Genome evolution and the formation of piRNA clusters in Drosophila

果蝇基因组进化和 piRNA 簇的形成

• 批准号: 458259436
• 负责人: Dr. Daniel Gebert
• 金额: --
• 依托单位: Department of Genetics
• 依托单位国家: 德国
• 项目类别: WBP Fellowship
• 财政年份: 2021
• 资助国家: 德国
• 起止时间: 2020-12-31 至 2023-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/458259436?language=en
• 关键词: Genome; evolution; formation; piRNA; clusters

Almost all life forms find themselves in a constant evolutionary arms race with transposable elements (TEs) in order to protect their genomes against uncontrolled proliferation of these mobile genetic units, as they pose a threat to genome integrity and stability. Since this is particularly critical in the germline where genomic changes are hereditary, organisms have evolved specialised defence systems, such as the metazoan PIWI/piRNA pathway. PIWI proteins are guided by small PIWI-interacting (pi-) RNAs through sequence complementarity to their TE targets and direct post-transcriptional and transcriptional silencing. In virtually all animals, large TE rich genomic loci, called piRNA clusters, produce the vast majority of piRNAs. It is commonly assumed that piRNA clusters play a central role in TE silencing as the main source for TE-targeting piRNAs.In our preliminary work, we studied the evolution of piRNA clusters in the Drosophila genus and found that fly piRNA clusters are exceptionally short-lived and often emerge at genomic loci that are marked by recurrent inversion breakpoints, which are regions that have been proposed to accumulate TE insertions in general. Since this lack of conservation is at odds with a critical function, we have put the model to the test by deleting three of the largest piRNA clusters of Drosophila melanogaster, which alone produce the majority of piRNAs. Surprisingly, this did not affect germ cell viability or lead to reactivation of any TEs, challenging the assumption of their functional relevance. This, however, poses the question why piRNA clusters, which appear to be highly ordered and distinct structures, emerge and how they develop, as they are a ubiquitous and recurrent characteristic of metazoan genomes.In this project, I will examine the influence of inversion-causing DNA fragility on the emergence of piRNA clusters, and the underlying impact of chromosome conformation and packaging, using Hi-C, ATAC-seq and ChIP-seq, across Drosophila species. Here, piRNA clusters will serve as a model study that will be expanded towards the broad-scale impact of genome conformation on genomic structural evolution. Further, I will bioinformatically analyse TE structures of piRNA clusters across species to understand their initial formation and development over time. Finally, since the PIWI system apparently functions independently of piRNA clusters, I will explore how new TEs are recognized without prior existence of complementary piRNAs, by introducing foreign TE copies into fly genomes followed by DNA and small RNA sequencing. Altogether, this project will provide valuable insights into the mutual impact of genomes and transposable elements and a deep view of genomic architecture evolution on an unprecedented scale.

几乎所有的生命形式都发现自己处于与转座因子（TE）的持续进化军备竞赛中，以保护其基因组免受这些移动的遗传单位的不受控制的增殖，因为它们对基因组的完整性和稳定性构成威胁。由于这在基因组变化具有遗传性的种系中特别关键，因此生物体已经进化出专门的防御系统，例如后生动物PIWI/皮尔纳途径。PIWI蛋白由小PIWI相互作用（pi-）RNA通过与其TE靶标的序列互补性以及直接转录后和转录沉默来引导。在几乎所有动物中，大的富含TE的基因组基因座（称为皮尔纳簇）产生绝大多数piRNA。通常认为皮尔纳簇作为TE靶向piRNA的主要来源在TE沉默中起着核心作用。在我们的初步工作中，我们研究了果蝇属中皮尔纳簇的进化，发现苍蝇皮尔纳簇的寿命非常短，并且经常出现在以反复倒位断点为标志的基因组位点，这是一般被认为积累TE插入的区域。由于这种保守性的缺乏与关键功能不一致，我们通过删除黑腹果蝇中最大的三个piRNA簇来测试该模型，这些piRNA簇单独产生大多数piRNA。令人惊讶的是，这并不影响生殖细胞的活力或导致任何TE的再活化，挑战了其功能相关性的假设。然而，这提出了一个问题，为什么皮尔纳簇，似乎是高度有序和独特的结构，出现和他们如何发展，因为他们是一个普遍存在的和经常性的后生动物基因组的特征。在这个项目中，我将研究导致倒位的DNA脆弱性对皮尔纳簇出现的影响，以及染色体构象和包装的潜在影响，使用Hi-C，ATAC-seq和ChIP-seq，果蝇物种之间的差异。在这里，皮尔纳簇将作为一个模型研究，将扩展到基因组构象对基因组结构进化的广泛影响。此外，我将从生物信息学角度分析跨物种皮尔纳簇的TE结构，以了解它们随着时间的推移的初始形成和发展。最后，由于PIWI系统显然独立于皮尔纳簇发挥作用，我将探索如何在没有互补piRNA的情况下识别新的TE，方法是将外源TE拷贝引入果蝇基因组，然后进行DNA和小RNA测序。总而言之，这个项目将提供有价值的见解，基因组和转座因子的相互影响和基因组结构的进化在一个前所未有的规模深入了解。