: 10.7150/ijbs.58514

Figure Lengend Snippet: YTHDC2 gene and protein expression is downregulated in patients with lung cancer from The Cancer Gene Atlas, Gene Expression Omnibus and Human Protein Atlas databases, as well as in CS-exposed cells. (A) Differential analysis of YTHDC2 mRNA expression in lung cancer tissues based on the Gene Expression Profiling Interactive Analysis tool. * P<0.05 vs. normal tissues. Differential analysis of YTHDC2 mRNA expression in lung cancer tissues from (B) GSE32665 and (C) GSE19188 datasets. (D) Representative IHC images showed that YTHDC2 staining was found in the cell cytoplasm in lung cancer and normal lung tissues. High expression of YTHDC2 could be found in adjacent normal tissues, while its expression was decreased in the majority of lung cancer tissues. (E) Differential analysis of YTHDC2 staining positive ratio quantitated by IHC Profiler in lung cancer tissue arrays. YTHDC2 staining positive ratio in lung cancer tissues with different (F) maximum diameter, (G) pathological stage and (H) invasion depth. (I) Relative mRNA expression level of YTHDC2 in CS-exposed cells (S10, S20 and S30) and normal BEAS-2B cells. Western blot analysis (J) and quantitative results (K) of YTHDC2 protein expression in CS-exposed cells (S10, S20 and S30) and normal BEAS-2B cells. S10, S20 and S30 represent BEAS-2B cells exposed to CS for 10, 20 and 30 passages, respectively. **P<0.01 vs. normal BEAS-2B cells. IHC, immunohistochemistry; YTHDC2, YTH domain containing 2; CS, cigarette smoke.

Article Snippet: TaqMan copy number assays for YTHDC2 (probe ID: Hs06075146_cn) were performed according to the manufacturer's instructions.

Techniques: Expressing, Gene Expression, Staining, Western Blot, Immunohistochemistry

Journal: International Journal of Biological Sciences

Article Title: Downregulation of m 6 A Reader YTHDC2 Promotes the Proliferation and Migration of Malignant Lung Cells via CYLD/NF-κB Pathway

doi: 10.7150/ijbs.58514

Figure Lengend Snippet: Proteomics analysis of YTHDC2-knockdown cells. Bubble chart showing the KEGG enrichment results of the genes associated with (A) LUAD and (B) LUSC. The larger the rich factor, the greater the degree of enrichment. The color gradient from red to green represents the P-value; the closer to green color, the lower the P-value and the higher the significance level corresponding to the enrichment. Volcano plots showing the tumor suppressor genes in YTHDC2-related genes in (C) LUAD and (D) LUSC. (E) Bar graph showing the number of upregulated and downregulated proteins in YTHDC2-knockdown cells. (F) Subcellular distribution of DEGs. (G) Biological process and (H) KEGG pathway analysis of DEGs. (I) The protein-protein interaction network of DEGs was constructed using Cytoscape. (J-L) Identification of the top 3 significant clusters using the plug-in Minimal Common Oncology Data Elements in Cytoscape, and biological process enrichment analysis of the proteins in these clusters. KEGG, Kyoto Encyclopedia of Genes and Genomes; YTHDC2, YTH domain containing 2; LUSC, lung squamous cell carcinoma; LUAD , lung adenocarcinoma; DEGs, differentially expressed genes.

Article Snippet: TaqMan copy number assays for YTHDC2 (probe ID: Hs06075146_cn) were performed according to the manufacturer's instructions.

Techniques: Knockdown, Construct

Journal: International Journal of Biological Sciences

Article Title: Downregulation of m 6 A Reader YTHDC2 Promotes the Proliferation and Migration of Malignant Lung Cells via CYLD/NF-κB Pathway

doi: 10.7150/ijbs.58514

Figure Lengend Snippet: YTHDC2 downregulation promotes lung cancer cell proliferation. Representative images and quantification results of cell cycle of (A) BEAS-2B and (C) H1299 cells transfected with siYTHDC2 and NC. Representative images and quantification results of cell cycle of (B) S30 and (D) H1299 cells transfected with overexpression vector (pYTHDC2) and blank vector. (E) Cell proliferation was measured in BEAS-2B and H1299 cells transfected with siYTHDC2 and NC by EdU cell proliferation assay. (F) Cell proliferation were evaluated in S30 and H1299 cells transfected with pYTHDC2 and blank control by EdU cell proliferation assay. (G) Representative immunofluorescence images and (I) quantification results of Ki67 in BEAS-2B and H1299 cells transfected with siYTHDC2 and NC. (H) Representative immunofluorescence images and (J) quantification results of Ki67 in S30 and H1299 cells transfected with pYTHDC2 and blank control. Relative mRNA expression level of cyclin D1 in (K) BEAS-2B and H1299 cells transfected with siYTHDC2 and NC, as well as in (L) S30 and H1299 cells transfected with pYTHDC2 and blank control. * P<0.05 vs. NC or blank control group, ** P<0.01 vs. NC or blank control group. YTHDC2, YTH domain containing 2; siRNA, small interfering RNA; NC, negative control; EdU, 5-ethynyl-2'-deoxyuridine.

Article Snippet: TaqMan copy number assays for YTHDC2 (probe ID: Hs06075146_cn) were performed according to the manufacturer's instructions.

