Regulation of ESCRT-III Activity in Yeast

描述（由申请人提供）：运输所需的内体分选复合物（escrt）包括从酵母到人保存的细胞机制。escrt通过执行腔内囊泡（ILVs）的形成在晚期内体多泡体（MVBs）的生物发生中发挥作用，这些囊泡（ILVs）直接从内体膜向腔室的管腔萌发。内溶酶体融合时，ilv携带不需要的、受损的或危险的跨膜蛋白，在溶酶体内部水解降解，在维持细胞稳态中发挥关键作用。ESCRT-III复合物的功能障碍与神经退行性疾病有关，其开发对HIV-1和其他逆转录病毒以及某些细菌病原体的传播至关重要。体外研究表明，ESCRT-III在MVBs的ILV出芽途径中催化膜断裂。本研究的目的是确定体内控制ESCRT-III活性的调控机制，以出芽酵母酿酒酵母为模型系统。核心假设是ESCRT-III活性在体内由控制该复合物组装和拆卸的蛋白质调节。本研究将研究酵母中ESCRT-III的结构和功能如何受到调节该复合物组装和拆卸的蛋白质的影响。使用的方法包括电子显微镜、光学显微镜、蛋白质生物化学和功能分析。这项研究的基本原理是，如果要开发治疗ESCRT-III功能障碍或对抗人类病原体对ESCRT-III的利用的治疗策略，必须确定ESCRT-III的调控。利用酵母作为模型系统来了解这一调控与人类健康直接相关，因为ESCRT-III的功能机制是高度保守的。至于预期的结果，预计这项研究的完成将揭示核心的、广泛保守的调控ESCRT-III活性的机制。这些结果预计将产生重要的积极影响，因为它们将对内吞途径中一个鲜为人知的调控步骤产生根本性的见解，揭示与ESCRT-III相关疾病的预防和治疗的潜在靶点，并激发新的和创新的方法来理解ESCRT-III的功能机制。

DESCRIPTION (provided by applicant): The endosomal sorting complexes required for transport (ESCRTs) comprise a cellular machinery conserved from yeast to man. ESCRTs function in the biogenesis of late endosomal multivesicular bodies (MVBs) by executing the formation of intralumenal vesicles (ILVs), which bud directly from the endosomal membrane toward the lumen of the compartment. ILVs perform a critical role in maintaining cellular homeostasis by carrying unneeded, damaged, or dangerous transmembrane proteins to be degraded in the hydrolytic interior of the lysosome upon endolysosomal fusion. Dysfunction of the ESCRT-III complex has been linked to neurodegenerative disease, and its exploitation is essential for the spread of HIV-1 and other retroviruses as well as certain bacterial pathogens. In vitro studies have shown that ESCRT-III catalyzes membrane scission in the ILV budding pathway at MVBs. The objective of this research is to determine the regulatory mechanisms that control ESCRT-III activity in vivo, using the budding yeast Saccharomyces cerevisiae as a model system. The central hypothesis is that ESCRT-III activity is regulated in vivo by proteins that control the assembly and disassembly of this complex. This research will study how the structure and function of ESCRT-III is impacted in yeast by proteins that regulate assembly and disassembly of the complex. The methodology to be used includes electron microscopy, light microscopy, protein biochemistry, and functional assays. The rationale for this research is that the regulation of ESCRT-III must be defined if therapeutic strategies are to be developed that will treat ESCRT-III dysfunction or combat exploitations of ESCRT-III by human pathogens. Using yeast as a model system to understand this regulation is directly relevant to human health because the mechanism of ESCRT-III function is highly conserved. With respect to expected outcomes, it is anticipated that completion of this research will reveal the core, broadly conserved mechanisms that regulate ESCRT-III activity. Such results are expected to have an important positive impact because they will yield fundamental insight into a poorly understood regulatory step in the endocytic pathway, reveal potential targets for the prevention and treatment of diseases linked to ESCRT-III, and inspire new and innovative approaches to understand the mechanisms of ESCRT-III function.