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rabbit anti fto antibody (Proteintech)

Name: rabbit anti fto antibody
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Proteintech rabbit anti fto antibody

<t>SRSF6</t> depletion inhibits tumor growth through the ablation of <t>FTO</t> expression in HNSC. A and B , dot blot assay analyzed the total RNA m6A abundance of CAL 27 or SCC-9 cells with knockdown of SRSF6 ( A ) or with forced overexpression of SRSF6 ( B ). siNC ( A ) and empty vector ( B ) were used as negative controls, respectively. Methylene blue (MB) staining served as a loading control. The histograms below summarize the statistical results of the dot blot assay. C and D , Western blot analysis of FTO and SRSF6 after CAL 27 or SCC-9 cells silencing ( C ) or overexpressing ( D ) SRSF6. siNC ( C ) and empty vector ( D ) were used as negative controls, respectively. GAPDH served as a loading control. The histograms below summarize the relative expression levels of FTO. E and F , the analysis of mRNA levels of FTO and SRSF6 in tumor tissues and normal tissues from the TCGA database. The gene expression data were downloaded from the oncoDB website. E , Spearman correlation between FTO and SRSF6 in cancers (including tumor tissues and normal tissues, Spearman r > 0.1, p < 0.05). F , statistical analysis of FTO mRNA expression in tumor tissues and normal tissues in HNSC. G – I , CAL 27 and SCC-9 cells were transfected with specific siRNA targeting FTO (siFTO) and negative control siRNA (siNC). G , cell proliferation curves. H , cell colony formation analysis. The colonies with more than 50 cells were counted. I , verification of FTO knockdown by siFTO using Western blot. GAPDH served as a loading control. J and K , CAL 27 and SCC-9 cells were treated with 0, 0.5, 2, and 5 μM FTO inhibitor FB23-2. J , cell colony formation analysis. Cell clusters with more than 50 cells were counted as clones. K , cell proliferation curves. L – O , CAL 27 cells stably expressing shSRSF6 or shNC were stably cotransfected with FTO-FLAG or empty vector control plasmids. Then the cells were injected into nude mice subcutaneously. L , Western blot analyzed the expression of FTO and SRSF6 in CAL 27 cells. GAPDH served as a loading control. M , growth curve of tumors. Tumor volume was calculated as (length × width 2 × π/6). N and O , the mice were sacrificed at day 47, and subcutaneous tumors were collected, weighed, and photographed immediately. Data are means ± SD. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. FTO, fat mass and obesity-associated protein; HNSC, head and neck squamous cell carcinoma; m6A, N6-methyladenosine; SRSF, serine and arginine rich splicing factor; TCGA, The Cancer Genome Atlas.

Rabbit Anti Fto Antibody, supplied by Proteintech, used in various techniques. Bioz Stars score: 96/100, based on 225 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "The splicing factor SRSF6 mediates ferroptosis resistance in head and neck squamous cell carcinoma through induction of stearoyl-CoA desaturase"

Article Title: The splicing factor SRSF6 mediates ferroptosis resistance in head and neck squamous cell carcinoma through induction of stearoyl-CoA desaturase

Journal: The Journal of Biological Chemistry

doi: 10.1016/j.jbc.2025.110509

SRSF6 depletion inhibits tumor growth through the ablation of FTO expression in HNSC. A and B , dot blot assay analyzed the total RNA m6A abundance of CAL 27 or SCC-9 cells with knockdown of SRSF6 ( A ) or with forced overexpression of SRSF6 ( B ). siNC ( A ) and empty vector ( B ) were used as negative controls, respectively. Methylene blue (MB) staining served as a loading control. The histograms below summarize the statistical results of the dot blot assay. C and D , Western blot analysis of FTO and SRSF6 after CAL 27 or SCC-9 cells silencing ( C ) or overexpressing ( D ) SRSF6. siNC ( C ) and empty vector ( D ) were used as negative controls, respectively. GAPDH served as a loading control. The histograms below summarize the relative expression levels of FTO. E and F , the analysis of mRNA levels of FTO and SRSF6 in tumor tissues and normal tissues from the TCGA database. The gene expression data were downloaded from the oncoDB website. E , Spearman correlation between FTO and SRSF6 in cancers (including tumor tissues and normal tissues, Spearman r > 0.1, p < 0.05). F , statistical analysis of FTO mRNA expression in tumor tissues and normal tissues in HNSC. G – I , CAL 27 and SCC-9 cells were transfected with specific siRNA targeting FTO (siFTO) and negative control siRNA (siNC). G , cell proliferation curves. H , cell colony formation analysis. The colonies with more than 50 cells were counted. I , verification of FTO knockdown by siFTO using Western blot. GAPDH served as a loading control. J and K , CAL 27 and SCC-9 cells were treated with 0, 0.5, 2, and 5 μM FTO inhibitor FB23-2. J , cell colony formation analysis. Cell clusters with more than 50 cells were counted as clones. K , cell proliferation curves. L – O , CAL 27 cells stably expressing shSRSF6 or shNC were stably cotransfected with FTO-FLAG or empty vector control plasmids. Then the cells were injected into nude mice subcutaneously. L , Western blot analyzed the expression of FTO and SRSF6 in CAL 27 cells. GAPDH served as a loading control. M , growth curve of tumors. Tumor volume was calculated as (length × width 2 × π/6). N and O , the mice were sacrificed at day 47, and subcutaneous tumors were collected, weighed, and photographed immediately. Data are means ± SD. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. FTO, fat mass and obesity-associated protein; HNSC, head and neck squamous cell carcinoma; m6A, N6-methyladenosine; SRSF, serine and arginine rich splicing factor; TCGA, The Cancer Genome Atlas.

