piRNA-independent PIWI function in spermatogenesis and male fertility.
piRNA-independent PIWI function in spermatogenesis and male fertility.
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不依赖于 piRNA 的 PIWI 在精子发生和男性生育能力中发挥作用。
DOI：
10.1093/biolre/iox055
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发表时间：
2017
影响因子：
3.6
通讯作者：
YanEditor-In-ChiefBiologyOfReproduction,Wei
中科院分区：
文献类型：
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作者：
YanEditor-In-ChiefBiologyOfReproduction,Wei
PIWI（果蝇中P元素诱导的懦弱睾丸）家族蛋白在进化上是保守的，并且在迄今为止研究的大多数动物物种中对雄性生育力具有重要作用[1]。所有三种鼠PIWI直系同源物（MIWI、MILI和MIWI 2）都是精子发生和男性生育力所需的，但四种人PIWI蛋白（HIWI、HILI、HIWI 2和PIWI 3）在人男性生育力中的相关性迄今尚未得到证实。刘莫方领导的研究小组最近在《细胞》杂志上发表了一篇论文[2]，报告称HIWI N末端D盒结构域的点突变通过阻碍精子发生晚期组蛋白与鱼精蛋白的交换而导致男性不育。由于这些点突变仅改变D-box区域中的一个或两个氨基酸，而已知对HIWI的皮尔纳功能至关重要的其他结构域（包括PAZ、MID和PIWI）保持完整，因此突变体HIWI蛋白的piRNA相关功能是正常的并不奇怪，这表明D-box结构域具有独立于已知皮尔纳功能的新作用。事实上，携带一个突变等位基因的敲入小鼠，其D盒中的点突变与人类相同，或过表达相同的突变MIWI蛋白，可以重现人类表型，即晚期精子发生和男性不育的损害，证明这些点突变和男性不育表型之间的因果关系。携带一个无效的Miwi等位基因（Miwi+/-）的雄性小鼠是可育的，而那些携带一个突变的Miwi等位基因的雄性小鼠是不育的，这些突变的Miwi等位基因含有点突变并在保守的D-box结构域中产生突变的MIWI蛋白。这一结果表明，野生型MIWI产生水平的降低（例如在Miwi或HIWI杂合子个体中）不会破坏精子发生，但具有D盒突变的突变MIWI/HIWI的产生会破坏精子发生，表明突变MIWI/HIWI蛋白在精子发生后期的显性负效应。这些D-box点突变是如何导致小鼠和人类的男性不育的？鉴于D-box结构域在APC/C泛素E3连接酶的底物之间共享，并且MIWI在延长精子细胞中被ACP/C降解[3]，D-box中的突变将阻碍泛素化介导的MIWI蛋白周转，导致突变MIWI在精子细胞中积累。事实上，MIWI D盒突变小鼠的MIWI睾丸水平更高。精子细胞中MIWI/HIWI水平过高和持续存在的后果是什么？通过生化分析，作者进一步发现MIWI与RNF 8相互作用，MIWI与RNF 8之间的结合通过将RNF 8隔离在细胞质中来抑制RNF 8泛素E3连接酶活性，从而阻止其进入细胞核并启动组蛋白降解程序[4]。这一观点进一步得到了实验的支持，该实验显示使用RNF 8 N-末端多肽阻断MIWI-RNF 8相互作用可以挽救D-盒突变小鼠中的缺陷。总之，作者令人信服地证明了MIWI/HIWI的D盒点突变导致MIWI/HIWI蛋白在延长精子细胞中的长期存在，这反过来又抑制了RNF 8介导的组蛋白降解，从而导致组蛋白与鱼精蛋白交换受损，从而导致男性不育。这一发现是新颖的，因为它首次揭示了HIWI蛋白的D-box结构域中的点突变通过阻碍组蛋白与鱼精蛋白的交换而不是通过影响皮尔纳的产生和功能来导致人类精子产生受损和男性不育。MIWI是数千种粗线期piRNA正常产生和功能所必需的，这些粗线期piRNA是由蛋白质组成的。
PIWI (P element-induced wimpy testis in Drosophila) family proteins are evolutionarily conserved and have an essential role in male fertility in most of the animal species studied so far [1]. All three murine PIWI orthologs (MIWI, MILI, and MIWI2) are required for spermatogenesis and male fertility, but the relevance of the four human PIWI proteins (HIWI, HILI, HIWI2, and PIWIL3) in human male fertility has not been demonstrated until now. A group led by Mofang Liu recently published a paper in Cell [2], reporting that point mutations in the D-box domain of the N-terminus of HIWI cause male infertility by impeding the histone-to-protamine exchange during late spermiogenesis. Since these point mutations only alter one or two amino acids in the D-box region while other domains known to be critical for HIWI’s piRNA functions, including PAZ, MID, and PIWI, remain intact, it is not surprising that the piRNA-related functions of the mutant HIWI protein are normal, suggesting that the D-box domain has a novel role independent of the known piRNA functions. Indeed, knockin mice carrying one mutant allele with the same point mutations in the D-box as those in humans, or overexpressing the same mutant MIWI proteins, can recapitulate the human phenotype, ie impairment in late spermiogenesis and male infertility, demonstrating the cause–effect relationship between these point mutations and the male infertility phenotype. The male mice carrying one null Miwi allele (Miwi+/−) are fertile, whereas those with one mutant Miwi allele containing point mutations and producing mutant MIWI protein in the conserved D-box domain are sterile. This result suggests that reduced levels of wild-type MIWI production (eg in Miwi or HIWI heterozygous individuals) do not disrupt spermiogenesis, but the production of the mutant MIWI/HIWI with D-box mutations does, indicating a dominantnegative effect of the mutant MIWI/HIWI proteins in late spermiogenesis. How do these D-box point mutations cause male infertility in both mice and humans? Given that the D-box domain is shared among the substrates for APC/C ubiquitin E3 ligase and MIWI is degraded by ACP/C in elongating spermatids [3], mutations in D-box would impeded ubiquitination-mediated MIWI protein turnover, leading to an accumulation of mutant MIWI in spermatids. Indeed, testicular levels of MIWI are higher in MIWI D-box mutant mice. What are the consequences of excessive levels and persistent presence of MIWI/HIWI in spermatids? Through biochemical analyses, the authors further discovered that MIWI interacts with RNF8 and the binding between MIWI and RNF8 inhibits RNF8 ubiquitin E3 ligase activity by sequestering RNF8 in the cytoplasm, thus preventing it from entering the nucleus and starting the histone degradation program [4]. This notion is further supported by the experiments showing that blocking MIWI-RNF8 interactions using a RNF8 N-terminus polypeptide can rescue the defects in the D-box mutant mice. Together, the authors have convincingly demonstrated that D-box point mutations of MIWI/HIWI lead to prolonged existence of MIWI/HIWI protein in elongating spermatids, which, in turn, inhibit the RNF8-mediated histone degradation, thus causing impaired histone-to-protamine exchange and consequently male infertility. The finding is novel because it reveals, for the first time, that point mutations in the D-box domain of HIWI proteins lead to impaired sperm production and male infertility in humans by impeding the histone-to-protamine exchange, rather than by affecting piRNA production and function.MIWI is required for normal production and function of thousands of pachytene piRNAs, which …