Research Paper Volume 13, Issue 19 pp 23361—23375

piRNA-36741 regulates BMP2-mediated osteoblast differentiation via METTL3 controlled m6A modification

piR-36741 suppressed METTL3-mediated BMP2 m6A methylation to promote BMP2 expression. (A) The m6A level of BMP2 mRNA in BMSCs treated with antagomir-piR-36741 or antagomir-NC was detected by using MeRIP-qPCR. (B, C) The mRNA and protein levels of BMP2 in BMSCs treated with mimic-piR-36741, antagomir-piR-36741 or respective controls were measured. (D) RIP assay was used to analyze the level of BMP2 mRNA binding to METTL3 using anti-IgG or anti-METTL3. BMSCs were transfected with METTL3 overexpression vector alone or together with mimic-piR-36741, and (E) the global m6A level and (F) BMP2 m6A level were detected by using MeRIP-qPCR. (G) RIP assay was performed to confirm the binding of YTHDF2 with BMP2 mRNA. (H) The mRNA level of BMP2 was determined in BMSCs treated with pcDNA-YTHDF2, YTHDF2 siRNA or respective controls. N = 5 in each group. *P

Figure 5. piR-36741 suppressed METTL3-mediated BMP2 m6A methylation to promote BMP2 expression. (A) The m6A level of BMP2 mRNA in BMSCs treated with antagomir-piR-36741 or antagomir-NC was detected by using MeRIP-qPCR. (B, C) The mRNA and protein levels of BMP2 in BMSCs treated with mimic-piR-36741, antagomir-piR-36741 or respective controls were measured. (D) RIP assay was used to analyze the level of BMP2 mRNA binding to METTL3 using anti-IgG or anti-METTL3. BMSCs were transfected with METTL3 overexpression vector alone or together with mimic-piR-36741, and (E) the global m6A level and (F) BMP2 m6A level were detected by using MeRIP-qPCR. (G) RIP assay was performed to confirm the binding of YTHDF2 with BMP2 mRNA. (H) The mRNA level of BMP2 was determined in BMSCs treated with pcDNA-YTHDF2, YTHDF2 siRNA or respective controls. N = 5 in each group. *P < 0.05. Each test was independently repeated at least three times.