ALKBH5 exerts an m6A-independent function to control the spindle assembly in oocytes through KIF22

ALKBH5 is a key determinant of oocyte quality by regulating N 6 -methyladenosine (m 6 A) to maintain maternal RNA stability in oocytes. However, the role of ALKBH5 in oocytes other than m 6 A has not been reported. Here, we document that ALKBH5 is expressed and localized to spindle during mouse oocyte meiotic maturation. The abnormal spindle in Alkbh5 -/- oocytes can be both partially restored by the ectopic expression of exogenous wild-type Alkbh5 and enzymatically dead Alkbh5-H205A . Moreover, we find that ALKBH5 binds to the mitotic kinesin motor protein KIF22, a key spindle assembly regulator. Mechanistically, ALKBH5 knockout led to proteasomal degradation of KIF22 protein (without altering its mRNA level) via the E3 ubiquitin ligase PJA1; ALKBH5 and KIF22 competed for binding to PJA1 at the shared HIS-10 site, and loss of ALKBH5 promoted PJA1-mediated KIF22 ubiquitination and degradation. Collectively, our findings uncover a novel nonenzymatic mechanism by which ALKBH5 modulates KIF22 protein stability through competing for PJA1 binding, thereby regulating spindle assembly and meiotic maturation in mouse oocytes. This study expands the biological functions of ALKBH5 and provides new insights into the molecular regulation of oocyte meiosis beyond its conventional role in m 6 A.