Journal: Stem Cells International

Article Title: YTHDC1 Promotes Stemness Maintenance and Malignant Progression in Head and Neck Squamous Cell Carcinoma

doi: 10.1155/2022/7494354

Figure Lengend Snippet: YTHDC1 aberrantly expressed in HNSCC and was associated with cancer cell stemness. (a) The relative mRNA expression of YTHDC1 in tumor and normal tissues in HNSCC in TCGA database. The comparison of the YTHDC1 mRNA expression in (b) individual tumor grades, (c) HPV infection status, and (d) different nodal metastasis status. (e) The Pearson correlation between YTHDC1 and SOX2 mRNA expression in HNSCC according to TCGA data. (f) The Pearson correlation between YTHDC1 and BMI1 mRNA expression in HNSCC according to the TCGA data.

Article Snippet: The following antibodies were used in the experiment: GAPDH (Proteintech, 10494-1-AP), YTHDC1 (Cell Signaling Technology, 54737S), BMI1 (Abcam, ab269678), SOX2 (Abcam, ab97959), OCT4 (Abcam, ab18976), and NANOG (Abcam, ab109250).

Techniques: Expressing, Comparison, Infection

Journal: Stem Cells International

Article Title: YTHDC1 Promotes Stemness Maintenance and Malignant Progression in Head and Neck Squamous Cell Carcinoma

doi: 10.1155/2022/7494354

Figure Lengend Snippet: YTHDC1 was overexpressed in HNSCC patients. (a) Representative graph of YTHDC1 staining in tumor and normal tissues of HNSCC patients (left) and the IHC score quantification (right) ( n = 6). Scale bar, 100 μ m. (b) The YTHDC1 expression in patients HNSCC tissues and normal tissues detected by immunoblotting analysis ( n = 3). (c) The SOX2 and BMI1 protein expression in patient HNSCC tissues and normal tissues detected by immunoblotting analysis ( n = 3).

Article Snippet: The following antibodies were used in the experiment: GAPDH (Proteintech, 10494-1-AP), YTHDC1 (Cell Signaling Technology, 54737S), BMI1 (Abcam, ab269678), SOX2 (Abcam, ab97959), OCT4 (Abcam, ab18976), and NANOG (Abcam, ab109250).

Techniques: Staining, Expressing, Western Blot

Journal: Stem Cells International

Article Title: YTHDC1 Promotes Stemness Maintenance and Malignant Progression in Head and Neck Squamous Cell Carcinoma

doi: 10.1155/2022/7494354

Figure Lengend Snippet: Inhibition of the YTHDC1 expression significantly limits tumor cell stemness maintenance, migration, and proliferation. (a) Western blotting of YTHDC1 in six HNSCC cell lines. (b) Western blotting of YTHDC1 in SCC9 and HN4 with or without YTHDC1 knockdown. (c) qPCR data of YTHDC1 in SCC9 and HN4 with or without YTHDC1 knockdown. (d) SOX2 and BMI1 protein expression in SCC9 and HN4 with or without YTHDC1 knockdown. (e) The OCT4 and NANOG protein expression in SCC9 and HN4 with or without YTHDC1 knockdown. (f). SCC9 and HN4 cell proliferation ability was suppressed after the YTHDC1 expression was decreased. (g) Sphere formation ability of SCC9 and HN4 cell lines was inhibited after the YTHDC1 expression was suppressed. (h) Representative images and quantification of migration assay of SCC9 and HN4 cell lines with or without YTHDC1 knockdown. (i) Representative images and quantification of colony formation assay of SCC9 and HN4 cell lines with or without YTHDC1 knockdown. (j). Tumor xenograft experiment in nude mice detects the effect of YTHDC1 on tumor growth in vivo ( n = 6 in each group). (k) Tumor growth curve. The quantitative value of tumor volumes is plotted, and tumor volumes are measured once two days after injected. (l) Scatter diagram shows tumor weight.

