Selective Autophagic Degradation of Peroxisomes in Yeast

描述(由申请人提供) 细胞中主要的降解隔间是溶酶体(或其 在植物和酵母中相当于液泡)。尽管细胞已经进化到 多种机制将细胞成分输送到溶酶体，溶酶体是主要的 途径是通过一系列统称为自噬的过程。频繁攻击目标 在这些过程中，有一种叫做过氧化体的细胞器，它含有必需的氧化作用。 酵素。过氧化体的数量、大小和酶含量变化很快。 为了响应细胞的新陈代谢需要，当某些特定的过氧化物体 新陈代谢类型不再有用，它们被有选择地通过自噬移除 流程。 这个项目的目标是了解控制 自噬。一种使用酵母毕赤酵母的遗传方法已经启动。 PASTORIS作为一个模型系统。这种酵母有大量的过氧化物体，当 在甲醇上培养，当转移到 要么是葡萄糖，要么是乙醇。在我们之前的FIRCA期间，我们 成功地从六个不同的菌株中分离和鉴定了巴斯德毕赤酵母突变体 自噬过氧化物体降解有缺陷的基因 葡萄糖。其中4个PDG基因被分离出来，DNA测序结果 表明这四种基因都编码未知功能的新蛋白质。 这项持续的FIRCA提案的具体目标是：(1) 确定四种PDG蛋白的亚细胞定位和搜索 与其产物相互作用的其他基因；(2)确定哪些其他基因 空泡分选途径受到每个基因突变的影响；以及(3) 开发和利用新的筛选和选择方法来分离 新的PDG基因的突变。这些研究的结果将提供重要的 对自噬机制的洞察。

DESCRIPTION (provided by applicant) The primary degradative compartment in the cell is the lysosome (or its equivalent in plants and yeasts, the vacuole). Although cells have evolved multiple mechanisms to deliver cellular components to the lysosome, the major route is via a set of processes collectively termed autophagy. Frequent targets of these processes are peroxisomes, organelles that contain essential oxidative enzymes. The number, size, and enzymatic content of peroxisomes change rapidly in response to a cell's metabolic needs, and when peroxisomes of a certain metabolic type are no longer useful, they are selectively removed by autophagic processes. The goal of this project is to understand the molecular mechanisms that control autophagy. A genetic approach has been initiated using the yeast Pichia pastoris as a model system. This yeast has large numerous peroxisomes when cultured on methanol, which are selectively autophagized upon transfer to either glucose or ethanol. During the period of our previous FIRCA, we succeeded in isolating and characterizing P. pastoris mutants in six different genes that are defective in autophagic peroxisome degradation in response to glucose. Four of these PDG genes were isolated, and DNA sequencing results indicate that all four encode novel proteins of unknown function. The specific aims of this continuing FIRCA proposal are: (1) to characterize the four PDG proteins by determining their subcellular location and searching for other genes whose products interact with them; (2) to determine what other vacuolar sorting pathways are affected by mutations in each gene; and (3) to develop and utilize novel screens and selection methods for the isolation of mutants in new PDG genes. Results of these studies will provide important insights into autophagic mechanisms.