SRA

Homologous pairing and synapsis are essential in meiotic prophase I during spermatogenesis. However, the underlying mechanisms of how alternative splicing (AS) functions in homologous pairing and synapsis remain largely unclear. We reveal that SRSF1 is essential for gene expression and AS in homologous pairing and synapsis. Conditional knockout (cKO) of Srsf1 in mouse germ cells impaired homologous pairing and synapsis, leading to non-obstructive azoospermia (NOA). SRSF1 was required for initial homology recognition, telomere-led chromosome movement, and synaptonemal complex (SC) assembly. Moreover, SRSF1 interacted with TRA2B and U2AF2, directly binding and regulating the expression of Dmc1, Sycp1, Sun1, and Majin via AS to implement homologous pairing and synapsis during the meiotic prophase I program. Altogether, our data reveal the critical role of an SRSF1-mediated post-transcriptional regulatory mechanism in homologous pairing and synapsis during meiotic prophase I, providing a framework for elucidating the molecular mechanisms underlying the post-transcriptional network of male meiosis. Overall design: Total cells were isolated from adult WT C57BL/6N mouse testes, and then the cells were cross-linked by ultraviolet light (254 nm) to maintain the covalent binding of RBPs to their cognate RNA. Subsequently, SRSF1 and cross-linked RNAs were immunoprecipitated with an anti-SRSF1 antibody and digested with micrococcal nuclease (EN0181, Thermo Fisher Scientific). An IR800-biotin adapter was ligated to the 3' ends of the RNA fragments. Then, the SRSF1/RNA complexes were separated by SDS?PAGE and transferred to a nitrocellulose membrane (HATF00010, Millipore). These RNA and protein complexes from approximately 47 to 62 kDa were extracted from the nitrocellulose membrane, after which proteinase K (9034, Takara) digestion was performed. RNA was isolated with saturated phenol (AM9712, Ambion), ligated with adaptors, and converted to cDNA with a SuperScript III First-Strand Kit (18080-051, Invitrogen). The cDNA was amplified by PCR to prepare the corresponding libraries and then sequenced on illumina NovaSeq 6000.