ANALYSIS OF GOLGI MEMBRANE PROTEIN LOCALIZATION IN YEAST

分泌和分泌细胞中蛋白质的分选和定位 内体途径将被调查，使用模型系统的定位 S.啤酒。PI的工作模型提出了两个 独立的机制，都发挥作用，以本地化蛋白质的TGN：选择性 从前液泡区室中回收，并降低运输速率 到TGN。初步数据表明，提取涉及的相互作用， 货物蛋白与Vps35p，一个前液泡外周膜蛋白，和这个 假设将在遗传抑制方法中进行测试，以及在 生物化学结合测定。建议从TGN缓慢退出， 磷酸化的居民TGN蛋白质的胞质结构域，这将是 使用生物化学方法进行研究，以确定 磷酸化，然后特异性位点诱变以改变这些 磷酸化位点。最后，几个基因已被确定在遗传 筛选为有缺陷的高尔基体保留蛋白质，其中四个将 进一步详细研究：grd 20，grd 21，grd 22和grd 23。

The sorting and localization of proteins in the secretory and endosomal pathways will be investigated, using the model system of localization of proteins to the TGN in S. cerevisiae. The PI's working model proposes two independent mechanisms that both act to localize proteins to the TGN: selective retrieval from the prevacuolar compartment and a reduced rate of transport out to the TGN. Preliminary data suggest that retrieval involves the interaction of cargo proteins with Vps35p, a prevacuolar peripheral membrane protein, and this hypothesis will be tested in a genetic suppressor approach, as well as in biochemical binding assays. Slow exit from the TGN is proposed to involve the phosphorylation of cytosolic domains of resident TGN proteins, and this will be investigated using biochemical approaches to identify the specific sites of phosphorylation followed by specific site mutagenesis to alter these phosphorylation sites. Finally, several genes have been identified in genetic screens as defective in Golgi retention of proteins, and four of these will be investigated in further detail: grd20, grd21, grd22 and grd23.