Decoding Stem Cell Chromatin Using Drosophila

描述(由申请人提供)：对干细胞的研究有望对包括癌症和器官衰竭在内的广泛的人类疾病产生重要的见解。此外，干细胞的再生潜力使其成为治疗应用的理想候选者。新出现的证据表明，染色质组织在建立干细胞身份和维持干细胞方面发挥着关键作用。我们的长期目标是识别和描述干细胞中负责基因调控的分子。作为一个模型，我们研究了成年果蝇的两种不同类型的干细胞，卵巢的生殖系干细胞和成年中肠的肠道干细胞。我们最初的努力集中在描述果蝇的瘦弱(Scny)基因。SCNY首先引起了我们的注意，因为它是一种保守的泛素特异性蛋白水解酶，它使组蛋白H2B去泛素化，并在基因沉默中发挥作用。我们的遗传分析表明，SCNY的破坏会导致多个组织中的干细胞丢失。在这项提案中，我们建立在这些初步发现的基础上。在目标1中，我们试图使用一些遗传方法来定义哪些细胞需要SCNY功能。在目标2中，我们从遗传学和生物化学的角度定义了SCNY蛋白的结构域。在目标3中，我们测试了SCNY是否直接调节候选基因的表达。最后，在目标4中，我们鉴定了PAF1和两个组蛋白H_2B泛素连接酶的功能，并测试了它们是否与SCNY相互作用。这项工作将有助于我们对SCNY功能的理解。考虑到组蛋白修饰层次的保守性，在这一提议下揭示的原理将为理解哺乳动物干细胞在正常和疾病状态下的内在程序提供基础。公共卫生相关性：干细胞的研究和临床应用将对医学科学产生深远的影响。这项建议侧重于以果蝇为模型系统来理解干细胞中控制基因表达的独特机制。我们相信，这项工作将揭示干细胞生物学的基本原理，从而加快这些细胞在人类疾病治疗中的使用。

DESCRIPTION (provided by applicant): The study of stem cells promises to yield important insights into a broad range of human diseases including cancer and organ failure. Furthermore, the regenerative potential of stem cells makes them ideal candidates for therapeutic applications. Emerging evidence suggests chromatin organization plays a pivotal role in establishing stem cell identity and maintenance. Our long-term goal is to identify and characterize the molecules responsible for gene regulation in stem cells. As a model, we study two different types of stem cells in adult Drosophila, the germline stem cells of the ovary and the intestinal stem cells of the adult midgut. We have focused our initial efforts on characterizing the Drosophila scrawny (scny) gene. SCNY first captured our attention because it is a conserved ubiquitin specific protease that deubiquitylates histone H2B and functions in gene silencing. Our genetic analysis showed that disruption of scny results in stem cell loss in multiple tissues. In this proposal we build upon these preliminary findings. In Aim 1, we seek to define what cells require scny function using a number of genetic approaches. In Aim 2, we genetically and biochemically define the domain structure of the SCNY protein. In Aim 3, we test if SCNY directly regulates the expression of candidate genes. Finally, in Aim 4 we characterize the function of PAF1 and two histone H2B ubiquitin ligases and test whether they interact with scny. This work will illuminate our understanding of scny function. Given the conservation of the histone modification hierarchy, principles revealed under this proposal will provide a basis for understanding the intrinsic programs of mammalian stem cells in both normal and diseased states. PUBLIC HEALTH RELEVANCE: The study and clinical use of stem cells is poised to have a profound impact on medical science. This proposal focuses on understanding the unique mechanisms that control gene expression in stem cells using Drosophila as a model system. We believe this work will reveal fundamental principles in stem cell biology and thus will accelerate the use of these cells in the treatment of human diseases.