Significant expression levels of transgenic PPP1CC2 in testis and sperm are required to overcome the male infertility phenotype of Ppp1cc null mice.
Significant expression levels of transgenic PPP1CC2 in testis and sperm are required to overcome the male infertility phenotype of Ppp1cc null mice.
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DOI：
10.1371/journal.pone.0047623
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Vijayaraghavan S
中科院分区：
文献类型：
--
作者：
Sinha N;Pilder S;Vijayaraghavan S
PPP1CC2是丝氨酸/苏氨酸蛋白磷酸酶PP1的四种异构体之一，是Ppp1cc基因的哺乳动物特有的剪接变异体，也是唯一几乎完全局限于生精细胞表达的异构体。此外，PPP1CC2是哺乳动物精子中发现的唯一异构体。尽管Ppp1cc的另一个产物PPP1CC1在包括睾丸在内的许多组织中都有表达，但Ppp1cc基因缺失导致的唯一表型是男性不育。为了确定Ppp1cc的哪种产物是男性生育所必需的，我们创建了两个PPP1CC2转基因基因ETG-G2和PTG-G2，其中Ppp1cc2的表达分别由Ppp1cc的内源启动子或睾丸特异性的人Pgk2启动子驱动。我们的结果表明，Ppp1cc结构基因上游2.6kb的基因组区域可以驱动Ppp1cc2的表达，并概括了PPP1CC2在转基因小鼠中的野生型组织特异性。更重要的是，我们发现单独通过任何一个启动子表达PPP1CC2不仅能够恢复正常的精子发生，而且能够恢复Ppp1cc缺失小鼠的生育能力，前提是转基因PPP1CC2在睾丸中的表达至少达到一个较低的阈值水平，相当于Ppp1cc+/−雄性小鼠表达的大约50%。我们的结论是，内源性Ppp1cc启动子在睾丸中正常发挥作用，以维持正常精子发生所需的足够水平的PPP1CC2，并且在完全缺乏PPP1CC1表达的情况下，能够产生能够在体内受精的精子。
PPP1CC2, one of four isoforms of the ser/thr protein phosphatase PP1, is a mammalian-specific splice variant of the Ppp1cc gene, and the only isoform whose expression is confined almost completely to spermatogenic cells. Additionally, PPP1CC2 is the sole isoform found in mammalian spermatozoa. Although PPP1CC1, the other Ppp1cc product, is expressed in many tissues including testis, the only phenotype resulting from deletion of Ppp1cc gene is male infertility. To determine which of the products of Ppp1cc is essential for male fertility, we created two PPP1CC2 transgenes, eTg-G2 and pTg-G2, where Ppp1cc2 expression was driven by the putative endogenous promoter of Ppp1cc or by the testis specific human Pgk2 promoter, respectively. Our results demonstrate that the 2.6-kb genomic region directly upstream of the Ppp1cc structural gene can drive expression of Ppp1cc2, and recapitulate the wild-type tissue specificity of PPP1CC2 in transgenic mice. More importantly, we show that expression of PPP1CC2 alone, via either promoter, is able not only to restore normal spermatogenesis, but the fertility of Ppp1cc null mice as well, provided that transgenic PPP1CC2 expression in testis reaches at least a lower threshold level equivalent to approximately 50% of its expression by a Ppp1cc +/− male. We conclude that the endogenous Ppp1cc promoter normally functions in the testis to maintain a sufficient level of PPP1CC2 expression for normal spermatogenesis to occur, and that production of spermatozoa capable of fertilization in vivo can take place in the complete absence of PPP1CC1 expression.
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