WTAP-mediated m6A modification modulates bone marrow mesenchymal stem cells differentiation potential and osteoporosis

Abstract

An imbalance in the differentiation potential of bone marrow mesenchymal stem cells (BMSCs) is an important pathogenic mechanism underlying osteoporosis (OP). N6-methyladenosine (m6A) is the most common post-transcriptional modification in eukaryotic cells. The role of the Wilms’ tumor 1-associated protein (WTAP), a member of the m6A functional protein family, in regulating BMSCs differentiation remains unknown. We used patient-derived and mouse model-derived samples, qRT-PCR, western blot assays, ALP activity assay, ALP, and Alizarin Red staining to determine the changes in mRNA and protein levels of genes and proteins associated with BMSCs differentiation. Histological analysis and micro-CT were used to evaluate developmental changes in the bone. The results determined that WTAP promoted osteogenic differentiation and inhibited adipogenic differentiation of BMSCs. We used co-immunoprecipitation (co-IP), RNA immunoprecipitation (RIP), methylated RNA immunoprecipitation (MeRIP), RNA pulldown, and dual-luciferase assay to explore the direct mechanism. Mechanistically, the expression of WTAP increased during osteogenic differentiation and significantly promoted pri-miR-181a and pri-miR-181c methylation, which was recognized by YTHDC1, and increased the maturation to miR-181a and miR-181c. MiR-181a and miR-181c inhibited the mRNA expression of SFRP1, promoting the osteogenic differentiation of BMSCs. Our results demonstrated that the WTAP/YTHDC1/miR-181a and miR-181c/SFRP1 axis regulated the differentiation fate of BMSCs, suggesting that it might be a potential therapeutic target for osteoporosis.

N6-Methyladenosine in Cell-Fate Determination of BMSCs: From Mechanism to Applications

“…In addition to abovementioned miR-320, further studies uncovered that METTL3 can methylate pri-miR-21 and facilitate the maturation of miR-21 [ 55 ], which potentiates the osteogenesis of BMSCs by activating the Smad1/5/8-RUNX2 pathway [ 56 ] and lowering the amount of hypoxia-inducible factor-1α (HIF-1α) [ 57 ]. WTAP, another critical component of m 6 A writers, also acts as a promoter of osteogenesis by encouraging mature miR-29b-3p [ 58 ] and miR-181a/c [ 59 ]. Similar phenomenon was observed between METTL14 and miR-873 [ 60 ].…”

Recent advances of m6A methylation in skeletal system disease

“…And whether BMSCs differentiate into osteogenic cells or adipocytes is linked to the pathogenesis of OP. Recent studies have revealed several mechanisms of m6A modifications to promoted osteogenic differentiation and inhibited adipogenic differentiation of BMSCs [ 137 , 141 , 147 ]. Growing evidences have shown the relationship between m6A modification and BMSC differentiation.…”

Roles of RNA Methylations in Cancer Progression, Autophagy, and Anticancer Drug Resistance

“…m6A modification is reversibly modulated through the installation of methyl transferases (writers), the removal of a methyl group by demethylases (erasers), and the recognition of m6A binding proteins (readers) [ 22 , 23 ]. m6A writers and associated proteins (m6A writer complex) include Vir-like m6A methyl transferase-associated protein (VIRMA) [ 24 ], methyl transferase-like protein 3 (METTL3) [ 25 ], METTL4 [ 26 ], MTTL5 [ 27 ], METTL14 [ 28 ], METTL 16 [ 29 ], Wilms’ tumor 1 associated protein (WTAP) [ 30 , 31 ], RNA-binding motif protein 15/15B (RBM15/15B) [ 32 ], casitas B-lineage lymphoma-transforming sequence-like protein 1 (CBLL1) [ 33 ], zinc finger CCCH-type containing 13 (ZC3H13), and zinc finger CCHC-type containing 4 (ZCCHC4) [ 5 , 34 , 35 , 36 , 37 , 38 ]. RBM15/15B, VIRMA, CBLL1, ZCCHC4, and ZC3H13 act as associated proteins of m6A writers.…”