Stress Granule clearance via Autophagy: Mechanism, Regulation and Consequences

 描述（由申请人提供）：应激颗粒（SG）是非翻译mRNA-蛋白质复合物（mRNP）的组装体，其在整个真核生物中是保守的，并且通常仅在细胞应激期间大量翻译被抑制时瞬时形成。SGS 螯合mRNP和信号传导因子，并通过调节基因表达和细胞信号传导在细胞应激反应中发挥关键作用。以前，我们确定SG是通过一种新的选择性自噬途径清除的，我们称之为颗粒吞噬。这一发现很重要，因为它证明了一种全新的SGs清除方式;以前认为SGs只能通过非破坏性手段分解。因此，颗粒吞噬可能影响mRNA水平的基因表达，并可以解释几种“包涵体”疾病的潜在病理学，如肌萎缩性侧索硬化症，额颞叶痴呆和佩吉特病。这些通常以受影响细胞中的组成性SG样聚集体和自噬缺陷为特征。我们理解的关键差距包括SGs是如何通过颗粒吞噬机制靶向的，这个过程是如何调节的，以及颗粒吞噬的生理相关性是什么。我们的长期目标是了解细胞组装、分解或清除mRNP颗粒的方式，这如何影响mRNA调控，以及以异常mRNP颗粒形成为特征的疾病的进展。本申请的总体目标是鉴定影响颗粒吞噬的基因，并确定颗粒吞噬活性所需的SG和自噬蛋白之间的蛋白质相互作用。我们还希望确定颗粒吞噬是如何调节的，以及颗粒吞噬对在包涵体疾病中观察到的SG样蛋白聚集体的mRNA稳定性和清除的影响。我们的中心假设是，选择性自噬受体蛋白与SG相互作用，并招募自噬机制，特别是在通过其他手段限制SG分解的条件下。使用酵母和细胞系模型系统，以及SG和自噬领域中已证实的遗传、生物化学和显微镜技术，我们将以以下三个目标来测试这一假设：什么基因促进颗粒吞噬，以及如何？2.）什么情况下会诱发嗜粒细胞吞噬，这是如何调节的？3.）第三章颗粒吞噬作用如何影响SG样疾病聚集体的mRNA衰减和清除？这一提议是创新的，因为它将增强我们对一种新的选择性自噬途径的理解，该途径靶向一种在疾病中具有重要意义的生理学底物（SGs）。这一贡献是重要的，因为更详细地了解粒细胞吞噬可能会提出新的范式的选择性自噬功能，并阐明新的模式的mRNA和细胞信号调节相关的一般细胞生物学，和疾病的特征或异常SG积累引起的。

 DESCRIPTION (provided by applicant): Stress granules (SGs) are assemblies of non-translating mRNA-protein complexes (mRNPs) that are conserved throughout eukaryotes, and which normally form only transiently when bulk translation is inhibited during cellular stress. SGs sequester mRNPs and signaling factors, and play key roles in cellular stress responses by modulating gene expression and cell signaling. Previously, we determined that SGs are cleared via a novel selective autophagic pathway that we termed granulophagy. This finding was important as it demonstrated a fundamentally new way in which SGs are cleared; previously SGs were thought to disassemble only by non-destructive means. Granulophagy thus likely impacts upon gene expression at the mRNA level, and could explain the underlying pathology of several "inclusion body" disorders such as Amyotrophic Lateral Sclerosis, Frontotemporal Lobar Dementia and Paget's disease. These are typically characterized by constitutive SG-like aggregates and autophagy defects in affected cells. Key gaps in our understanding include how are SGs mechanistically targeted by granulophagy, how is this process regulated, and what is the physiological relevance of granulophagy. Our long-term goal is to understand the means by which cells assemble, disassemble or clear mRNP granules, how this affects mRNA regulation, and the progression of diseases characterized by aberrant mRNP granule formation. The overall objectives of this application are to identify genes affecting granulophagy and determine protein interactions between SG and autophagy proteins that are required for granulophagy activity. We also wish to determine how granulophagy is regulated, and what the consequences of granulophagy are on both mRNA stability and clearance of SG-like protein aggregates observed in inclusion body disorders. Our central hypothesis is that selective autophagy receptor proteins interact with SGs and recruit the autophagic machinery, especially under conditions that limit SG disassembly via other means. Using yeast and cell line model systems, and proven genetic, biochemical and microscopy techniques in the SG and autophagy fields, we shall test this hypothesis with the following three aims: 1.) What genes promote granulophagy, and how? 2.) What conditions induce granulophagy, and how is this regulated? 3.) How does granulophagy affect mRNA decay and clearance of SG-like disease aggregates? This proposal is innovative, as it will enhance our understanding of a novel selective autophagic pathway that targets a physiologically important substrate (SGs) with implications in disease. This contribution is important because understanding granulophagy in greater detail may suggest new paradigms of selective autophagic function, and illuminate new modes of mRNA and cell signaling regulation relevant to both general cell biology, and to diseases characterized or caused by aberrant SG accumulation.