Techniques: Transfection, Over Expression, Plasmid Preparation, Proliferation Assay, Control, Immunofluorescence, Expressing, Small Interfering RNA, Negative Control

Journal: International Journal of Biological Sciences

Article Title: Downregulation of m 6 A Reader YTHDC2 Promotes the Proliferation and Migration of Malignant Lung Cells via CYLD/NF-κB Pathway

doi: 10.7150/ijbs.58514

Figure Lengend Snippet: YTHDC2 downregulation promotes lung cancer cell migration. (A) Representative images and quantification of the wound healing assay showing that cell migration was significantly increased at 24 and 48 h after transfection with siYTHDC2 in BEAS-2B and H1299 cells, as well as following transfection with (B) the overexpressing vector pYTHDC2 and a blank vector. Representative images and quantification of the Transwell migration assay of BEAS-2B and H1299 cells transfected with (C) siYTHDC2 and (D) pYTHDC2. (E) Quantitative PCR analysis of CDH1 and CDH2 in normal BEAS-2B andH1299 cells transfected with siYTHDC2, (F) as well as in S30 and H1299 cells transfected with pYTHDC2. (G) Western blot analysis and (I) quantitative results of EMT markers in BEAS-2B and H1299 cells transfected with siYTHDC2. (H) Western blot analysis and (J) quantitative results of EMT markers in S30 and H1299 cells transfected with pYTHDC2. *P<0.05 vs. NC (blank vector) group, **P<0.01 vs. NC (blank vector) group. YTHDC2, YTH domain containing 2; siRNA, small interfering RNA; EMT, epithelial-mesenchymal transition; NC: negative control; CDH1: E-cadherin; CDH2: N-cadherin.

Article Snippet: TaqMan copy number assays for YTHDC2 (probe ID: Hs06075146_cn) were performed according to the manufacturer's instructions.

Techniques: Migration, Wound Healing Assay, Transfection, Plasmid Preparation, Transwell Migration Assay, Real-time Polymerase Chain Reaction, Western Blot, Small Interfering RNA, Negative Control

Journal: International Journal of Biological Sciences

Article Title: Downregulation of m 6 A Reader YTHDC2 Promotes the Proliferation and Migration of Malignant Lung Cells via CYLD/NF-κB Pathway

doi: 10.7150/ijbs.58514

Figure Lengend Snippet: YTHDC2 overexpression suppresses H1299 cells growth in vivo . (A) Images of the xenograft tumors formed in nude mice injected with YTHDC2-overexpressing and control cells. (B) Volume and (C) weight of xenograft tumors isolated from nude mice. (D) Representative images and (E) quantification of the results of cell cycle analysis of single cell suspensions yielded from xenograft tumors. (F) Representative images of hematoxylin and eosin staining, and immunohistochemical staining of YTHDC2, Ki-67, cyclin D1, E-cadherin and N-cadherin in xenograft tumors derived from nude mice. Scale bar, 50 µm; **P<0.01 vs. blank group. YTHDC2, YTH domain containing 2.

Article Snippet: TaqMan copy number assays for YTHDC2 (probe ID: Hs06075146_cn) were performed according to the manufacturer's instructions.

Techniques: Over Expression, In Vivo, Injection, Control, Isolation, Cell Cycle Assay, Staining, Immunohistochemical staining, Derivative Assay

Journal: International Journal of Biological Sciences

Article Title: Downregulation of m 6 A Reader YTHDC2 Promotes the Proliferation and Migration of Malignant Lung Cells via CYLD/NF-κB Pathway

doi: 10.7150/ijbs.58514

Figure Lengend Snippet: YTHDC2 mRNA expression was regulated by gene amplification. Distribution of patients with (A) LUAD and (D) LUSC with different YTHDC2 amplification status. YTHDC2 mRNA expression in (B) LUAD and (E) LUSC tissues with different YTHDC2 amplification status. Different letters (a, b, c and d) represent statistically significant group differences. Pearson's correlation analysis revealed a significant positive correlation between YTHDC2 mRNA expression and copy numbers in (C) LUAD and (F) LUSC. The line represents linear regression of data (LUAD: y=1.065x+9.177, R 2 =0.385; LUSC: y=0.965x+9.318, R 2 =0.198). (G) The Oncomine datasets for the corresponding YTHDC2 copy numbers in lung cancer were obtained with a threshold P=0.001 and ≥2 fold-change. The data in the graphic show significant downregulation (blue column) of YTHDC2 copy numbers in lung cancer versus normal tissue. The intensity of the blue color represents the respective levels of YTHDC2 copy number. (H) Copy number variation in LUAD samples with different smoking histories. Different letters (a, b, c and d) represent statistically significant group differences. (I) Copy number variation of YTHDC2 in BEAS-2B cells and cigarette smoke-exposed cells (grey block), as well as in two lung cancer cell lines (black block). The dotted line (copy number = 2) represents the copy number of the reference gene RNase P. * P<0.05, ** P<0.01 vs. BEAS-2B cells. YTHDC2, YTH domain containing 2; LUSC, lung squamous cell carcinoma; LUAD , lung adenocarcinoma.

Article Snippet: TaqMan copy number assays for YTHDC2 (probe ID: Hs06075146_cn) were performed according to the manufacturer's instructions.