Figure Legend Snippet: SRSF6 depletion inhibits tumor growth through the ablation of FTO expression in HNSC. A and B , dot blot assay analyzed the total RNA m6A abundance of CAL 27 or SCC-9 cells with knockdown of SRSF6 ( A ) or with forced overexpression of SRSF6 ( B ). siNC ( A ) and empty vector ( B ) were used as negative controls, respectively. Methylene blue (MB) staining served as a loading control. The histograms below summarize the statistical results of the dot blot assay. C and D , Western blot analysis of FTO and SRSF6 after CAL 27 or SCC-9 cells silencing ( C ) or overexpressing ( D ) SRSF6. siNC ( C ) and empty vector ( D ) were used as negative controls, respectively. GAPDH served as a loading control. The histograms below summarize the relative expression levels of FTO. E and F , the analysis of mRNA levels of FTO and SRSF6 in tumor tissues and normal tissues from the TCGA database. The gene expression data were downloaded from the oncoDB website. E , Spearman correlation between FTO and SRSF6 in cancers (including tumor tissues and normal tissues, Spearman r > 0.1, p < 0.05). F , statistical analysis of FTO mRNA expression in tumor tissues and normal tissues in HNSC. G – I , CAL 27 and SCC-9 cells were transfected with specific siRNA targeting FTO (siFTO) and negative control siRNA (siNC). G , cell proliferation curves. H , cell colony formation analysis. The colonies with more than 50 cells were counted. I , verification of FTO knockdown by siFTO using Western blot. GAPDH served as a loading control. J and K , CAL 27 and SCC-9 cells were treated with 0, 0.5, 2, and 5 μM FTO inhibitor FB23-2. J , cell colony formation analysis. Cell clusters with more than 50 cells were counted as clones. K , cell proliferation curves. L – O , CAL 27 cells stably expressing shSRSF6 or shNC were stably cotransfected with FTO-FLAG or empty vector control plasmids. Then the cells were injected into nude mice subcutaneously. L , Western blot analyzed the expression of FTO and SRSF6 in CAL 27 cells. GAPDH served as a loading control. M , growth curve of tumors. Tumor volume was calculated as (length × width 2 × π/6). N and O , the mice were sacrificed at day 47, and subcutaneous tumors were collected, weighed, and photographed immediately. Data are means ± SD. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. FTO, fat mass and obesity-associated protein; HNSC, head and neck squamous cell carcinoma; m6A, N6-methyladenosine; SRSF, serine and arginine rich splicing factor; TCGA, The Cancer Genome Atlas.

Techniques Used: Expressing, Dot Blot, Knockdown, Over Expression, Plasmid Preparation, Staining, Control, Western Blot, Gene Expression, Transfection, Negative Control, Clone Assay, Stable Transfection, Injection

FTO promotes ferroptosis resistance in HNSC cells. A and B , analysis of cytosolic ROS generation ( A ) or lipid peroxidation ( B ) by flow cytometry after CAL 27 and SCC-9 cells transfected with siFTO or siNC. C and D , measurement of cytosolic ROS generation ( C ) and lipid peroxidation ( D ) by flow cytometry after CAL 27 and SCC-9 cells treated with FB23-2 or DMSO. E – I , after transfected with siFTO and siNC ( E – G ), or treated with FB23-2 and DMSO ( H and I ), CAL 27 and SCC-9 cells were treated with Fer-1 and DMSO. Flow cytometry was used to detect the cytosolic ROS ( E and H ) and lipid peroxidation ( F and I ). Western blot was used to detect the knockdown efficiency of siFTO and GAPDH served as a loading control ( G ). J – L , CAL 27 and SCC-9 cells stably overexpressing FTO or empty control vector were treated with RSL3 or DMSO. Flow cytometry was used to analyze the cytosolic ROS ( J ) and lipid peroxidation ( K ). Western blot was used to analyze the overexpression of FTO-FLAG ( L ). Data are means ± SD, n = 3 or 4. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. DMSO, dimethyl sulfoxide; Fer-1, ferrostatin-1; FTO, fat mass and obesity-associated protein; HNSC, head and neck squamous cell carcinoma; ROS, reactive oxygen species; RSL3, RAS-selective lethal 3.

