The Cutoff protein regulates piRNA cluster expression and piRNA production in the Drosophila germline. Drosophila melanogaster

In a broad range of organisms, Piwi-interacting RNAs (piRNAs) have emerged as core components of a surveillance system that protects the genome by silencing transposable and repetitive elements. A vast proportion of piRNAs is produced from discrete genomic loci, termed piRNA clusters. The molecular mechanisms and the factors that govern the expression of these loci are largely unknown. We have preciously shown the Cutoff (Cuff), a protein with similarity to yeast Rai1, is a component of the piRNA pathway. In order to understand the function of the Cuff protein in piRNA production, we produced small RNA libraries from cn, bw (wt) and cuffwm25 mutant ovaties. The analysis of these libraries revealed that approximately 80% of the total piRNA population is depleted in the absence of a functional Cuff protein. We also determined that Cuff is mostly a nuclear protein and is enriched at the level of certain piRNA clusters. Our results point to a role for Cuff in the transcriptional regulation of piRNA generating loci and in the production of the proper piRNA complement during Drosophila oogenesis. Overall design: piRNA profiling in ovaries from cn, bw and cuffwm25 mutant ovaries

在广泛的生物类群中，Piwi互作RNA（Piwi-interacting RNAs，piRNAs）已被证实为一类基因组监控系统的核心组分，该系统通过沉默转座因子与重复序列来维护基因组稳定性。绝大多数piRNAs源自特定的基因组位点，这类位点被称为piRNA簇（piRNA clusters）。目前，调控这些位点表达的分子机制与相关因子仍在很大程度上未被阐明。此前我们已证实，与酵母Rai1蛋白具有序列同源性的Cutoff（简称Cuff），是piRNA通路的组分之一。为解析Cuff蛋白在piRNA生成过程中的功能，我们分别以野生型cn、bw品系以及cuffwm25突变型果蝇的卵巢为材料，构建了小分子RNA文库。对这些文库的分析结果显示，当缺乏功能性Cuff蛋白时，总piRNA群体中约80%的组分出现显著缺失。我们还证实，Cuff主要定位于细胞核，且在特定piRNA簇区域呈现富集状态。本研究结果表明，Cuff在果蝇卵子发生过程中，参与调控piRNA生成位点的转录，并参与合成完整的piRNA组分。实验整体设计：以cn、bw野生型品系及cuffwm25突变型果蝇的卵巢为材料，开展piRNA表达谱分析。