Techniques: Expressing, Amplification, Blocking Assay

Journal: International Journal of Biological Sciences

Article Title: Downregulation of m 6 A Reader YTHDC2 Promotes the Proliferation and Migration of Malignant Lung Cells via CYLD/NF-κB Pathway

doi: 10.7150/ijbs.58514

Figure Lengend Snippet: YTHDC2 promotes CYLD mRNA stability and inhibits NF-κB activity. (A) Scatter plot showing the mRNA transcripts identified by YTHDC2 RIP-sequencing, and transcripts that were significantly enriched are marked in red. (B) Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis of the mRNAs significantly enriched by YTHDC2 RIP. (C) Venn diagram showing the intersection of the YTHDC2 RIP-enriched mRNAs, YTHDC2-related mRNAs, upregulated differentially expressed proteins and tumor suppressor genes. (D) Relative mRNA expression level of CYLD in YTHDC2-overexpressing and knocked down H1299 cells. ** P<0.01 vs. blank group, ## P<0.01 vs. negative control group. (E) Relative protein expression level of CYLD in YTHDC2-overexpressing and knocked down H1299 cells. (F) RNA decay assay for CYLD mRNA stability upon YTHDC2 overexpression and knockdown in H1299 cells. (G) Immunostaining analysis of NF-κB p65 and CYLD in YTHDC2-overexpressing and -knockdown H1299 cells. (H) The DNA-binding activity of NF-κB in YTHDC2-overexpressing and -knockdown H1299 cells was measured by electrophoretic mobility shift assay using the biotin-labeled consensus NF-κB-binding sequence. (I) Immunohistochemistry staining of NF-κB p65 and CYLD in xenograft tumors derived from nude mice. RIP, RNA immunoprecipitation; YTHDC2, YTH domain containing 2; CYLD, cylindromatosis.

Article Snippet: TaqMan copy number assays for YTHDC2 (probe ID: Hs06075146_cn) were performed according to the manufacturer's instructions.

Techniques: Activity Assay, Sequencing, Expressing, Negative Control, Over Expression, Knockdown, Immunostaining, Binding Assay, Electrophoretic Mobility Shift Assay, Labeling, Immunohistochemistry, Staining, Derivative Assay, RNA Immunoprecipitation

Journal: International Journal of Biological Sciences

Article Title: Downregulation of m 6 A Reader YTHDC2 Promotes the Proliferation and Migration of Malignant Lung Cells via CYLD/NF-κB Pathway

doi: 10.7150/ijbs.58514

Figure Lengend Snippet: YTHDC2 regulates the stability of CYLD through m 6 A modification. (A) Prediction score of m 6 A distribution in CYLD sequence according to the sequence-based RNA adenosine methylation site predictor online tool. (B) meRIP-quantitative PCR and (C) meRIP-PCR results showed amplification of sites 2 to 5, indicating m 6 A modification in these four segments. Anti-IgG antibody was used as control. (D and E) Western blot results showing the relative protein expression level of CYLD in METTL3 , METTL14 , FTO and ALKBH5 knocked down H1299 cells. (F and G) Western blot result showing the relative protein expression level of CYLD in H1299 cells treated with DAA, a global methylation inhibitor, and with meclofenamic acid, a FTO inhibitor. (H and I) Western blot results showing the relative protein expression level of CYLD in YTHDC2-overexpressing H1299 cells treated with or without 3-DAA. YTHDC2, YTH domain containing 2; CYLD, cylindromatosis; meRIP, m 6 A methylated RNA immunoprecipitation; m 6 A, N6-methyladenosine; METTL, methyltransferase-like; FTO, fat mass and obesity-associated protein; DAA, 3-deazaadenosine.

Article Snippet: TaqMan copy number assays for YTHDC2 (probe ID: Hs06075146_cn) were performed according to the manufacturer's instructions.

Techniques: Modification, Sequencing, Methylation, Real-time Polymerase Chain Reaction, Amplification, Control, Western Blot, Expressing, RNA Immunoprecipitation

Journal: Cell Death Discovery

Article Title: RNA m 6 A involves in regulation of oxidative stress and apoptosis may via NF-kB pathway in cadmium-induced lung cells

doi: 10.1038/s41420-024-02284-w

Figure Lengend Snippet: The blue fluorescence represents the nucleus, the pink fluorescence represents the FTO protein, the red fluorescence represents the NF-κB p65 protein, the green fluorescence represents the YTHDC2 protein, and the intensity of fluorescence represents the level of protein expression. The overlaps of others and blue fluorescence represent the localization of the protein in the nucleus, while red and green fluorescence overlap represent the co-localization of YTHDC2 and NF-κB p65 protein in the cytoplasm (white arrow). A Immunofluorescence map of different protein expression in lung tissue (10×). B Protein immunofluorescence quantitative map. C Nuclear and cytoplasmic distribution of Nf-κB p65 protein (400×). D Nuclear and cytoplasmic distribution of FTO protein (400×). E Nucleoplasmic distribution of YTHDC2 protein (400×). F Difference plots of co-localization of FTO and YTHDC2 with NF-κB in different dose groups (300×).

Article Snippet: YTHDC2 Polyclonal antibody , Proteintech Group, Inc , 27779-1-AP , 1:2000 , Rabbit , 160.

Techniques: Fluorescence, Expressing, Immunofluorescence

Journal: Cell Death Discovery

Article Title: RNA m 6 A involves in regulation of oxidative stress and apoptosis may via NF-kB pathway in cadmium-induced lung cells

doi: 10.1038/s41420-024-02284-w

Figure Lengend Snippet: Antigen repair condition.

Article Snippet: YTHDC2 Polyclonal antibody , Proteintech Group, Inc , 27779-1-AP , 1:2000 , Rabbit , 160.

Techniques: Labeling

Journal: Cell Death Discovery

Article Title: RNA m 6 A involves in regulation of oxidative stress and apoptosis may via NF-kB pathway in cadmium-induced lung cells

doi: 10.1038/s41420-024-02284-w

Figure Lengend Snippet: Primer sequences and lengths of target genes.

Article Snippet: YTHDC2 Polyclonal antibody , Proteintech Group, Inc , 27779-1-AP , 1:2000 , Rabbit , 160.