Figure Legend Snippet: FTO promotes ferroptosis resistance in HNSC cells. A and B , analysis of cytosolic ROS generation ( A ) or lipid peroxidation ( B ) by flow cytometry after CAL 27 and SCC-9 cells transfected with siFTO or siNC. C and D , measurement of cytosolic ROS generation ( C ) and lipid peroxidation ( D ) by flow cytometry after CAL 27 and SCC-9 cells treated with FB23-2 or DMSO. E – I , after transfected with siFTO and siNC ( E – G ), or treated with FB23-2 and DMSO ( H and I ), CAL 27 and SCC-9 cells were treated with Fer-1 and DMSO. Flow cytometry was used to detect the cytosolic ROS ( E and H ) and lipid peroxidation ( F and I ). Western blot was used to detect the knockdown efficiency of siFTO and GAPDH served as a loading control ( G ). J – L , CAL 27 and SCC-9 cells stably overexpressing FTO or empty control vector were treated with RSL3 or DMSO. Flow cytometry was used to analyze the cytosolic ROS ( J ) and lipid peroxidation ( K ). Western blot was used to analyze the overexpression of FTO-FLAG ( L ). Data are means ± SD, n = 3 or 4. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. DMSO, dimethyl sulfoxide; Fer-1, ferrostatin-1; FTO, fat mass and obesity-associated protein; HNSC, head and neck squamous cell carcinoma; ROS, reactive oxygen species; RSL3, RAS-selective lethal 3.

Techniques Used: Flow Cytometry, Transfection, Western Blot, Knockdown, Control, Stable Transfection, Plasmid Preparation, Over Expression

FTO promotes the expression of SRSF6 through modulating RNA m6A modification. A , MeRIP-PCR analyzed the m6A modification on SRSF6 mRNA. B , the diagram of the predicted m6A site in SRSF6 mRNA 3′ UTR near the stop codon. The SRSF6 expression plasmids containing the wild-type (T7-SRSF6-m6A-wt) or mutant (T7-SRSF6-m6A-mt) m6A sites were constructed. C , MeRIP-PCR analyzed the m6A modification on mRNA transcripts of T7-SRSF6 in HEK 293 cells transduced with T7-SRSF6-m6A-wt or T7-SRSF6-m6A-mt plasmids. D – I , CAL 27 and SCC-9 cells were treated with FB23-2 and DMSO ( D and G ), transfected with siFTO and siNC ( E and H ), or transfected with FTO-FLAG and empty vector control plasmids ( F and I ). D – F , RT-PCR analyzed the mRNA expression of SRSF6, and β-actin served as a loading control. G – I , the protein expression of SRSF6 ( G – I ) and FTO ( H and I ) was analyzed by Western blot, and GAPDH served as a loading control. J , MeRIP-PCR analyzed the m6A modification on SRSF6 mRNA after cells transfected with siFTO and siNC. K , RT-qPCR analysis of the mRNA stability of SRSF6 in CAL 27 cells transfected with siNC or siFTO. β-actin served as a loading control. Data are means ± SD. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. DMSO, dimethyl sulfoxide; FTO, fat mass and obesity-associated protein; m6A, N6-methyladenosine; RT-qPCR, real-time quantitative reverse transcription PCR; RT-PCR, reverse transcription PCR; SRSF, serine and arginine rich splicing factor.

Figure Legend Snippet: FTO promotes the expression of SRSF6 through modulating RNA m6A modification. A , MeRIP-PCR analyzed the m6A modification on SRSF6 mRNA. B , the diagram of the predicted m6A site in SRSF6 mRNA 3′ UTR near the stop codon. The SRSF6 expression plasmids containing the wild-type (T7-SRSF6-m6A-wt) or mutant (T7-SRSF6-m6A-mt) m6A sites were constructed. C , MeRIP-PCR analyzed the m6A modification on mRNA transcripts of T7-SRSF6 in HEK 293 cells transduced with T7-SRSF6-m6A-wt or T7-SRSF6-m6A-mt plasmids. D – I , CAL 27 and SCC-9 cells were treated with FB23-2 and DMSO ( D and G ), transfected with siFTO and siNC ( E and H ), or transfected with FTO-FLAG and empty vector control plasmids ( F and I ). D – F , RT-PCR analyzed the mRNA expression of SRSF6, and β-actin served as a loading control. G – I , the protein expression of SRSF6 ( G – I ) and FTO ( H and I ) was analyzed by Western blot, and GAPDH served as a loading control. J , MeRIP-PCR analyzed the m6A modification on SRSF6 mRNA after cells transfected with siFTO and siNC. K , RT-qPCR analysis of the mRNA stability of SRSF6 in CAL 27 cells transfected with siNC or siFTO. β-actin served as a loading control. Data are means ± SD. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. DMSO, dimethyl sulfoxide; FTO, fat mass and obesity-associated protein; m6A, N6-methyladenosine; RT-qPCR, real-time quantitative reverse transcription PCR; RT-PCR, reverse transcription PCR; SRSF, serine and arginine rich splicing factor.

