Novel Role for Piwi/piRNA pathway in developmental robustness

项目摘要/摘要： 我们建议的长期目标是描述piwi/piRNA途径的机制。 使由严重环境压力和固有的基因变异引起的表型变异正常化。 管道化或发育稳健性提供了一个框架，在这个框架中，有机体拒绝展示 面对组合的基因变异和严重的环境压力的表型变异。近期 研究表明，Hsp90是一种分子伴侣，在管道形成过程中起着关键作用。尽管已知 对于Hsp90的作用，缺乏一个详细的分子机制来支持导管形成，这引发了巨大的争论 几十年来。我们的实验室最近证实了“管化”的存在，并发现了一个主要的分子 涉及Piwi、Hsp90和Hop的通路。使用敏感化的眼睛发育测试，我们显示出 减少果蝇Piwi和Aubergine的母体剂量，这与新的种系丰富相结合 被称为piRNAs的小的非编码RNA可以诱导表型变异，这种变异可以在群体中固定，并且 稳定地遗传给后代。研究还表明，Piwi与Hop和Hsp90直接相互作用，并 Hsp90在抑制表型变异方面的作用与Hsp90相同。此外，我们还演示了 这是Piwi第一次以Hsp90依赖的方式磷酸化，从而提供了对如何 HSP90可能参与调节Piwi的通道化功能。至关重要的是，这项研究还揭示了两个相关但截然不同的 抑制表型变异的Piwi/piRNA途径依赖机制：表观遗传学 沉默固有的基因变异和抑制转座子介导的突变。我们的工作 揭示了抑制表型变异的途径的框架。我们的工作假设是 HSP90和Hop通过其磷酸化来调节Piwi功能，进而在 通道化。通过进一步解开这一途径每一步的内部工作原理，我们的目标是阐明 调节这一非常重要但知之甚少的细胞现象的机制。该计划的目标是 建议是-1)发现新的成分，在管道中调节和/或调节Piwi功能，使用 遗传筛选，2)揭示Piwi介导的表观遗传调控的生化机制 最后，了解Hsp90-piwi/piRNA通路如何抵消由 工业污染物，如六价铬。Piwi蛋白存在于从原生动物到人类的 在干细胞自我更新和生殖系维持中的保守功能。然而，他们在镇压 表型变异是一种新的功能，需要进一步的表征。通过以下方式解开这个机制 以果蝇为模型系统的Piwi在导管形成中的作用 将为以下内容提供必要的上下文 了解它在人类中的作用，并了解这一过程的功能障碍可能如何影响人类 发展和引发疾病。

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.