Human X chromosome exome sequencing identifies BCORL1 as contributor to spermatogenesis

Background Infertility affects approximately 15% of couples worldwide with male infertility being responsible for approximately 50% of cases. Although accumulating evidence demonstrates the critical role of the X chromosome in spermatogenesis during the last few decades, the expression patterns and potential impact of the X chromosome, together with X linked genes, on male infertility are less well understood. Methods We performed X chromosome exome sequencing followed by a two-stage independent population validation in 1333 non-obstructive azoospermia cases and 1141 healthy controls to identify variant classes with high likelihood of pathogenicity. To explore the functions of these candidate genes in spermatogenesis, we first knocked down these candidate genes individually in mouse spermatogonial stem cells (SSCs) using short interfering RNA oligonucleotides and then generated candidate genes knockout mice by CRISPR-Cas9 system. Results Four low-frequency variants were identified in four genes ( BCORL1 , MAP7D3 , ARMCX4 and H2BFWT ) associated with male infertility. Functional studies of the mouse SSCs revealed that knocking down Bcorl1 or Mtap7d3 could inhibit SSCs self-renewal and knocking down Armcx4 could repress SSCs differentiation in vitro. Using CRISPR-Cas9 system, Bcorl1 and Mtap7d3 knockout mice were generated. Excitingly, Bcorl1 knockout mice were infertile with impaired spermatogenesis. Moreover, Bcorl1 knockout mice exhibited impaired sperm motility and sperm cells displayed abnormal mitochondrial structure. Conclusion Our data indicate that the X-linked genes are associated with male infertility and involved in regulating SSCs, which provides a new insight into the role of X-linked genes in spermatogenesis.