INTRACELLULAR TRANSPORT--MANNOSE-PHOSPHATE RECEPTOR

在细胞内运输过程中，蛋白质被运往质膜， 分泌囊泡和溶酶体必须在内部彼此分离， 高尔基复合体，并发送到适当的地址。长期 这项研究的目的是阐明分子机制， 蛋白质被靶向特定的和不同的细胞内目的地。 蛋白质靶向溶酶体是目前最好的理解 排序过程。可溶性溶酶体酶获得甘露糖6-磷酸 （man 6P）标签;细胞内受体识别含man 6P的蛋白质， 并以某种方式促进它们运输到溶酶体。的具体目标 这些建议是：1.研究反式高尔基体网络中的蛋白质分选 (TGN)，将分离携带man 6P受体的网格蛋白包被的囊泡 通过免疫吸附。它们的溶酶体膜糖蛋白含量 然后将被分析以确定这两种蛋白质是否被运输 从TGN到前溶酶体。 2.为了研究前溶酶体中的蛋白质分选， 从前溶酶体到TGN的溶酶体膜糖蛋白将是 研究了可溶性溶酶体酶被转运的能力 TGN也将被审查。 3.研究前溶酶体囊泡转运的分子基础 TGN，分析促进这一点的胞质因子， 这一过程将使用在此开发的无细胞系统启动。 实验室，纯化N-乙基马来酰亚胺敏感因子将 企图。将进行实验，以检查 网格蛋白和称为“NSF”的运输因子在该运输过程中。 最后，将产生抗肽抗血清以鉴定家族 促进细胞内转运的GTP结合蛋白。 希望这些细胞生物学和生物化学实验将 进一步了解蛋白质的分子机制， 在细胞质内的膜结合区室之间运输 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.