Mitochondria-anchored protein complexes in piRNA biogenesis and function

项目总结 Pachytene piRNAs是哺乳动物特有的一组小调节RNA， 调节精子发生和生育。在小鼠中，粗线虫piRNAs是由 通过被piRNA加工机械切割的长piRNA前体和 被加载到两个细胞质PIWI蛋白MIWI和MILI上。然而，这一机制通过 哪些PIWI蛋白被招募到piRNA加工机制中参与piRNA 生物发生仍然难以捉摸。在这里，我们提供了初步数据，揭示了一种新的基因 MIWI和MILI的分离，以差异进入pIRNA加工机械。 这种分离是由线粒体锚定蛋白复合体介导的 专门与MIWI互动，但不与MILI互动。我们假设不同的线粒体- 锚定复合体不同地引导不同的PIWI蛋白进入piRNA途径 在粗线期的piRNA加工过程中。为了验证这一假设，我们将使用生物化学 和小鼠遗传学方法：1)了解特定基因的功能重要性 基于线粒体的蛋白质相互作用在引导piRNA生物发生和生殖细胞中的作用 功能；2)确定PIWI蛋白募集和下游的偶联机制 3)探索一种特定的MILI招募机制，以进入 PiRNA途径。这些研究将为组织原则提供新的见解 PIRNA加工机制及对其潜在机制的理解 粗线虫piRNA的生物发生及其与正常精子发生和生育的关系。

PROJECT SUMMARY Pachytene piRNAs are a population of small regulatory RNAs unique to mammals that regulates spermatogenesis and fertility. In mice, pachytene piRNAs are generated from long piRNA precursors through cleavage by the piRNA processing machinery and loaded onto two cytoplasmic PIWI proteins MIWI and MILI. However, the mechanism by which PIWI proteins are recruited to piRNA processing machinery to participate in piRNA biogenesis remains elusive. Here we provide preliminary data that reveal a novel genetic separation of MIWI and MILI to differentially enter into the piRNA processing machinery. This segregation is mediated by a mitochondria-anchored protein complex that specifically interacts with MIWI but not MILI. We hypothesize that distinct mitochondria- anchored complexes differentially direct different PIWI proteins into the piRNA pathway during pachytene piRNA processing. To test this hypothesis, we will use biochemical and mouse genetic approaches to: 1) understand the functional importance of specific mitochondria-based protein interactions in directing piRNA biogenesis and germ cell function; 2) define the coupling mechanism for PIWI protein recruitment and downstream piRNA processing; 3) explore a specific MILI recruiting mechanism to enter into the piRNA pathway. These studies will provide novel insight into the organizing principle of the piRNA processing machinery and the understanding of mechanism underlying pachytene piRNA biogenesis and its relevance to normal spermatogenesis and fertility.