Techniques Used: Expressing, Modification, Mutagenesis, Construct, Transduction, Transfection, Plasmid Preparation, Control, Reverse Transcription Polymerase Chain Reaction, Western Blot, Quantitative RT-PCR, Reverse Transcription

FTO or SRSF6 silencing inhibits SREBF1/SCD expression. A – F , RNA sequencing (RNA-seq) and bioinformatic analysis in CAL 27 cells transfected with siFTO, siSRSF6-1#, and siNC. Three biological replicates were conducted per group. A , flow chart of the RNA-seq. B , the Venn diagram of the differentially expressed genes (DEGs) was created through the online tool EVenn. The thresholds of significant DEGs were | log 2 Fold Change (log 2 FC) | > 0.3 and P adj < 0.05. C , the WikiPathways (WP) enrichment analysis of the common downstream genes of SRSF6 and FTO analyzed by the Venn diagram in ( B ) was conducted using the online website DAVID. The top 20 signaling pathways ( p < 0.05) were selected to draw the bubble diagram using the website Hiplot. D , the GSEA enrichment analysis of all genes in “WP-ferroptosis” pathways using the GSEA tool. E , heatmap of the log 2 FC values of genes in “WP-Sterol regulatory element-binding proteins (SREBP) signaling” and “WP-ferroptosis” pathways. P adj < 0.05. F , the GSEA enrichment analysis of all genes in “WP-Sterol regulatory element-binding proteins (SREBP) signaling” pathways using the GSEA tool. G , MeRIP-PCR analyzed the m6A modification on SREBF1 and SCD mRNA after CAL 27 cells transfected with siFTO and siNC. H – L , RT-PCR analysis of SREBF1 and SCD expression after cells treated with FB23-2 and DMSO ( H ), or transfected with siFTO and siNC ( I ), FTO-FLAG and control vector plasmids ( J ), siSRSF6 and siNC ( K ), as well as T7-SRSF6 and control vector plasmids ( L ). β-actin served as a loading control. Data are means ± SD, n = 3, 4, or 5. M , RT-PCR analysis of SREBF1 and SCD expression after CAL 27 cells stably cotransfected with T7-SRSF6 (empty vector as control) and shFTO (shNC as control) plasmids. β-actin served as a loading control. N and O , RT-qPCR analysis of the mRNA stability of SREBF1 ( N ) and SCD ( O ) in CAL 27 cells transfected with siNC or siFTO. β-actin served as a loading control. Data are means ± SD. P and Q , the analysis of mRNA levels of SREBF1 ( P ) and SCD ( Q ) in tumor tissues and normal tissues of HNSC from the TCGA database. The gene expression data were downloaded from the oncoDB website. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. DMSO, dimethyl sulfoxide; FTO, fat mass and obesity-associated protein; GSEA, Gene Set Enrichment Analysis; HNSC, head and neck squamous cell carcinoma; m6A, N6-methyladenosine; RT-qPCR, real-time quantitative reverse transcription PCR; RT-PCR, reverse transcription PCR; SCD, stearoyl-CoA desaturase; SRSF, serine and arginine rich splicing factor; SREBF, sterol regulatory element binding transcription factor; TCGA, The Cancer Genome Atlas.

Figure Legend Snippet: FTO or SRSF6 silencing inhibits SREBF1/SCD expression. A – F , RNA sequencing (RNA-seq) and bioinformatic analysis in CAL 27 cells transfected with siFTO, siSRSF6-1#, and siNC. Three biological replicates were conducted per group. A , flow chart of the RNA-seq. B , the Venn diagram of the differentially expressed genes (DEGs) was created through the online tool EVenn. The thresholds of significant DEGs were | log 2 Fold Change (log 2 FC) | > 0.3 and P adj < 0.05. C , the WikiPathways (WP) enrichment analysis of the common downstream genes of SRSF6 and FTO analyzed by the Venn diagram in ( B ) was conducted using the online website DAVID. The top 20 signaling pathways ( p < 0.05) were selected to draw the bubble diagram using the website Hiplot. D , the GSEA enrichment analysis of all genes in “WP-ferroptosis” pathways using the GSEA tool. E , heatmap of the log 2 FC values of genes in “WP-Sterol regulatory element-binding proteins (SREBP) signaling” and “WP-ferroptosis” pathways. P adj < 0.05. F , the GSEA enrichment analysis of all genes in “WP-Sterol regulatory element-binding proteins (SREBP) signaling” pathways using the GSEA tool. G , MeRIP-PCR analyzed the m6A modification on SREBF1 and SCD mRNA after CAL 27 cells transfected with siFTO and siNC. H – L , RT-PCR analysis of SREBF1 and SCD expression after cells treated with FB23-2 and DMSO ( H ), or transfected with siFTO and siNC ( I ), FTO-FLAG and control vector plasmids ( J ), siSRSF6 and siNC ( K ), as well as T7-SRSF6 and control vector plasmids ( L ). β-actin served as a loading control. Data are means ± SD, n = 3, 4, or 5. M , RT-PCR analysis of SREBF1 and SCD expression after CAL 27 cells stably cotransfected with T7-SRSF6 (empty vector as control) and shFTO (shNC as control) plasmids. β-actin served as a loading control. N and O , RT-qPCR analysis of the mRNA stability of SREBF1 ( N ) and SCD ( O ) in CAL 27 cells transfected with siNC or siFTO. β-actin served as a loading control. Data are means ± SD. P and Q , the analysis of mRNA levels of SREBF1 ( P ) and SCD ( Q ) in tumor tissues and normal tissues of HNSC from the TCGA database. The gene expression data were downloaded from the oncoDB website. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. DMSO, dimethyl sulfoxide; FTO, fat mass and obesity-associated protein; GSEA, Gene Set Enrichment Analysis; HNSC, head and neck squamous cell carcinoma; m6A, N6-methyladenosine; RT-qPCR, real-time quantitative reverse transcription PCR; RT-PCR, reverse transcription PCR; SCD, stearoyl-CoA desaturase; SRSF, serine and arginine rich splicing factor; SREBF, sterol regulatory element binding transcription factor; TCGA, The Cancer Genome Atlas.

