Novel Role for Piwi/piRNA pathway in developmental robustness

Piwi/piRNA 通路在发育稳健性中的新作用

• 批准号: 8787737
• 负责人: VAMSI GANGARAJU
• 金额: $ 24.9万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-12-20 至 2016-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8787737
• 关键词: Affect; Apoptosis; Aubergine; Binding; Biochemical; Biogenesis; Biological Assay; Biological Models; Cell Nucleus; Cell physiology; Cells; Chromatin; Column Chromatography; Complex; DNA Fragmentation; Development; Developmental Biology; Disease; Drosophila genus; Embryo; Embryonic Development; Enhancers; Environment; Environmental Pollutants; Evolution; Eye Development; Female sterility; Foundations; Functional disorder; Future; Gene Expression Profile; Generations; Genes; Genetic; Genetic Screening; Genotype; Germ Lines; Goals; Human; Human Development; Industrial Waste; Inherited; Learning; Light; Maintenance; Mediating; Molecular; Molecular Chaperones; Mutagenesis; Organism; Pathway interactions; Peptides; Phosphorylation; Play; Population; Process; Proteins; Research; Resources; Role; Stem cells; Stress; Testing; Time; Toxic effect; Untranslated RNA; Variant; Work; abstracting; chromium hexavalent ion; combinatorial; dosage; epigenetic regulation; epigenetic variation; genetic analysis; genome-wide; human disease; in vivo; insight; novel; piRNA; pollutant; prevent; protein crosslink; research study; self-renewal; tool

Project Summary/Abstract: The long term goal of our proposal is to delineate the mechanisms by which Piwi/piRNA pathway normalizes phenotypic variation induced by severe environmental stress and inherent genotype variations. Canalization or developmental robustness provides a framework in which organisms resist displaying phenotypic variation in the face of combined genotype variations and severe environmental stress. Recent research has shown that Hsp90, a molecular chaperone, plays a critical role in canalization. Despite the known role for Hsp90, the lack of a detailed molecular mechanism underlying canalization has provoked great debate for decades. Our lab recently validated the existence of 'canalization' and uncovered a major molecular pathway involving Piwi, Hsp90 and Hop. Using a sensitized eye development assay, we showed that a reduction in the maternal dosage of Drosophila Piwi and Aubergine, which bind to novel germline-enriched small non-coding RNAs called piRNAs, induces phenotypic variations that can be fixed in a population and stably inherited in later generations. The study also showed that Piwi directly interacts with Hop and Hsp90 and functions in the same pathway as Hsp90 in suppressing phenotypic variations. Further, we demonstrate for the first time that Piwi becomes phosphorylated in an Hsp90 dependent manner, thus providing insights into how Hsp90 may modulate Piwi's function in canalization. Crucially, this study also revealed two related yet distinct Piwi/piRNA pathway-dependent mechanisms responsible for suppression of phenotypic variation: epigenetic silencing of inherent genotype variations and suppression of transposon-mediated mutagenesis. Our work revealed a framework of a pathway that suppresses phenotypic variation. Our working hypothesis is that Hsp90 and Hop modulate Piwi function via its phosphorylation which then modulates Piwi function in canalization. By further unraveling the inner workings of each step of this pathway, we aim to shed light on the mechanisms that mediate this very important, yet poorly understood cellular phenomenon. The aims of the proposal are to- 1) Discover new components that mediate and/or regulate Piwi function in canalization using genetic screens, 2) Unravel the biochemical mechanism of Piwi mediated epigenetic regulation in canalization and finally 3) Understand how Hsp90-Piwi/piRNA pathway counteracts environmental stress induced by industrial pollutants like hexavalent chromium. Piwi proteins are present from protozoans to humans with conserved functions in stem cell self-renewal and germ line maintenance. However their role in suppression of phenotypic variation is a novel function that requires further characterization. Unraveling the mechanism by which Piwi functions in canalization using Drosophila as a model system will provide the necessary context for understanding its roles in humans and learning how the dysfunction of this process might affect human development and cause diseases.

项目摘要/摘要： 我们提案的长期目标是描述 Piwi/piRNA 途径的机制 使由严重环境应激和固有基因型变异引起的表型变异正常化。 运河化或发育稳健性提供了一个框架，其中生物体抵抗展示 面对组合基因型变异和严重环境压力时的表型变异。最近的 研究表明，热休克蛋白90（Hsp90）作为一种分子伴侣，在渠道化过程中发挥着关键作用。尽管已知 对于 Hsp90 的作用，由于缺乏详细的运河化分子机制，引起了很大的争论 几十年来。我们的实验室最近验证了“管道化”的存在，并发现了一种主要分子 途径涉及 Piwi、Hsp90 和 Hop。使用敏化眼发育试验，我们发现 减少与富含新种系的果蝇 Piwi 和茄子结合的母体剂量 称为 piRNA 的小非编码 RNA 可诱导表型变异，这些变异可以在群体中固定下来， 并在后世稳定传承。研究还表明 Piwi 直接与 Hop 和 Hsp90 相互作用， 其功能与 Hsp90 抑制表型变异的途径相同。进一步，我们证明了 Piwi 首次以 Hsp90 依赖性方式磷酸化，从而提供了关于如何磷酸化的见解 Hsp90 可能调节 Piwi 的运河化功能。至关重要的是，这项研究还揭示了两个相关但又不同的现象 负责抑制表型变异的 Piwi/piRNA 途径依赖机制：表观遗传 固有基因型变异的沉默和转座子介导的诱变的抑制。我们的工作 揭示了抑制表型变异的途径框架。我们的工作假设是 Hsp90 和 Hop 通过其磷酸化调节 Piwi 功能，然后调节 Piwi 功能 运河化。通过进一步揭示该途径每一步的内部运作，我们的目标是阐明 介导这种非常重要但人们知之甚少的细胞现象的机制。该组织的目标 提议是 - 1) 发现在运河化过程中介导和/或调节 Piwi 功能的新成分 遗传筛选，2）揭示Piwi介导的渠道化表观遗传调控的生化机制 最后 3) 了解 Hsp90-Piwi/piRNA 通路如何抵消由 六价铬等工业污染物。 Piwi 蛋白从原生动物到人类均存在 干细胞自我更新和生殖系维持的保守功能。然而，它们在抑制 表型变异是一种新功能，需要进一步表征。揭开机制 Piwi 在以果蝇为模型系统的运河化中发挥作用 将提供必要的背景 了解其在人类中的作用并了解该过程的功能障碍如何影响人类 发育并引发疾病。