Techniques:

Journal: Cell Death Discovery

Article Title: RNA m 6 A involves in regulation of oxidative stress and apoptosis may via NF-kB pathway in cadmium-induced lung cells

doi: 10.1038/s41420-024-02284-w

Figure Lengend Snippet: Target protein manufacturers and dilutions.

Article Snippet: YTHDC2 Polyclonal antibody , Proteintech Group, Inc , 27779-1-AP , 1:2000 , Rabbit , 160.

Techniques: Recombinant

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) Illustration of functional domains in the mouse YTHDC2 protein. (B) Schematic diagram of mouse Ythdc2 gene structure, targeting vector, and targeted alleles. Ythdc2 consists of 30 exons. Exons 13-16 are flanked by loxP sites. Deletion of exons 13-16 results in a deletion of 149 amino acids (aa 578 – 726) and a frameshift in the mutant transcript. Coding exons are shown as black bars. 5’ and 3’ untranslated regions are shown as gray boxes. HyTK is a double selection marker and enables hygromycin positive selection and thymidine kinase-negative selection in ES cells. (C) Histology (H&E staining) of 3-month-old wild-type and Ythdc2 -/- testes. Arrowheads indicate abnormal metaphase-like spermatocytes. (D) Reduced testis size in 6-week-old Ythdc2 fl/- Stra8 -Cre males. (E) Histology (H&E staining) of 6-week-old control and Ythdc2 fl/- Stra8 -Cre testes. Arrowheads indicate abnormal metaphase-like spermatocytes. (F) Reduced testis size in 8-week-old Ythdc2 fl/em1 Ngn3 -Cre mice. (G) Periodic acid Schiff-stained sections of Bouin’s-fixed testes from 5-month-old Ythdc2 fl/em1 Ngn3 -Cre and Ythdc2 +/em1 mice. Ythdc2 em1 is a deletion allele . In the higher magnification image of the boxed region, arrowheads annotate abnormal metaphase-like cells. Scale bars (C, E, and G left panels), 50 μm; (G right panel), 20μm.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques: Functional Assay, Plasmid Preparation, Mutagenesis, Selection, Marker, Staining

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) Regimen of tamoxifen treatment in ≥8-week-old Ythdc2 fl/- Ddx4 -Cre ERT2 male mice. See also . (B) Testis weight of control ( Ythdc2 fl/- ) and Ythdc2 iKO males (mean ± s.d.; n ≥ 3). (C) Western blot analysis of cell cycle regulators in control ( Ythdc2 fl/- ), Ythdc2 iKO , and Ythdc2 -/- testes. (D) Histology of control and Ythdc2 iKO (2 dpt) testes. Five stages of seminiferous tubules are shown (VIII-XII). Abbreviations: PreL, preleptotene; Lep, leptotene; Zyg, zygotene; Pa, pachytene; Dip, diplotene; Met, metaphase; RS, round spermatid; ES, elongating spermatid; CS, condensing spermatids. Arrowheads indicate apparently apoptotic pachytene spermatocytes. Loss of diplotene spermatocytes at the stage XI Ythdc2 iKO tubule is demarcated by a dashed blue line. Scale bar, 50 µm. See also . (E) Surface nuclear spread analysis of control and Ythdc2 iKO (2 dpt) spermatocytes. Absence of TC pachytene cells in control is marked with a large symbol. A severe loss of diplotene cells in Ythdc2 iKO (2 dpt) is indicated by a small symbol. TC, telomere clustering. The percentage of each type of spermatocytes in control and Ythdc2 iKO testes (mean ± s.d.) is shown in plot (F). >80 cells per mouse and three males per genotype per timepoint were analyzed. Scale bar, 10 µm.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques: Western Blot

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) Histology of stage VIII, IX, X, XI, and XII tubules are shown. For 6, 8, and 10 dpt, stages XI and XII could not be distinguished from each other, because of a total loss of diplotene and metaphase spermatocytes. Note a partial loss of zygotene spermatocytes in stages XI/XII at 6 dpt and a complete loss of zygotene spermatocytes in stages XI/XII at 8 and 10 dpt. Arrowheads indicate the apoptotic spermatocytes. Abbreviations: PreL, preleptotene; Lep, leptotene; Zyg, zygotene; Pa, pachytene; Dip, diplotene; Met, metaphase I or II spermatocyte; RS, round spermatid; ES, elongating spermatid; CS, condensing spermatid. Scale bar, 50 μm. (B) Time-dependent progressive loss of spermatocytes in adult tamoxifen-treated Ythdc2 iKO testes. Lines indicate the presence of spermatocytes. Dash lines indicate reduction in the indicated type of spermatocytes. A thinner dash line at 2 dpt indicates a severe reduction in the number of diplotene spermatocytes.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques:

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) TUNEL analysis of paraffin-embedded tissue sections from control and Ythdc2 iKO (2 dpt) testes. The acrosome morphology shown by SP10 immunofluorescence staining was used for seminiferous tubule staging. Nuclear DNA was stained with DAPI. Arrowheads indicate TUNEL-positive pachytene cells. The dashed line in stage IX Ythdc2 iKO demarcates pachytene cells (the inner layer) from leptotene cells (the outer layer). Abbreviations: PreL, preleptotene; Lep, leptotene; Pa, pachytene; RS, round spermatid; ES, elongating spermatid. Scale bar, 50 μm. (B) Percentage of TUNEL-positive tubules from control and Ythdc2 iKO testes at 2 dpt. The mean ± s.d. values were plotted. (C) Quantification of TUNEL-positive cells in TUNEL-positive tubules (mean ± s.d.) from control and Ythdc2 iKO at 2 dpt. (B, C) Two males per genotype (control and Ythdc2 iKO ) were analyzed. At least 200 tubules were counted for each mouse.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques: TUNEL Assay, Immunofluorescence, Staining

