INTRACELLULAR TRANSPORT--MANNOSE-PHOSPHATE RECEPTOR

在细胞内运输过程中，流向质膜的蛋白质， 分泌囊泡和溶酶体必须在内部相互分类 高尔基建筑群，并被送到他们适当的地址。从长远来看 本研究的目的是阐明细胞免疫的分子机制。 蛋白质的目标是特定的和不同的细胞内目的地。 目前对溶酶体靶向蛋白质的了解最多。 分类过程。可溶性溶酶体酶可获得甘露糖6-磷酸 (Man6P)标签；细胞内受体识别含man6P的蛋白质， 并以某种方式促进它们向溶酶体的运输。的具体目标 这项建议是：1.研究蛋白质在跨高尔基网络中的分选 (TGN)，将分离携带man6P受体的笼状蛋白包裹的囊泡 通过免疫吸附。它们的溶酶体膜糖蛋白含量 然后进行分析以确定这两种蛋白质是否被运输 从TGN到前溶酶体合在一起。 2.为了研究前溶酶体中蛋白质的分选，潜在的循环利用 溶酶体膜糖蛋白从溶酶体前到TGN将被 调查过了。一种可溶性溶酶体酶的转运能力 也将对TGN进行审查。 3.研究溶酶体前体囊泡转运的分子基础 对TGN来说，对促进这一点的细胞质因素的分析 将使用在此开发的无细胞系统启动该过程 实验室提纯N-乙基马来酰亚胺敏感因子Will 已尝试。我们将进行实验，以检验其潜在的作用 在这个转运过程中，有一种被称为“NSF”的转运因子。 最后，将产生抗肽抗血清来鉴定一个家族 促进细胞内转运的GTP结合蛋白。 希望这些细胞生物和生化实验将 进一步我们对蛋白质形成的分子机制的理解 在细胞质内的膜结合的隔间之间运输 真核细胞。

During intracellular transport, proteins destined for the plasma membrane, secretory vesicles, and lysosomes must be sorted from one another within the Golgi complex, and sent to their appropriate addresses. The long term goal of this research is to elucidate the molecular mechanisms by which proteins are targeted to specific and distinct intracellular destinations. The targeting of proteins to lysosomes is presently the best understood sorting process. Soluble lysosomal enzymes acquire a mannose 6-phosphate (man6P) tag; intracellular receptors recognize man6P-containing proteins, and somehow facilitate their transport to lysosomes. The specific aims of this proposal are: 1. To study protein sorting in the trans Golgi network (TGN), clathrin coated vesicles carrying man6P receptors will be isolated by immunoadsorption. Their content of a lysosomal membrane glycoprotein will then be analyzed to determine if these two proteins are transported together from the TGN to pre-lysosomes. 2. To study protein sorting in pre-lysosomes, the potential recycling of a lysosomal membrane glycoprotein from pre-lysosomes to the TGN will be investigated. The ability of a soluble lysosomal enzyme to be transported to the TGN will also be examined. 3. To study the molecular basic of vesicular transport from pre-lysosomes to the TGN, an analysis of the cytosolic factors that facilitate this process will be initiated using a cell-free system developed in this laboratory, Purification of an N-ethylmaleimide-sensitive factor will attempted. Experiments will be carried out to examine the potential role of clathrin and a transport factor termed "NSF" in this transport process. Finally, an anti-peptide antiserum will be generated to identify a family of GTP binding proteins that facilitates intracellular transport. It is hoped that these cell biological and biochemical experiments will further our understanding of the molecular mechanisms by which proteins are transported between membrane-bound compartments within the cytoplasm of eukaryotic cells.