Article Snippet: The following antibodies were used in the experiment: GAPDH (Proteintech, 10494-1-AP), YTHDC1 (Cell Signaling Technology, 54737S), BMI1 (Abcam, ab269678), SOX2 (Abcam, ab97959), OCT4 (Abcam, ab18976), and NANOG (Abcam, ab109250).

Techniques: Inhibition, Expressing, Migration, Western Blot, Knockdown, Colony Assay, In Vivo, Injection

Journal: Stem Cells International

Article Title: YTHDC1 Promotes Stemness Maintenance and Malignant Progression in Head and Neck Squamous Cell Carcinoma

doi: 10.1155/2022/7494354

Figure Lengend Snippet: Validation of the role of YTHDC1 in regulating stemness maintenance at the single cell level. (a) The UMAP plot of scRNA-seq cell data labeled by cell type. (b) The UMAP plot of the YTHDC1 expression in all the clusters. (c) The UMAP plot of cancer cells from HNSCC tissues shows the YTHDC1 expression pattern. (d) The Pearson correlation between YTHDC1 and SOX2 mRNA expression according to the single cell RNA data. (e) The Pearson correlation between YTHDC1 and BMI1 mRNA expression according to the single cell RNA data. (f) Volcano plot shows differentially expressed genes between YTHDC1 high and YTHDC1 low cells. (g) GO analysis of the upregulated genes in the YTHDC1 high expression cell population. (h) KEGG analysis of the upregulated genes in the YTHDC1 high expression cell population. (i) Violin plot depicts the expression of stem cell-related markers of YTHDC1 high and YTHDC1 low cells.

Article Snippet: The following antibodies were used in the experiment: GAPDH (Proteintech, 10494-1-AP), YTHDC1 (Cell Signaling Technology, 54737S), BMI1 (Abcam, ab269678), SOX2 (Abcam, ab97959), OCT4 (Abcam, ab18976), and NANOG (Abcam, ab109250).

Techniques: Biomarker Discovery, Labeling, Expressing

Journal: Oxidative Medicine and Cellular Longevity

Article Title: The Effect of N6-Methyladenosine Regulators and m6A Reader YTHDC1-Mediated N6-Methyladenosine Modification Is Involved in Oxidative Stress in Human Aortic Dissection

doi: 10.1155/2023/3918393

Figure Lengend Snippet: qRT-PCR primer sequences.

Article Snippet: After cell spreading, HUVECs were transfected with 75 nM YTHDC1 siRNAs or NC siRNAs for 48 h by riboFECT™ CP Transfection Kit (C10511-05, RiboBio, Guangzhou, China).

Techniques: Sequencing

Journal: Oxidative Medicine and Cellular Longevity

Article Title: The Effect of N6-Methyladenosine Regulators and m6A Reader YTHDC1-Mediated N6-Methyladenosine Modification Is Involved in Oxidative Stress in Human Aortic Dissection

doi: 10.1155/2023/3918393

Figure Lengend Snippet: Human aortic tissue validation, WB results of YTHDC1. (a) Western blot in AD and normal. (b) Significance test of difference: “ ∗ ” means P < 0.05.

Article Snippet: After cell spreading, HUVECs were transfected with 75 nM YTHDC1 siRNAs or NC siRNAs for 48 h by riboFECT™ CP Transfection Kit (C10511-05, RiboBio, Guangzhou, China).

Techniques: Western Blot

Journal: Oxidative Medicine and Cellular Longevity

Article Title: The Effect of N6-Methyladenosine Regulators and m6A Reader YTHDC1-Mediated N6-Methyladenosine Modification Is Involved in Oxidative Stress in Human Aortic Dissection

doi: 10.1155/2023/3918393

Figure Lengend Snippet: IF staining of DAPI, YTHDC1, and CD31 and their merged images.

Article Snippet: After cell spreading, HUVECs were transfected with 75 nM YTHDC1 siRNAs or NC siRNAs for 48 h by riboFECT™ CP Transfection Kit (C10511-05, RiboBio, Guangzhou, China).