Techniques Used: Expressing, RNA Sequencing, Transfection, Protein-Protein interactions, Binding Assay, Modification, Reverse Transcription Polymerase Chain Reaction, Control, Plasmid Preparation, Stable Transfection, Quantitative RT-PCR, Gene Expression, Reverse Transcription

Knockdown of SRSF6 or FTO induces ferroptosis through the inhibition of SREBF1/SCD. A – F , CAL 27 cells with or without SREBF1-FLAG overexpression were transfected with siSRSF6 ( A – C ) or siFTO ( D – F ). The cytosolic ROS level ( A and D ) and lipid peroxidation ( B and E ) were detected by flow cytometry. The overexpression of SREBF1-FLAG ( C and F ) and the knockdown efficiency of siSRSF6 ( C ) or siFTO ( F ) were analyzed by Western blot. GAPDH served as a loading control. G – L , Knockdown of SRSF6 ( G – I ) or FTO ( J – L ) was conducted by siRNAs in CAL 27 cells with or without SCD-3×FLAG overexpression. Flow cytometry was used to analyze the cytosolic ROS level ( G and J ) and lipid peroxidation ( H and K ). Western blot analyzed the overexpression of SCD-3×FLAG ( I and L ) and the silence efficiency of siSRSF6 ( I ) or siFTO ( L ), and GAPDH served as a loading control. Data are means ± SD, n = 4. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. FTO, fat mass and obesity-associated protein; ROS, reactive oxygen species; SCD, stearoyl-CoA desaturase; SREBF, sterol regulatory element binding transcription factor; SRSF, serine and arginine rich splicing factor.

Figure Legend Snippet: Knockdown of SRSF6 or FTO induces ferroptosis through the inhibition of SREBF1/SCD. A – F , CAL 27 cells with or without SREBF1-FLAG overexpression were transfected with siSRSF6 ( A – C ) or siFTO ( D – F ). The cytosolic ROS level ( A and D ) and lipid peroxidation ( B and E ) were detected by flow cytometry. The overexpression of SREBF1-FLAG ( C and F ) and the knockdown efficiency of siSRSF6 ( C ) or siFTO ( F ) were analyzed by Western blot. GAPDH served as a loading control. G – L , Knockdown of SRSF6 ( G – I ) or FTO ( J – L ) was conducted by siRNAs in CAL 27 cells with or without SCD-3×FLAG overexpression. Flow cytometry was used to analyze the cytosolic ROS level ( G and J ) and lipid peroxidation ( H and K ). Western blot analyzed the overexpression of SCD-3×FLAG ( I and L ) and the silence efficiency of siSRSF6 ( I ) or siFTO ( L ), and GAPDH served as a loading control. Data are means ± SD, n = 4. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. FTO, fat mass and obesity-associated protein; ROS, reactive oxygen species; SCD, stearoyl-CoA desaturase; SREBF, sterol regulatory element binding transcription factor; SRSF, serine and arginine rich splicing factor.

Techniques Used: Knockdown, Inhibition, Over Expression, Transfection, Flow Cytometry, Western Blot, Control, Binding Assay

Schematic model for the mechanism by which HNSC cells escape from ferroptosis through the SRSF6/FTO/SREBF1/SCD axis. FTO, fat mass and obesity-associated protein; HNSC, head and neck squamous cell carcinoma; SCD, stearoyl-CoA desaturase; SREBF, sterol regulatory element binding transcription factor; SRSF, serine and arginine rich splicing factor.

Figure Legend Snippet: Schematic model for the mechanism by which HNSC cells escape from ferroptosis through the SRSF6/FTO/SREBF1/SCD axis. FTO, fat mass and obesity-associated protein; HNSC, head and neck squamous cell carcinoma; SCD, stearoyl-CoA desaturase; SREBF, sterol regulatory element binding transcription factor; SRSF, serine and arginine rich splicing factor.

Techniques Used: Binding Assay

Image Search Results

VP1 up-regulated the expressions of key elements involved in m 6 A modification in MSCs. A-I, qRT-PCR analysis of the mRNA levels of EV71-VP1, METTLE 3/14, YTHDC1, YTHDF 1/2/3, FTO and ALKBH5 in MSCs transfected with PEGFP-C3-VP1 plasmid for 24 h. N = 3. * p < 0.05 compared with the NC group. J-K, Western blot analysis and quantification of protein expressions of EV71-VP1 and key elements involved in m 6 A modification in MSCs transfected with PEGFP-C3-VP1 plasmid for 48 h, including ALKBH5, FTO, METTLE 3/14, YTHDF 1/2/3, and YTHDC1. GAPDH was used as internal control. N = 3. * p < 0.05 compared with the NC group.

