The N-methyladenosine demethylase ALKBH5 negatively regulates the osteogenic differentiation of mesenchymal stem cells through PRMT6.

N-methyladenosine (mA) modification is widespread in messenger RNAs and increasing evidence suggests the crucial roles of mA in cell differentiation and tissue development. However, whether mA modulates the osteogenic differentiation of mesenchymal stem cells (MSCs) has not been fully elucidated. Here we show that conditional knockout of the demethylase Alkbh5 in bone marrow MSCs strengthened bone mass in mice. Loss- and gain-of-function studies demonstrated that ALKBH5 negatively regulates the osteogenic differentiation of MSCs in vitro. At a mechanistic level, meRIP-seq and RNA-seq in MSCs following knockdown of ALKBH5 revealed changes in transcripts of PRMT6 containing consensus mA motifs required for demethylation by ALKBH5. Furthermore, we found that ALKBH5 accelerates the degradation rate of PRMT6 mRNA in an mA-dependent manner, and that the ALKBH5-PRMT6 axis regulates the osteogenesis of MSCs, mainly through activation of the PI3K/AKT pathway. Thus, our work reveals a different facet of the novel ALKBH5-PRMT6 axis that modulates the osteogenic differentiation of MSCs, which can serve as a target to improve the clinical use of MSCs.