Dey Souvik, Nofal Wesam, Brothag Cameron, Kabi Mustfa, Khamamkar Aditi, Choudhari Neha, Vijayaraghavan Srinivasan
Reproduction. 2025 Jan 21;169(2). doi: 10.1530/REP-24-0256. Print 2025 Feb 1.
Protein phosphatase 1 catalytic subunit gamma isoform 2 (PP1γ2) is a unique phosphatase expressed only in mammalian testes and sperm cells. The PP1γ2 isoform is indispensable for sperm motility and fertility and cannot be replaced by the PP1γ1 isoform for these functions.
The serine-threonine phosphatase has four paralogs - PP1α, PP1β, PP1γ1 and PP1γ2 - encoded by three genes, Ppp1ca, Ppp1cb and Ppp1cc. Protein phosphatase PP1γ2, one of two isoforms of the gene Ppp1cc, is expressed in spermatogenic cells in the testes and sperm, while PP1γ1 is found in somatic cells. The two PP1γ isoforms, formed by alternate splicing that occurs only in mammals, are identical except at their C-termini. Global or testis-specific knockout of Ppp1cc in mice results in male infertility due to disrupted spermiation and mid-to-late spermiogenesis. Transgenic expression of PP1γ2, driven by a testis-specific promoter in differentiating spermatogenic cells, rescues spermatogenesis and fertility in the Ppp1cc-null mice. Why PP1γ2 is essential and present only in mammalian sperm is a mystery. We have generated a knock-in mouse where the Ppp1cc gene is edited to express only PP1γ1. Spermatogenesis was normal in knock-in mice. Testis-expressed PP1γ1 in the knock-in mice and PP1γ2 in the wild-type mice were incorporated in equal amounts into sperm. Sperm bearing PP1γ1 have reduced flagellar beat amplitude and motility, and male mice were severely sub-fertile. Although in the wild-type mice, PP1γ2 is present in both the head and tail, in the knock-in mice, PP1γ1 is absent in sperm heads, leading to an altered intra-sperm protein phosphatase landscape. Phosphoproteomic analysis of sperm proteins suggested a plausible molecular basis for compromised PP1γ1 functions: it identified GSK3α, a known substrate of PP1, to be dysregulated in knock-in sperm. This study provides a preliminary explanation for the isoform-specific requirement of PP1γ2 for male fertility.
蛋白磷酸酶1催化亚基γ2型（PP1γ2）是一种仅在哺乳动物睾丸和精子细胞中表达的独特磷酸酶。PP1γ2亚型对于精子活力和生育能力不可或缺，并且PP1γ1亚型无法替代其在这些功能中的作用。
丝氨酸 - 苏氨酸磷酸酶有四个旁系同源物——PP1α、PP1β、PP1γ1和PP1γ2——由三个基因Ppp1ca、Ppp1cb和Ppp1cc编码。蛋白磷酸酶PP1γ2是基因Ppp1cc的两种亚型之一，在睾丸生精细胞和精子中表达，而PP1γ1存在于体细胞中。这两种PP1γ亚型由仅在哺乳动物中发生的可变剪接形成，除了它们的C末端外其余相同。在小鼠中全局或睾丸特异性敲除Ppp1cc会导致雄性不育，原因是精子释放和精子发生中后期过程受到破坏。在分化的生精细胞中由睾丸特异性启动子驱动的PP1γ2转基因表达可挽救Ppp1cc基因敲除小鼠的精子发生和生育能力。为什么PP1γ2是必需的且仅存在于哺乳动物精子中仍是一个谜。我们构建了一种基因敲入小鼠，其中Ppp1cc基因被编辑为仅表达PP1γ1。基因敲入小鼠的精子发生正常。基因敲入小鼠睾丸中表达的PP1γ1和野生型小鼠中的PP1γ2以等量掺入精子中。携带PP1γ1的精子鞭毛摆动幅度和活力降低，雄性小鼠严重亚生育。虽然在野生型小鼠中，PP1γ2存在于头部和尾部，但在基因敲入小鼠中，精子头部不存在PP1γ1，导致精子内蛋白磷酸酶格局改变。对精子蛋白的磷酸化蛋白质组分析为受损的PP1γ1功能提出了一个合理的分子基础：它确定PP1的已知底物GSK3α在基因敲入精子中失调。这项研究为PP1γ2对雄性生育能力的亚型特异性需求提供了初步解释。