Journal: Virus Research

Article Title: Enterovirus 71 structural viral protein 1 promotes the expression of PMP22 through m 6 A modification in mouse Schwann cells

doi: 10.1016/j.virusres.2025.199590

Figure Lengend Snippet: VP1 up-regulated the expressions of key elements involved in m 6 A modification in MSCs. A-I, qRT-PCR analysis of the mRNA levels of EV71-VP1, METTLE 3/14, YTHDC1, YTHDF 1/2/3, FTO and ALKBH5 in MSCs transfected with PEGFP-C3-VP1 plasmid for 24 h. N = 3. * p < 0.05 compared with the NC group. J-K, Western blot analysis and quantification of protein expressions of EV71-VP1 and key elements involved in m 6 A modification in MSCs transfected with PEGFP-C3-VP1 plasmid for 48 h, including ALKBH5, FTO, METTLE 3/14, YTHDF 1/2/3, and YTHDC1. GAPDH was used as internal control. N = 3. * p < 0.05 compared with the NC group.

Article Snippet: The primary antibodies used in this study were: anti-GAPDH antibody (1:1000, Abcam, Cat. # ab181602); Enterovirus 71 VP1 monoclonal antibody (1:1000, Thermo Fisher, Cat. # MA5–33,257); Rabbit anti-METTL14 antibody (1:1000, Bioss, Cat. # bs-17608R); Rabbit anti-FTO antibody (1:1000, Bioss, Cat. # bs-7056R); Rabbit anti-ALKBH5 antibody (1:1000, Bioss, Cat. # bs-20540R); Recombinant anti-METTL3 antibody [ EPR18810 ] (1:1000, Abcam, Cat. # ab195352); Recombinant Anti-YTHDF1 antibody [ EPR22349 –16] (1:1000, Abcam, Cat. # ab252346); Recombinant Anti-YTHDF2 antibody [ EPR20318 ] (1:1000, Abcam, Cat. # ab220163); Recombinant Anti-YTHDF3 antibody [ EPR21912 –3] (1:1000, Abcam, Cat. # ab220161); Recombinant Anti-YTHDC1 antibody [ EPR21821 ] (1:1000, Abcam, Cat. # ab220159).

Techniques: Modification, Quantitative RT-PCR, Transfection, Plasmid Preparation, Western Blot, Control

Journal: The Journal of Biological Chemistry

Article Title: The splicing factor SRSF6 mediates ferroptosis resistance in head and neck squamous cell carcinoma through induction of stearoyl-CoA desaturase

doi: 10.1016/j.jbc.2025.110509

Figure Lengend Snippet: SRSF6 depletion inhibits tumor growth through the ablation of FTO expression in HNSC. A and B , dot blot assay analyzed the total RNA m6A abundance of CAL 27 or SCC-9 cells with knockdown of SRSF6 ( A ) or with forced overexpression of SRSF6 ( B ). siNC ( A ) and empty vector ( B ) were used as negative controls, respectively. Methylene blue (MB) staining served as a loading control. The histograms below summarize the statistical results of the dot blot assay. C and D , Western blot analysis of FTO and SRSF6 after CAL 27 or SCC-9 cells silencing ( C ) or overexpressing ( D ) SRSF6. siNC ( C ) and empty vector ( D ) were used as negative controls, respectively. GAPDH served as a loading control. The histograms below summarize the relative expression levels of FTO. E and F , the analysis of mRNA levels of FTO and SRSF6 in tumor tissues and normal tissues from the TCGA database. The gene expression data were downloaded from the oncoDB website. E , Spearman correlation between FTO and SRSF6 in cancers (including tumor tissues and normal tissues, Spearman r > 0.1, p < 0.05). F , statistical analysis of FTO mRNA expression in tumor tissues and normal tissues in HNSC. G – I , CAL 27 and SCC-9 cells were transfected with specific siRNA targeting FTO (siFTO) and negative control siRNA (siNC). G , cell proliferation curves. H , cell colony formation analysis. The colonies with more than 50 cells were counted. I , verification of FTO knockdown by siFTO using Western blot. GAPDH served as a loading control. J and K , CAL 27 and SCC-9 cells were treated with 0, 0.5, 2, and 5 μM FTO inhibitor FB23-2. J , cell colony formation analysis. Cell clusters with more than 50 cells were counted as clones. K , cell proliferation curves. L – O , CAL 27 cells stably expressing shSRSF6 or shNC were stably cotransfected with FTO-FLAG or empty vector control plasmids. Then the cells were injected into nude mice subcutaneously. L , Western blot analyzed the expression of FTO and SRSF6 in CAL 27 cells. GAPDH served as a loading control. M , growth curve of tumors. Tumor volume was calculated as (length × width 2 × π/6). N and O , the mice were sacrificed at day 47, and subcutaneous tumors were collected, weighed, and photographed immediately. Data are means ± SD. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. FTO, fat mass and obesity-associated protein; HNSC, head and neck squamous cell carcinoma; m6A, N6-methyladenosine; SRSF, serine and arginine rich splicing factor; TCGA, The Cancer Genome Atlas.

Article Snippet: After blocking with 5% skim milk, the membrane was probed with different specific primary antibodies as follows: rabbit anti-SRSF6 antibody (#ab140623, 1:2000, Abcam), rabbit anti-FTO antibody (#27226, 1:2000, Proteintech), rabbit anti-Flag antibody (#20543, 1:2000, Proteintech), mouse anti-GAPDH antibody (#sc-47724, 1:1000, Santa Cruz Biotechnology).