Techniques: Staining

Journal: Oxidative Medicine and Cellular Longevity

Article Title: The Effect of N6-Methyladenosine Regulators and m6A Reader YTHDC1-Mediated N6-Methyladenosine Modification Is Involved in Oxidative Stress in Human Aortic Dissection

doi: 10.1155/2023/3918393

Figure Lengend Snippet: Effects of YTHDC1 on ROS and inflammation. “ ∗ ” means P < 0.05, “ ∗∗ ” means P < 0.01, and “ ∗∗∗∗ ” means P < 0.0001. (a) YTHDC1 knockdown efficiency. (b) ROS levels of the si-YThDC1 groups and si-NC groups. (c) SOD2 relative expression of the si-YTHDC1 groups and si-NC groups. (d) PRR (NOD1, NOD2, and NLRP3) relative expression of the si-YTHDC1 groups and si-NC groups.

Article Snippet: After cell spreading, HUVECs were transfected with 75 nM YTHDC1 siRNAs or NC siRNAs for 48 h by riboFECT™ CP Transfection Kit (C10511-05, RiboBio, Guangzhou, China).

Techniques: Knockdown, Expressing

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: 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: Representative histology sections show high ( A , D , G ) and low ( B , E , H ) expression levels of METTL3, METTL14 and YTHDC1 visualized by immunohistochemistry; hematoxylin (blue) was used for nuclear staining (bright field image, 400xmagnification). Kaplan–Meier estimates show a significantly shorter 5-year survival ( p < 0.05) in patients with high expression of METTL3, ( F ) METTL14, and (I) YTHDC1. Prognostic significance remained after correction for multiple testing ( q < 0.1). Scale bar = 20 um

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, Immunohistochemistry, Staining

Journal: Biological & pharmaceutical bulletin

Article Title: N 6 -Methyladenosine Reader Protein YTHDC1 Supports Proper Mitotic Progression Partly through Regulation of TPX2-Aurora A Signaling.

doi: 10.1248/bpb.b24-00542

Figure Lengend Snippet: Fig. 2. YTHDC1 Knockdown Prolongs the Duration of the Early Stage of Mitosis

Article Snippet: Doxycycline (Dox)-inducible cell lines expressing FLAG and HA-tagged wild-type or mutant YTHDC1 (A549/YTHDC1-WT or -W377A/W428A) were established as previously described.11) Plasmids pcDNA3-FLAG-HA-hYTHDC1 was a gift from Samie Jaffrey (Addgene plasmid # 85167; http://n2t.net/ addgene:85167; RRID:Addgene_85167).12) FLAG-HA epitopetagged YTHDC1 was subcloned into pENTR4-no-ccDB (686-1) [a gift from Eric Campeau & Paul Kaufman (Addgene plasmid # 17424; http://n2t.net/addgene:17424; RRID: Addgene_17424)].13) The small interfering RNA (siRNA)-resistant YTHDC1 plasmid was created by introducing a silent mutation into the siYTHDC1#1 target site by PCR using the following primers: (sense) 5′-CTGGTCTCCAAGCCACTGAGCTCATCTGTTAGC-3′ and (antisense) 5′-CTGCCTCGAATGGACAGAAGGCTTTTGTCG-3′.

Techniques: Knockdown

Journal: Biological & pharmaceutical bulletin

Article Title: N 6 -Methyladenosine Reader Protein YTHDC1 Supports Proper Mitotic Progression Partly through Regulation of TPX2-Aurora A Signaling.

doi: 10.1248/bpb.b24-00542

Figure Lengend Snippet: Fig. 3. The Spindle Assembly Checkpoint Is Involved in the YTHDC1 Knockdown-Induced Delay in Mitotic Progression

