A non-canonical bivalent gene Wfdc15a controls spermatogenic protease and immune homeostasis [P25]

Male infertility can be caused by chromosomal abnormalities, mutations, and epigenetic defects. Epigenetic modifiers pre-program hundreds of spermatogenic genes in spermatogonial stem cells (SSCs) for expression later in spermatids, but it remains mostly unclear whether and how those genes are involved in fertility. Here, we report that Wfdc15a, a WFDC family protease inhibitor pre-programmed by KMT2B, is essential for spermatogenesis. We found that Wfdc15a is a non-canonical bivalent gene carrying both H3K4me3 and facultative H3K9me3 in SSCs but is later activated along with the loss of H3K9me3 and acquisition of H3K27ac during meiosis. We show that WFDC15A deficiency causes defective spermiogenesis at the beginning of spermatid elongation. Notably, depletion of WFDC15A causes substantial disturbance of the testicular protease-antiprotease network and leads to an orchitis-like inflammatory response associated with TNFa expression in round spermatids. Together, our results reveal a unique epigenetic program regulating innate immunity crucial for fertility. To investigate the function of Wfdc15a for spermatogenesis, we performed RNA-seq using whole testis from P25 Wfdc15a mice.男性不育可由染色体异常、基因突变及表观遗传缺陷诱发。表观遗传修饰因子可在精原干细胞（spermatogonial stem cells, SSCs）中对数百个生精相关基因完成预编程，使其在精子细胞发育阶段后续得以表达，但目前学界尚未明确此类基因是否以及如何参与生育调控过程。本研究报道了由KMT2B预编程的WFDC家族蛋白酶抑制剂Wfdc15a，其对精子发生过程至关重要。研究发现，Wfdc15a属于非经典二价基因，在精原干细胞中同时携带H3K4me3与兼性H3K9me3组蛋白修饰；而在减数分裂阶段，该基因会被激活，同时伴随H3K9me3的丢失与H3K27ac修饰的获得。实验结果显示，WFDC15A缺失会导致精子细胞伸长起始阶段的生精功能缺陷。值得关注的是，WFDC15A敲除会显著扰乱睾丸内的蛋白酶-抗蛋白酶网络平衡，并引发以圆形精子细胞中TNFα表达上调为特征的类睾丸炎炎症反应。综上，本研究揭示了一套独特的表观遗传编程机制，其通过调控天然免疫过程对生育能力起到关键支撑作用。为探究Wfdc15a在精子发生中的功能，我们对P25龄Wfdc15a小鼠的全睾丸组织开展了RNA测序（RNA-seq）实验。