Techniques: Expressing, Dot Blot, Knockdown, Over Expression, Plasmid Preparation, Staining, Control, Western Blot, Gene Expression, Transfection, Negative Control, Clone Assay, Stable Transfection, Injection

Journal: The Journal of Biological Chemistry

Article Title: The splicing factor SRSF6 mediates ferroptosis resistance in head and neck squamous cell carcinoma through induction of stearoyl-CoA desaturase

doi: 10.1016/j.jbc.2025.110509

Figure Lengend Snippet: FTO promotes ferroptosis resistance in HNSC cells. A and B , analysis of cytosolic ROS generation ( A ) or lipid peroxidation ( B ) by flow cytometry after CAL 27 and SCC-9 cells transfected with siFTO or siNC. C and D , measurement of cytosolic ROS generation ( C ) and lipid peroxidation ( D ) by flow cytometry after CAL 27 and SCC-9 cells treated with FB23-2 or DMSO. E – I , after transfected with siFTO and siNC ( E – G ), or treated with FB23-2 and DMSO ( H and I ), CAL 27 and SCC-9 cells were treated with Fer-1 and DMSO. Flow cytometry was used to detect the cytosolic ROS ( E and H ) and lipid peroxidation ( F and I ). Western blot was used to detect the knockdown efficiency of siFTO and GAPDH served as a loading control ( G ). J – L , CAL 27 and SCC-9 cells stably overexpressing FTO or empty control vector were treated with RSL3 or DMSO. Flow cytometry was used to analyze the cytosolic ROS ( J ) and lipid peroxidation ( K ). Western blot was used to analyze the overexpression of FTO-FLAG ( L ). Data are means ± SD, n = 3 or 4. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. DMSO, dimethyl sulfoxide; Fer-1, ferrostatin-1; FTO, fat mass and obesity-associated protein; HNSC, head and neck squamous cell carcinoma; ROS, reactive oxygen species; RSL3, RAS-selective lethal 3.

Techniques: Flow Cytometry, Transfection, Western Blot, Knockdown, Control, Stable Transfection, Plasmid Preparation, Over Expression

Journal: The Journal of Biological Chemistry

Article Title: The splicing factor SRSF6 mediates ferroptosis resistance in head and neck squamous cell carcinoma through induction of stearoyl-CoA desaturase

doi: 10.1016/j.jbc.2025.110509

Figure Lengend Snippet: FTO promotes the expression of SRSF6 through modulating RNA m6A modification. A , MeRIP-PCR analyzed the m6A modification on SRSF6 mRNA. B , the diagram of the predicted m6A site in SRSF6 mRNA 3′ UTR near the stop codon. The SRSF6 expression plasmids containing the wild-type (T7-SRSF6-m6A-wt) or mutant (T7-SRSF6-m6A-mt) m6A sites were constructed. C , MeRIP-PCR analyzed the m6A modification on mRNA transcripts of T7-SRSF6 in HEK 293 cells transduced with T7-SRSF6-m6A-wt or T7-SRSF6-m6A-mt plasmids. D – I , CAL 27 and SCC-9 cells were treated with FB23-2 and DMSO ( D and G ), transfected with siFTO and siNC ( E and H ), or transfected with FTO-FLAG and empty vector control plasmids ( F and I ). D – F , RT-PCR analyzed the mRNA expression of SRSF6, and β-actin served as a loading control. G – I , the protein expression of SRSF6 ( G – I ) and FTO ( H and I ) was analyzed by Western blot, and GAPDH served as a loading control. J , MeRIP-PCR analyzed the m6A modification on SRSF6 mRNA after cells transfected with siFTO and siNC. K , RT-qPCR analysis of the mRNA stability of SRSF6 in CAL 27 cells transfected with siNC or siFTO. β-actin served as a loading control. Data are means ± SD. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. DMSO, dimethyl sulfoxide; FTO, fat mass and obesity-associated protein; m6A, N6-methyladenosine; RT-qPCR, real-time quantitative reverse transcription PCR; RT-PCR, reverse transcription PCR; SRSF, serine and arginine rich splicing factor.

Techniques: Expressing, Modification, Mutagenesis, Construct, Transduction, Transfection, Plasmid Preparation, Control, Reverse Transcription Polymerase Chain Reaction, Western Blot, Quantitative RT-PCR, Reverse Transcription

