The contribution of the secretory pathway to macroautophagy

 描述(申请人提供)：当细胞受到压力时，被称为大自噬的高度保守的分解代谢途径上调。这一途径的上调导致自噬小体的形成，自噬小体是以蛋白质和细胞器为目标进行降解的不同隔室。在细胞应激过程中，对制造自噬小体的膜的高需求导致了细胞内膜的戏剧性重组。因此，分泌途径被下调，分泌途径中的膜被重定向到宏观自噬途径。最近的研究发现，内质网来源的COPII囊泡是自噬小体形成的膜来源，这表明细胞应激时分泌途径的下调是COPII囊泡从ER-高尔基体途径转移到宏观自噬途径的结果。了解COPII囊泡是如何从它们的典型路径重定向到另一条途径的，对于阐明自噬领域长达40年的谜团至关重要，自噬是在诱导巨型自噬时发生的膜重组事件的启动。在这项提案中，我提出了三个具体的目标，以解决磷酸化在靶向COPII囊泡到宏观自噬途径中所起的作用。1.我们已经确定了COPII涂层上的三个对细胞活力重要的磷酸化位点。将使用多种方法来确定一种蛋白的磷酸化 或更多的这些位点在将COPII囊泡转移到宏观自噬途径中发挥作用。2.我们将确定将COPII囊泡重定向到宏观自噬途径的激酶。这些研究可能会确定将流量重定向到宏自噬途径的激酶与调节诱导宏自噬的TORC1相互作用网络之间的联系。3.我们已经在内质网高尔基体系绳中确定了一个结构域，Uso1/p115，它是将COPII囊泡定向到高尔基体的关键。我们将确定该结构域的磷酸化是否有助于将COPII囊泡重定向到自噬途径。

 DESCRIPTION (provided by applicant): When cells are stressed, the highly conserved catabolic pathway called macroautophagy is upregulated. The upregulation of this pathway induces the formation of autophagosomes, distinct compartments that target proteins and organelles for degradation. The high demand for membrane to make autophagosomes during cell stress leads to a dramatic reorganization of intracellular membranes. Consequently, the secretory pathway is downregulated and membrane from the secretory pathway is redirected to the macroautophagy pathway. The recent finding that ER-derived COPII coated vesicles are a membrane source for autophagosome formation suggests that the down regulation of the secretory pathway during cell stress is the result of diverting COPII vesicles from the ER-Golgi pathway to the macroautophagy pathway. Understanding how COPII vesicles are redirected from their canonical route to an alternate pathway is critical to elucidating a 40 year mystery in the autophagy field, the initiation of the membrane reorganization events that occur when macroautophagy is induced. In this proposal I present three specific aims to address the role that phosphorylation plays in targeting COPII vesicles to the macroautophagy pathway. 1. We have identified three patches of phosphorylation sites on the COPII coat that are important for cellular viability. A variety of approaches will be used to determine if the phosphorylation of one or more of these sites plays a role in diverting a COPII vesicle to the macroautophagy pathway. 2. We will identify the kinases that redirect COPII vesicles to the macroautophagy pathway. These studies are likely to define the connection between the kinases that redirect traffic to the macroautophagy pathway and the TORC1 interaction network that regulates the induction of macroautophagy. 3. We have identified a domain in the ER-Golgi tether, Uso1/p115, that is critical for directing a COPII vesicle to the Golgi. We will determine if phosphorylation of this domain facilitates the redirection of COPII vesicles to the autophagic pathway.