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) Immunofluorescence analysis of pH3 (green) in seminiferous tubules from adult control ( Ythdc2 fl/- ) and Ythdc2 iKO testes at 2, 4, and 6 dpt. The acrosome (red) was stained with anti-SP10 antibody. Nuclear DNA was counterstained with DAPI (blue). Abbreviations: PreL, preleptotene; Lep, leptotene; Zyg, zygotene; Pa, pachytene; RS, round spermatid; ES, elongating spermatid; CS, condensed spermatid; Met, metaphase I or II spermatocytes; Met-like, metaphase-like spermatocyte. Inset shows an enlarged view of the boxed pH3-positive metaphase-like spermatocyte. Scale bar, 50 μm. (B) Percentage of pH3-positive tubules from adult control, 2 dpt Ythdc2 iKO , and 4 dpt Ythdc2 iKO testes. The mean ± s.d. values were plotted. (C) The number of pH3-positive cells in pH3-positive tubules (mean ± s.d.) from adult control, 2 dpt Ythdc2 iKO , and 4 dpt Ythdc2 iKO testes. (B, C) Two males per genotype were analyzed. At least 200 tubules were counted for each mouse. The stages of seminiferous tubules are shown in Roman numerals.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques: Immunofluorescence, Staining

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) Assessment of purity of Sta-put isolated control ( Ythdc2 fl/- ) and Ythdc2 iKO pachytene spermatocytes by bright view and spread analysis. (B) qRT-PCR analysis of a set of 8 upregulated genes and 8 downregulated genes in Ythdc2 iKO pachytene cells. (C) Correlation of expression changes of 8 upregulated and 8 downregulated genes between RNA-seq and qRT-PCR. (D) Western blot analysis of selected proteins in purified pachytene cells. Western blotting was performed on independent samples twice. ACTB serves as a loading control. Quantification of each protein is normalized to ACTB and set at 1.0 in control pachytene cells.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques: Isolation, Quantitative RT-PCR, Expressing, RNA Sequencing Assay, Western Blot, Purification

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) Volcano plot of expression changes in Ythdc2 iKO vs control ( Ythdc2 fl/- ; HT) pachytene spermatocytes. Only coding genes (13,754) with an average expression of ≥50 across the samples are plotted. (B) MA plot of expression changes. The differentially expressed (up and down regulated) genes are shown as red dots. See also Figures S4 and S5. (C) Expression of differentially expressed genes in various wild-type spermatogenic populations. SSC, spermatogonial stem cells; SC, spermatocytes; RS, round spermatids. (D) Expression comparison of genes in Ythdc2 iKO vs control ( Ythdc2 fl/- ; HT) pachytene cells that are enriched or depleted of m 6 A in juvenile testes . (E) Overlap of YTHDC2 RIP-seq targets with the differentially expressed genes in Ythdc2 iKO pachytene cells. (F) Overlap of YTHDC2 RIP-seq targets with the differentially expressed genes in Ythdc2 iKO pachytene cells.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques: Expressing

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) Comparison of Ythdc2 expression in individual samples. (B) Gene ontology analysis of the up-regulated genes in Ythdc2 iKO pachytene cells. (C) Gene ontology analysis of the down-regulated genes in Ythdc2 iKO pachytene cells. (D) Expression changes based on individual chromosomes. Mitochondria are included. (E) The number of upregulated vs downregulated genes on each chromosome and mitochondria.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques: Expressing

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) Immunofluorescence of α-tubulin, SUN1, SYCP2 in intact control ( Ythdc2 fl/- ) and Ythdc2 iKO (2 dpt) pachytene spermatocytes. Scale bar, 10 μm. See also . (B) Nuclear spread analysis of pachytene, diplotene, and diakinesis/metaphase I (dia/met) spermatocytes after treatment with DMSO or nocodazole. TC, telomere clustering. Scale bar, 10 μm. (C) Percentage of spermatocytes (mean ± s.d.) in control ( Ythdc2 fl/- ) and Ythdc2 iKO (2 dpt) after treatment with DMSO or nocodazole. Only pachytene, diplotene, and diakinesis/metaphase I spermatocytes are included. At least 500 cells were counted per genotype per treatment group. Two males per genotype per experiment were used. The experiments were performed three times (n=3). See also .

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques: Immunofluorescence

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: Immunofluorescence of α-tubulin, SUN1, SYCP2 was performed in intact control ( Ythdc2 fl/- ) and Ythdc2 iKO (2 dpt) pachytene spermatocytes. Scale bar, 10 μm.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques: Immunofluorescence

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) Nuclear spread analysis of pachytene, diplotene, and diakinesis/metaphase I (dia/met) spermatocytes from control ( Ythdc2 fl/- ) and Ythdc2 iKO (2 dpt) testes after treatment with DMSO, cytochalasin D, or latrunculin A. TC, telomere clustering. Scale bar, 10 μm. (B) Percentage of spermatocytes (mean ± s.d.). Only pachytene, diplotene, and diakinesis/metaphase I spermatocytes are included. At least 350 cells were counted per genotype per treatment group. Two males per genotype per experiment were used. The experiments were performed twice.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques:

