RNA binding protein RBM20 regulates turtle temperature-dependent sex determination by repressing the splicing of Wt1...

Accumulating correlative studies suggest a potential role of alternative splicing in vertebrate sex determination; however, the underlying mechanism remains elusive. Herein, we show that RNA binding motif protein 20 (RBM20) exhibits temperature-dependent sexually dimorphic expression in early embryonic gonads of the red-eared slider turtle Trachemys scripta, preceding the onset of gonadal differentiation. Knockdown of Rbm20 at male-producing temperature induces male-to-female sex reversal, whereas Rbm20 overexpression at female-producing temperature triggers male development. RBM20 represses the splicing of Wilms tumor suppressor WT1 at KTS sites, thereby governing the balance of +KTS and -KTS isoforms between different temperatures. In addition, functional analysis reveals that the Wt1 +KTS variant is both necessary and sufficient to drive male development in T. scripta. This study demonstrates that RBM20 activates the male pathway by repressing the Wt1 KTS splicing in T. scripta, thus establishing causality and a direct genetic link between alternative splicing mechanism and vertebrate sex determination.