Journal: The Journal of Biological Chemistry

Article Title: The splicing factor SRSF6 mediates ferroptosis resistance in head and neck squamous cell carcinoma through induction of stearoyl-CoA desaturase

doi: 10.1016/j.jbc.2025.110509

Figure Lengend Snippet: FTO or SRSF6 silencing inhibits SREBF1/SCD expression. A – F , RNA sequencing (RNA-seq) and bioinformatic analysis in CAL 27 cells transfected with siFTO, siSRSF6-1#, and siNC. Three biological replicates were conducted per group. A , flow chart of the RNA-seq. B , the Venn diagram of the differentially expressed genes (DEGs) was created through the online tool EVenn. The thresholds of significant DEGs were | log 2 Fold Change (log 2 FC) | > 0.3 and P adj < 0.05. C , the WikiPathways (WP) enrichment analysis of the common downstream genes of SRSF6 and FTO analyzed by the Venn diagram in ( B ) was conducted using the online website DAVID. The top 20 signaling pathways ( p < 0.05) were selected to draw the bubble diagram using the website Hiplot. D , the GSEA enrichment analysis of all genes in “WP-ferroptosis” pathways using the GSEA tool. E , heatmap of the log 2 FC values of genes in “WP-Sterol regulatory element-binding proteins (SREBP) signaling” and “WP-ferroptosis” pathways. P adj < 0.05. F , the GSEA enrichment analysis of all genes in “WP-Sterol regulatory element-binding proteins (SREBP) signaling” pathways using the GSEA tool. G , MeRIP-PCR analyzed the m6A modification on SREBF1 and SCD mRNA after CAL 27 cells transfected with siFTO and siNC. H – L , RT-PCR analysis of SREBF1 and SCD expression after cells treated with FB23-2 and DMSO ( H ), or transfected with siFTO and siNC ( I ), FTO-FLAG and control vector plasmids ( J ), siSRSF6 and siNC ( K ), as well as T7-SRSF6 and control vector plasmids ( L ). β-actin served as a loading control. Data are means ± SD, n = 3, 4, or 5. M , RT-PCR analysis of SREBF1 and SCD expression after CAL 27 cells stably cotransfected with T7-SRSF6 (empty vector as control) and shFTO (shNC as control) plasmids. β-actin served as a loading control. N and O , RT-qPCR analysis of the mRNA stability of SREBF1 ( N ) and SCD ( O ) in CAL 27 cells transfected with siNC or siFTO. β-actin served as a loading control. Data are means ± SD. P and Q , the analysis of mRNA levels of SREBF1 ( P ) and SCD ( Q ) in tumor tissues and normal tissues of HNSC from the TCGA database. The gene expression data were downloaded from the oncoDB website. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. DMSO, dimethyl sulfoxide; FTO, fat mass and obesity-associated protein; GSEA, Gene Set Enrichment Analysis; HNSC, head and neck squamous cell carcinoma; m6A, N6-methyladenosine; RT-qPCR, real-time quantitative reverse transcription PCR; RT-PCR, reverse transcription PCR; SCD, stearoyl-CoA desaturase; SRSF, serine and arginine rich splicing factor; SREBF, sterol regulatory element binding transcription factor; TCGA, The Cancer Genome Atlas.

Techniques: Expressing, RNA Sequencing, Transfection, Protein-Protein interactions, Binding Assay, Modification, Reverse Transcription Polymerase Chain Reaction, Control, Plasmid Preparation, Stable Transfection, Quantitative RT-PCR, Gene Expression, Reverse Transcription

Journal: The Journal of Biological Chemistry

Article Title: The splicing factor SRSF6 mediates ferroptosis resistance in head and neck squamous cell carcinoma through induction of stearoyl-CoA desaturase

doi: 10.1016/j.jbc.2025.110509

Figure Lengend Snippet: Knockdown of SRSF6 or FTO induces ferroptosis through the inhibition of SREBF1/SCD. A – F , CAL 27 cells with or without SREBF1-FLAG overexpression were transfected with siSRSF6 ( A – C ) or siFTO ( D – F ). The cytosolic ROS level ( A and D ) and lipid peroxidation ( B and E ) were detected by flow cytometry. The overexpression of SREBF1-FLAG ( C and F ) and the knockdown efficiency of siSRSF6 ( C ) or siFTO ( F ) were analyzed by Western blot. GAPDH served as a loading control. G – L , Knockdown of SRSF6 ( G – I ) or FTO ( J – L ) was conducted by siRNAs in CAL 27 cells with or without SCD-3×FLAG overexpression. Flow cytometry was used to analyze the cytosolic ROS level ( G and J ) and lipid peroxidation ( H and K ). Western blot analyzed the overexpression of SCD-3×FLAG ( I and L ) and the silence efficiency of siSRSF6 ( I ) or siFTO ( L ), and GAPDH served as a loading control. Data are means ± SD, n = 4. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. FTO, fat mass and obesity-associated protein; ROS, reactive oxygen species; SCD, stearoyl-CoA desaturase; SREBF, sterol regulatory element binding transcription factor; SRSF, serine and arginine rich splicing factor.

Techniques: Knockdown, Inhibition, Over Expression, Transfection, Flow Cytometry, Western Blot, Control, Binding Assay

Journal: The Journal of Biological Chemistry

Article Title: The splicing factor SRSF6 mediates ferroptosis resistance in head and neck squamous cell carcinoma through induction of stearoyl-CoA desaturase

doi: 10.1016/j.jbc.2025.110509

Figure Lengend Snippet: Schematic model for the mechanism by which HNSC cells escape from ferroptosis through the SRSF6/FTO/SREBF1/SCD axis. FTO, fat mass and obesity-associated protein; HNSC, head and neck squamous cell carcinoma; SCD, stearoyl-CoA desaturase; SREBF, sterol regulatory element binding transcription factor; SRSF, serine and arginine rich splicing factor.

Techniques: Binding Assay

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