Article Snippet: Doxycycline (Dox)-inducible cell lines expressing FLAG and HA-tagged wild-type or mutant YTHDC1 (A549/YTHDC1-WT or -W377A/W428A) were established as previously described.11) Plasmids pcDNA3-FLAG-HA-hYTHDC1 was a gift from Samie Jaffrey (Addgene plasmid # 85167; http://n2t.net/ addgene:85167; RRID:Addgene_85167).12) FLAG-HA epitopetagged YTHDC1 was subcloned into pENTR4-no-ccDB (686-1) [a gift from Eric Campeau & Paul Kaufman (Addgene plasmid # 17424; http://n2t.net/addgene:17424; RRID: Addgene_17424)].13) The small interfering RNA (siRNA)-resistant YTHDC1 plasmid was created by introducing a silent mutation into the siYTHDC1#1 target site by PCR using the following primers: (sense) 5′-CTGGTCTCCAAGCCACTGAGCTCATCTGTTAGC-3′ and (antisense) 5′-CTGCCTCGAATGGACAGAAGGCTTTTGTCG-3′.

Techniques: Knockdown

Journal: Biological & pharmaceutical bulletin

Article Title: N 6 -Methyladenosine Reader Protein YTHDC1 Supports Proper Mitotic Progression Partly through Regulation of TPX2-Aurora A Signaling.

doi: 10.1248/bpb.b24-00542

Figure Lengend Snippet: Fig. 4. YTHDC1 Supports Mitotic Progression Partly through Regulation of Centrosomal Abundance of Aurora A

Article Snippet: Doxycycline (Dox)-inducible cell lines expressing FLAG and HA-tagged wild-type or mutant YTHDC1 (A549/YTHDC1-WT or -W377A/W428A) were established as previously described.11) Plasmids pcDNA3-FLAG-HA-hYTHDC1 was a gift from Samie Jaffrey (Addgene plasmid # 85167; http://n2t.net/ addgene:85167; RRID:Addgene_85167).12) FLAG-HA epitopetagged YTHDC1 was subcloned into pENTR4-no-ccDB (686-1) [a gift from Eric Campeau & Paul Kaufman (Addgene plasmid # 17424; http://n2t.net/addgene:17424; RRID: Addgene_17424)].13) The small interfering RNA (siRNA)-resistant YTHDC1 plasmid was created by introducing a silent mutation into the siYTHDC1#1 target site by PCR using the following primers: (sense) 5′-CTGGTCTCCAAGCCACTGAGCTCATCTGTTAGC-3′ and (antisense) 5′-CTGCCTCGAATGGACAGAAGGCTTTTGTCG-3′.

Techniques:

Journal: Biological & pharmaceutical bulletin

Article Title: N 6 -Methyladenosine Reader Protein YTHDC1 Supports Proper Mitotic Progression Partly through Regulation of TPX2-Aurora A Signaling.

doi: 10.1248/bpb.b24-00542

Figure Lengend Snippet: Fig. 5. YTHDC1 Represses the Centrosomal Abundance of Aurora A through the Regulation of TPX2 Expression

Article Snippet: Doxycycline (Dox)-inducible cell lines expressing FLAG and HA-tagged wild-type or mutant YTHDC1 (A549/YTHDC1-WT or -W377A/W428A) were established as previously described.11) Plasmids pcDNA3-FLAG-HA-hYTHDC1 was a gift from Samie Jaffrey (Addgene plasmid # 85167; http://n2t.net/ addgene:85167; RRID:Addgene_85167).12) FLAG-HA epitopetagged YTHDC1 was subcloned into pENTR4-no-ccDB (686-1) [a gift from Eric Campeau & Paul Kaufman (Addgene plasmid # 17424; http://n2t.net/addgene:17424; RRID: Addgene_17424)].13) The small interfering RNA (siRNA)-resistant YTHDC1 plasmid was created by introducing a silent mutation into the siYTHDC1#1 target site by PCR using the following primers: (sense) 5′-CTGGTCTCCAAGCCACTGAGCTCATCTGTTAGC-3′ and (antisense) 5′-CTGCCTCGAATGGACAGAAGGCTTTTGTCG-3′.

Techniques: Expressing