Fig 1: IGF2BP3 was involved in m6A methylation modification in LSCC. a RIP-qPCR was showing the enrichment of IGF2BP3 binding to TMBIM6 m6A modification sites. b and c After knocked down or overexpressed IGF2BP3, qRT-PCR evaluated the expression of IGF2BP3 in LSCC cells. d and e MeRIP assays were performed to identify variation in m6A modification enrichment in TMBIM6 after silencing or overexpressing IGF2BP3 in LSCC cells. f and g qRT-PCR of TMBIM6 mRNA after IGF2BP3 inhibition or overexpression in LSCC cells. h RIP-qPCR unveiled the interaction within IGF2BP3 and TMBIM6 mRNA after RBM15 inhibition. i The luciferase activities in AMC-HN-8 cells co-transfected with TMBIM6-WT or TMBIM6-Mut together with shCtrl or shIGF2BP3. j The luciferase activities in TU-212 cells co-transfected with TMBIM6-WT or TMBIM6-Mut together with Vector or IGF2BP3. k qRT-PCR was used to detect IGF2BP3 expression in 34 pairs of LSCC tissues. l Results based on the TCGA and GEPIA databases showed the expression level of IGF2BP3 in HNSC. m IHC staining of IGF2BP3 in LSCC tumour specimens. n Kaplan-Meier survival analysis showed that the expression level of IGF2BP3 in LSCC patients was significantly related to the OS. o and p Data from the TCGA database showed that the expression level of IGF2BP3 in HNSC patients was significantly related to OS and RFS. q Correlation analysis was conducted on the expression levels of TMBIM6 and IGF2BP3 in 34 cases of LSCC tissues. r Correlation analysis was performed on the expression levels of TMBIM6 and IGF2BP3 in the TCGA database
Fig 2: RBM15 regulates LSCC growth and apoptosis in mice. a Inhibition of RBM15 impaired the growth of xenografted tumors. b Overexpression of RBM15 stimulated the growth of xenografted tumors. c and d Representative images of TUNEL staining quantification of RBM15 positive staining in xenografted tumours after transfection with shCtrl and/or shRBM15 and/or RBM15 vectors
Fig 3: Quantification and concentration-dependence analysis of the RNA methyltransferase expression levels after CTX treatment in vitro. (A) The mRNA expression levels of METTL3, METTL14, and WTAP in mouse ovary were measured after treatment with CTX (40, 80, and 120 mg/kg). (B) The mRNA expression levels of ZC3H13, KIAA1429, and RBM15 in mouse ovary were measured after treatment with CTX (40, 80, and 120 mg/kg). (C) The protein expression levels of METTL3, METTL14, and WTAP in mouse ovary were measured after treatment with CTX (40, 80, and 120 mg/kg). (D) The protein expression levels of ZC3H13, KIAA1429 and RBM15 in mouse ovary were measured after treatment with CTX (40, 80, and 120 mg/kg). All experiments were performed three times. The error bars indicate SD; **p < 0.01; ***p < 0.001 (compared with the control group); ##p < 0.01 (compared with the 80 mg/kg CTX concentration group).
Fig 4: RBM15 accelerates LSCC malignant progression by upregulating TMBIM6. a Western blot was performed to investigate the expression of RBM15 and TMBIM6 in AMC-HN-8 cells after transfection with shRBM15–2 and/or TMBIM6. b Western blot was conducted to examine the expression of RBM15 and TMBIM6 proteins in TU-212 cells after transfection with shTMBIM6–2 and/or RBM15. c Transwell assay was performed to measure the cell invasion ability of AMC-HN-8 cells after transfection with shRBM15–2 and/or TMBIM6. Hoechst staining assay was performed to measure cell apoptosis of AMC-HN-8 cells after transfection with shRBM15–2 and/or TMBIM6. d Transwell assay was performed to measure the cell invasion ability of TU-212 cells after transfection with shTMBIM6–2 and/or RBM15. Hoechst staining assay was performed to measure the cell apoptosis of TU-212 cells after transfection with shTMBIM6–2 and/or RBM15. e Knockdown of RBM15 inhibited TMBIM6-induced AMC-HN-8 cells subcutaneously tumor growth in nude mice. f Knockdown of TMBIM6 inhibited RBM15-induced TU-212 cells subcutaneously tumor growth in nude mice
Fig 5: Western blot results suggested that after both RBM15 and IGF2BP3 were knocked down in AMC-HN-8 cells, the TMBIM6 protein level was significantly reduced, and the TMBIM6 protein expression increased after both RBM15 and IGF2BP3 were overexpressed, but was restored after the knockdown of RBM15 or IGF2BP3
Supplier Page from Abcam for Anti-Rbm15/OTT antibody