Journal: bioRxiv

Article Title: YTHDC2 Is Essential for Pachytene Progression and Prevents Aberrant Microtubule-Driven Telomere Clustering in Male Meiosis

doi: 10.1101/2021.04.12.439470

Figure Lengend Snippet: (A) Requirement of YTHDC2 at the leptotene and pachytene stages. The red cross designates the stage of meiotic arrest in the Ythdc2 or Meioic mutants. Ythdc2 or Meioc global knockout mutants display the same early meiotic arrest, which is also present in the Ythdc2 iKO ( Ythdc2 fl/- Ddx4 -Cre ERT2 ) mutant. In contrast, Ythdc2 iKO pachytene cells undergo apoptosis at the late pachytene stage. The seminiferous tubule stage is shown in Roman numerals. See also . (B) Illustration of telomere distribution in wild type and Ythdc2 iKO pachytene spermatocytes. Two pairs of homologous chromosomes are depicted. The SUN/KASH proteins that connect telomeres with microtubules are not shown. In contrast with the random distribution of telomeres in wild type, telomeres in the Ythdc2 iKO pachytene cell are clustered to one pole, where microtubules converge and the cytoplasm is expanded.

Article Snippet: The cDNA fragment encoding the N-terminal 239 aa of mouse YTHDC2 was amplified by PCR from the Ythdc2 plasmid (NM_001163013) (Cat. No. MR226474, OriGene Technologies) using these two primers: 5’-CGCGGATCCATGTCCAGGCCGAGCAG-3’ and 5’-CCCAAGCTTTGTGGTCTTTCCAGACCCAGT-3’, and was cloned into the Bam HI and Hind III sites of the pQE30 vector.

Techniques: Knock-Out, Mutagenesis

Journal: PLoS ONE

Article Title: Identification and Validation of a Multigene Predictor of Recurrence in Primary Laryngeal Cancer

doi: 10.1371/journal.pone.0070429

Figure Lengend Snippet: mRNA target selection from the 30 probes included in the prognostically relevant signature and TaqMan® gene expression assay description.

Article Snippet: 10 , 205836_s_at , YTH domain containing 2 , NM_022828 , YTHDC2 , 64848 , , 0,0981884 , 1,04 , yes , , NM_022828.3 , Hs00967276_g1 , 75 , 29–30 , Accepted.

Techniques: Selection, Gene Expression, Membrane, Binding Assay, Sequencing

Journal: PLoS ONE

Article Title: Identification and Validation of a Multigene Predictor of Recurrence in Primary Laryngeal Cancer

doi: 10.1371/journal.pone.0070429

Figure Lengend Snippet: 30-probe set model annotations.

Article Snippet: 10 , 205836_s_at , YTH domain containing 2 , NM_022828 , YTHDC2 , 64848 , , 0,0981884 , 1,04 , yes , , NM_022828.3 , Hs00967276_g1 , 75 , 29–30 , Accepted.

Techniques: Binding Assay, Membrane, Sequencing

Journal: PLoS ONE

Article Title: Identification and Validation of a Multigene Predictor of Recurrence in Primary Laryngeal Cancer

doi: 10.1371/journal.pone.0070429

Figure Lengend Snippet: Individual and profiled gene expression effects on DFS.

Article Snippet: 10 , 205836_s_at , YTH domain containing 2 , NM_022828 , YTHDC2 , 64848 , , 0,0981884 , 1,04 , yes , , NM_022828.3 , Hs00967276_g1 , 75 , 29–30 , Accepted.

Techniques: Gene Expression

Journal: Frontiers in Oncology

Article Title: m 6 A Reader YTHDC2 Promotes Radiotherapy Resistance of Nasopharyngeal Carcinoma via Activating IGF1R/AKT/S6 Signaling Axis

doi: 10.3389/fonc.2020.01166

Figure Lengend Snippet: YTHDC2 is upregulated in radiation-resistance NPC cells. (A) Heat map showing the expression of m6A relevant genes in CNE2 and CEN2-IRR cells. The shades of red and blue represent increases and decreases in the expression of the corresponding genes, respectively. Data were generated in independent experiments (1 and 2). (B,C) mRNA (B) and protein (C) expression of the m6A relevant genes in CNE2, CNE2-IRR, HK1, and HK1-IRR cells. Data are presented as mean ± SD from n = 3. * P < 0.05, ** P < 0.01, *** P < 0.001, student's t test. (D) H-Score of YTHDC2 in NPC sample with complete response and with partial response after radiotherapy. Up-panel showed representative images of YTHDC2 protein level examined in NPC CR and PR tissues using immunohistochemistry (shown in 100×magnifications). *** P < 0.001, student's t test. (E) scatter plots comparing levels of YTHDC2 mRNA in different clinical stage groups of NPC samples from GSE102349. * P < 0.05, Student's t -test. (F) Recurrence-free survival rate analysis base on YTHDC2 mRNA expression in NPC patients from GSE102349.

Article Snippet: Moreover, Tanabe et al. found that knockdown of YTHDC2 inhibited the expression of several proteins involved in the metastasis of colon cancer ( ).

Techniques: Expressing, Generated, Immunohistochemistry

Journal: Frontiers in Oncology

Article Title: m 6 A Reader YTHDC2 Promotes Radiotherapy Resistance of Nasopharyngeal Carcinoma via Activating IGF1R/AKT/S6 Signaling Axis

doi: 10.3389/fonc.2020.01166

Figure Lengend Snippet: Clinical characteristics of 105 NPC patients according to YTHDC2 expression levels.

Article Snippet: Moreover, Tanabe et al. found that knockdown of YTHDC2 inhibited the expression of several proteins involved in the metastasis of colon cancer ( ).

