酿酒酵母中ESCRT复合体在自噬前体闭合中的作用及机制研究

Autophagy is a conserved degradation pathway for eukaryotic cells to recycle cytoplasmic constitutes which are enwrapped by membrane structures, and degraded and released by lysosome. Autophagy dysfunction will affect cellular homeostasis and may result in abnormal growth, development and disease. The importance of autophagy study was further underscored due to the Nobel Prize in Physiology or Medicine in 2016 for autophagy. In the past two decades, great progresses were made in autophagy study. However, there are still lots of unanswered critical questions in the autophagy field, e.g., which proteins regulate phagophore closure/sealing and what the underlying mechanism is after the expansion of the phagophore in the autophagy process. We recently reported that the Vps21/Rab5 module is important for phagophore closure in budding yeast, although the molecular mechanism is still unknown. Our latest preliminary data indicate that the ESCRT complex subunits, which are the effectors of Vps21, are also important for phagophore closure. In this proposal, we are going to explore the molecular mechanism for phagophore closure under the coordination of the Vps21/Rab5 module and the ESCRT complex with multiple methods. We will determine whether the ESCRT complex is recruited to the phagophore for phagophore closure/sealing through the Vps21-dependent interactions between the ESCRT complex subunit Snf7 and Atg proteins. The obtaining results will greatly contribute to our understanding on the regulation mechanism of phagophore closure/sealing.

自噬是真核细胞将待降解物通过膜结构包裹，经溶酶体降解释放后循环利用的一种保守途径。自噬障碍会影响细胞稳态，导致生长发育异常及病变。自噬研究的重要性也因2016年的诺贝尔生理或医学奖颁发给自噬领域愈发凸显。过去的二十多年，自噬的研究取得了重要的进展，但仍有许多关键的问题没有回答，如自噬过程中自噬前体延伸后是什么蛋白通过什么分子机制将自噬前体闭合形成自噬体？我们最近报道了酿酒酵母中Vps21/Rab5模块蛋白对自噬前体闭合非常重要，但分子机制不清楚。我们的最新研究发现Vps21的效应物ESCRT复合体亚基缺失也影响自噬前体闭合。本项目将通过多重方法探讨Vps21和ESCRT复合体协同影响自噬前体闭合的分子机制。解析Vps21通过调节ESCRT亚基Snf7与自噬蛋白Atg17等的互作，将ESCRT复合体招募至自噬前体上行使封口功能的可能性。研究结果将对揭示自噬前体闭合的调控机理做出重大贡献。

自噬是真核细胞将待降解物通过膜结构包裹，经溶酶体降解释放后循环利用的一种保守途径。自噬障碍会影响细胞稳态，导致生长发育异常及病变。自噬研究的重要性也因2016年的诺贝尔生理或医学奖颁发给自噬领域愈发凸显。过去的二十多年，自噬的研究取得了重要的进展，但仍有许多关键的问题没有回答，如自噬过程中自噬前体延伸后是什么蛋白通过什么分子机制将自噬前体闭合形成自噬体？我们前期报道了酿酒酵母中Vps21/Rab5模块蛋白对自噬前体闭合非常重要，但分子机制不清楚。基于ESCRT复合体对膜性细胞器具有封口功能及囊泡运输过程中Vps21/Rab5模块蛋白调控ESCRT复合体对多囊泡体封口，我们检测了ESCRT复合体亚基缺失时的自噬发生情况，发现ESCRT复合体亚基缺失也影响自噬前体闭合并用多种方法验证了这一特性。我们更是发现这两类蛋白缺失后出现类似性状的原因是Vps21缺失后使ESCRT复合体亚基Snf7和Vps4无法定位到自噬体上，更是使得Snf7与Atg17的互作与共定位显著减少，而人为迫使Vps21缺失菌株中Snf7与Atg17共定位可修复自噬缺陷，说明Vps21通过调节ESCRT亚基Snf7与自噬蛋白Atg17的互作，将ESCRT复合体招募至自噬前体上行使封口功能。研究结果对揭示自噬前体闭合的调控机理做出了重大贡献，发表于细胞生物学顶级期刊Journal of Cell Biology上并被收集到自噬历史文献专集JCB65：Autophagy中。