YTHDF2 is essential for spermatogenesis and fertility through mediating a wave of transcript transition during spermatogonial differentiation

The dynamic and reversible regulation roles of m6A modification, and the characterization of m6A readers have provided new insights into spermatogenesis at post-transcriptional level. YTHDF2 has been reported to recognize and mediate the m6A-containing transcripts decay during the mouse oocyte mature, embryonic stem cell differentiation, neural development, and zebrafish maternal-to-zygotic transition. However, the roles of YTHDF2 in mammalian spermatogenesis are uncertain. Here, we generated germ cell-specific Ythdf2 mutants (Ythdf2-vKO) at a C57BL/6J background, and demonstrated that YTHDF2 was essential for mouse spermatogenesis and fertility. Ythdf2-vKO provided oligoasthenoteratozoospermia (OAT) phenotype with increased apoptosis in germ cells. High-throughput RNA-seq of the testis tissue showed the failure of the degradation of a wave of YTHDF2 target mRNA. Interestingly, RNA-seq analysis combined with our previous single-cell transcriptomics data of mouse spermatogenesis pointed out the failure of a wave of transcript transition during the spermatogenesis of Ythdf2-vKO, which was confirmed by gene expression analysis of diplotene spermatocytes and round spermatids obtained through fluorescence-activated cell sorting using qPCR. Our study demonstrates the fundamental role of YTHDF2 during mouse spermatogenesis and provides a potential candidate for the diagnosis of male infertility with OAT syndrome. Author summaryMale infertility is becoming a worldwide health problem. Male gamete is generated through spermatogenesis, which is a complicated developmental process with dynamic transcriptome changes. RNA m6A modification has been reported as the most prominently internal mRNA modification, which control the tune of gene expression through mRNA splicing, export, translation and decay. RNA m6A modification is catalyzed by "writers", and could be removed by "erasers". The m6A modification enzymes are reported to play important roles during spermatogenesis. Given that the biological function of m6A modification are mediated through its "readers", its readers might be proposed to be involved in the regulation of spermatogenesis. YTHDF2, as a reader of m6A modification, has been reported to mediate the m6A-containing transcripts decay. To explore the roles of YTHDF2 in spermatogenesis, we used germ cell-specific mutation strategy to knock out the mouse Ythdf2. The mutants provided oligoasthenoteratozoospermia phenotype. Our study demonstrate that YTHDF2 is essential for mouse spermatogenesis. YTHDF2 could be a potential candidate for the diagnosis of oligoasthenoteratozoospermia syndrome.