Techniques: Expressing

Journal: Frontiers in Oncology

Article Title: m 6 A Reader YTHDC2 Promotes Radiotherapy Resistance of Nasopharyngeal Carcinoma via Activating IGF1R/AKT/S6 Signaling Axis

doi: 10.3389/fonc.2020.01166

Figure Lengend Snippet: Loss of function of YTHDC2 enhances radiosensitivity of NPC cell in vitro . (A) Demonstration of shRNA efficiency in down-regulation of YTHDC2 in CNE2-IRR and HK1-IRR cells. (B) CCK8 assays of CNE2-IRR and HK1-IRR cells with knockdown of YTHDC 2 with or without irradiation treatment. Data are presented as mean ± SD from n = 3. *** P < 0.001, student's t -test. (C) Images from soft agar colony formation assay in CNE2-IRR and HK1-IRR cells with YTHDC2 knockdown after expose to radiation were shown in 2 different magnifications (40×and 200×). Data are presented as mean ± SD from n = 4. (D) Quantitative analyses of colony size (50–100 μm and >100 μm) in CNE2-IRR and HK1-IRR cells with YTHDC2 knocking down after exposure to radiation. (E) Survival rate in CNE2-IRR cells expose to 0, 2, 4, 6, 8 Gy radiation after YTHDC2 was knocked down. Data are presented as mean ± SD from n = 3. ** P < 0.01, *** P < 0.001, student's t -test. (F) The apoptosis assay of CNE2-IRR with YTHDC2 knockdown after 0 or 6 Gy radiation by flow cytometric analysis (left) and the statistical analysis of the data (right). Data are presented as mean ± SD from n = 3, student's t -test.

Article Snippet: Moreover, Tanabe et al. found that knockdown of YTHDC2 inhibited the expression of several proteins involved in the metastasis of colon cancer ( ).

Techniques: In Vitro, shRNA, Knockdown, Irradiation, Soft Agar Assay, Apoptosis Assay

Journal: Frontiers in Oncology

Article Title: m 6 A Reader YTHDC2 Promotes Radiotherapy Resistance of Nasopharyngeal Carcinoma via Activating IGF1R/AKT/S6 Signaling Axis

doi: 10.3389/fonc.2020.01166

Figure Lengend Snippet: Overexpression of YTHDC2 attenuates radiosensitivity of NPC cell in vitro . (A) Demonstration of YTHDC2 over-expression in CNE2 and HK1 cells. (B) CCK8 assay indicated overexpression of YTHDC2 weaken radiotherapy effect in CNE2 cells. Data are presented as mean ± SD from n = 3. *** P < 0.001, student's t test. (C) Images of soft agar colony formation assay in CNE2 cells after expose to radiation were shown in 2 different magnifications (40×and 200×) (left) and statistical analysis of colony numbers (50–100 μm and >100 μm) (right). Data are presented as mean ± SD from n = 4. (D) The colonies (left) and survival rate (right) in CNE2 and HK1 cells expose to 0, 2, 4, 6, 8 Gy radiation with overexpression of YTHDC2. Data are presented as mean ± SD from n = 3. ** P < 0.01, student's t -test. (E) The apoptosis assay of CNE2 with overexpression of YTHDC2 after 0 or 6 Gy radiation exposure by flow cytometric analysis (left) and data analysis (right). Data are presented as mean ± SD from n = 3, student's t -test. * P < 0.05, student's t -test.

Article Snippet: Moreover, Tanabe et al. found that knockdown of YTHDC2 inhibited the expression of several proteins involved in the metastasis of colon cancer ( ).

Techniques: Over Expression, In Vitro, CCK-8 Assay, Soft Agar Assay, Apoptosis Assay

Journal: Frontiers in Oncology

Article Title: m 6 A Reader YTHDC2 Promotes Radiotherapy Resistance of Nasopharyngeal Carcinoma via Activating IGF1R/AKT/S6 Signaling Axis

doi: 10.3389/fonc.2020.01166

Figure Lengend Snippet: IGF1R is a key downstream gene of YTHDC2. (A) Fifty-six genes related to the PI3K-AKT/S6 signaling pathway were found to be upregulated in the GSE48501 dataset, among which IGF1R is the key downstream target gene of YTHDC2 by using m6A prediction database (m6avar.renlab.org) (B) PI3K-AKT signaling was activated in radiation-resistance NPC cells. (C) Western blot analyses of PI3K-AKT/S6 signaling activation in radiation resistant cells. (D) A positive correlation between the expressions of YTHDC2 and IGF1R in NPC tissues was found by analyzing GSE102349 dataset. (E) Knockdown of YTHDC2 in CNE2-IRR cells downregulated IGF1R protein expression and inactivated PI3K-AKT/S6 signaling. (F) Overexpression of YTHDC2 in CNE2 and HK1 cells upregulated IGF1R protein expression and activated PI3k-AKT signaling. All experiments were repeated twice.

Article Snippet: Moreover, Tanabe et al. found that knockdown of YTHDC2 inhibited the expression of several proteins involved in the metastasis of colon cancer ( ).

Techniques: Western Blot, Activation Assay, Knockdown, Expressing, Over Expression

Journal: Frontiers in Oncology

Article Title: m 6 A Reader YTHDC2 Promotes Radiotherapy Resistance of Nasopharyngeal Carcinoma via Activating IGF1R/AKT/S6 Signaling Axis

doi: 10.3389/fonc.2020.01166

Figure Lengend Snippet: Promoter methylation regulates gene expression level of YTHDC2. (A) Schematic representation of the CpG distribution in promoter region of the YTHDC2 gene from the CpG Island Searcher ( https://www.urogene.org/cgi-bin/methprimer/methprimer.cgi ). (B) Methylation-specific PCR (MSP) analysis of YTHDC2 promoter methylation status in two pairs of cell lines using 2 serials of primers. (C) YTHDC2 expression in CNE2-IRR and HK1-IRR cells treated with gemcitabine with different concentrations for 48 hr and subjected to immunoblotting and qPCR assay. Data are presented as mean ± SD from n = 3. * P < 0.05, ** P < 0.01, *** P < 0.001, student's t -test. (D) CNE2 and HK1 cells treated with 5-aza-2'-deoxycytidine with different concentrations for 48 hr and subjected to immunoblotting and qPCR assay of YTHDC2 expression. Data are presented as mean ± SD from n = 3. * P < 0.05, ** P < 0.01, *** P < 0.001, student's t test. (E) MSP analysis of YTHDC2 promoter methylation in paraffin-embeded sections of radiosensitivity and radioresistance NPC tissues. Data are presented as mean ± SD from n = 3. ** P < 0.01, *** P < 0.001, student's t -test. (F) CNE2-IRR cells were treated with radiotherapy, gemcitabine, radiotherapy plus gemcitabine, respectively. The same number of cells were seeded and the colonies formed were counted 14 days later.

Article Snippet: Moreover, Tanabe et al. found that knockdown of YTHDC2 inhibited the expression of several proteins involved in the metastasis of colon cancer ( ).

Techniques: Methylation, Gene Expression, Expressing, Western Blot

Journal: Frontiers in Oncology

Article Title: m 6 A Reader YTHDC2 Promotes Radiotherapy Resistance of Nasopharyngeal Carcinoma via Activating IGF1R/AKT/S6 Signaling Axis

doi: 10.3389/fonc.2020.01166

Figure Lengend Snippet: YTHDC2 regulates translation of IGF1R. (A) Verification of efficiency of FLAG-immunoprecipitation by western blot. (B) RNA immunoprecipitation of FLAG-YTHDC2 interacted with IGF1R in vivo in CNE2 cells. Protein–RNA complexes immunoprecipitated by anti-FLAG or IgG were determined by RT-PCR using specific primers for IGF1R or Hprt1 (negative control). (C) Cell lysate derived from shCON, shYTHDC2-1, and shYTHDC2-3 of CNE2-IRR cells were loaded on a linear gradient of 15–40% sucrose and centrifuged at 38,000 rpm at 4°C for 4 hr. After centrifugation, samples were then fractionated into 24 fractions (0.5 mL per fraction), respectively and were monitored at 260 nm. (D) The amount of IGF1R mRNA in 40S, 60S, 80S, polysome fractionated RNA and total RNA derived from shCON, shYTHDC2-1, and shYTHDC2-3 of CNE2-IRR cells were quantified by quantitative RT-PCR. Data are presented as mean ± SD from n = 3. ** P < 0.001; *** P < 0.001, student's t -test.

Article Snippet: Moreover, Tanabe et al. found that knockdown of YTHDC2 inhibited the expression of several proteins involved in the metastasis of colon cancer ( ).

Techniques: Immunoprecipitation, Western Blot, RNA Immunoprecipitation, In Vivo, Reverse Transcription Polymerase Chain Reaction, Negative Control, Derivative Assay, Centrifugation, Quantitative RT-PCR

Journal: Frontiers in Oncology

Article Title: m 6 A Reader YTHDC2 Promotes Radiotherapy Resistance of Nasopharyngeal Carcinoma via Activating IGF1R/AKT/S6 Signaling Axis

doi: 10.3389/fonc.2020.01166

Figure Lengend Snippet: Knockdown of YTHDC2 enhances radiotherapy effect in vivo . (A) Knockdown of YTHDC2 significantly enhanced radiotherapy effect ( n = 5). One dose of irradiation was given at the day 6 when tumor volumes reached around 100 mm 3 . Photos were taken from tumor samples collected at the end of experiment (day 21). (B) The growth curve of tumors from CEN2-IRR cells transduced with viral vectors expressing shRNA-1, shRNA-3, or control sequence ( n = 5). (C) Tumors weight measured at the end of experiment (day 21) ( n = 5). (D) The YTHDC2 expression and inhibition of PI3K-AKT/S6 pathway in tumors was determined by western blot. (E) Representative images of immunohistostaining for examining the expression of YTHDC2, IGF1R, p-AKT, and p-S6 in xenograft tissues. Images were shown in 2 different magnifications (100×and 200×). (F) Quantitative analysis of YTHDC2, IGF1R, p-AKT, and p-S6 expression in the xenograft tumors. All experiments were repeated twice. * P < 0.05, ** P < 0.01, *** P < 0.001, student's t -test.

Article Snippet: Moreover, Tanabe et al. found that knockdown of YTHDC2 inhibited the expression of several proteins involved in the metastasis of colon cancer ( ).

Techniques: Knockdown, In Vivo, Irradiation, Transduction, Expressing, shRNA, Control, Sequencing, Inhibition, Western Blot

Journal: Frontiers in Oncology

Article Title: m 6 A Reader YTHDC2 Promotes Radiotherapy Resistance of Nasopharyngeal Carcinoma via Activating IGF1R/AKT/S6 Signaling Axis

doi: 10.3389/fonc.2020.01166

Figure Lengend Snippet: A schematic diagram showing YTHDC2 upregulated expression of IGF1R attenuate radiotherapy response via mediated RNA translation.

Article Snippet: Moreover, Tanabe et al. found that knockdown of YTHDC2 inhibited the expression of several proteins involved in the metastasis of colon cancer ( ).

Techniques: Expressing