Mechanisms of Transport in the Golgi Apparatus

本研究的总体目的是了解在高尔基体分泌的糖蛋白的末端糖基化过程中底物获得糖基转移酶的分子机制。包括本实验室在内的几个实验室的研究表明，特定的载体蛋白参与了将核苷酸糖运送到高尔基体管腔和将转移酶的核苷酸产物运送出管腔的过程。该项目的具体目标是：(1)确定负责udp -半乳糖和UMP跨高尔基膜运输的蛋白质；(2)分离和表征转运蛋白；(3)确定转运蛋白在膜中的定向，以及它们相对于半乳糖转移酶的分布。也许关于活细胞最引人注目的事情是生命过程发生所必需的高度有序和组织性。细胞被划分为执行特定功能的特定区域，这种划分允许对这些功能进行高度调节，而对其他细胞功能的干扰最小。其中一些“隔室”由膜封闭的胞内细胞器组成，特定的生化反应在其中发生。在“分泌”蛋白（包括膜蛋白和溶酶体蛋白，其生物合成也利用“分泌途径”）的翻译后加工的情况下，通过添加末端糖（和其他取代基）的后期翻译后修饰发生在高尔基体的管腔内。新生蛋白质被运输到的一组膜结合的生物合成室。糖转移酶反应需要膜结合转移酶、新生糖蛋白和核苷酸连接的“活化”糖底物的参与。激活的核苷酸糖不是在高尔基腔内生物合成的，而是在细胞的其他地方（通常在细胞质中）生物合成的。这些化合物不容易穿过生物膜。同样，转移酶反应的产物游离核苷酸也不容易穿过细胞膜。然而，核苷酸糖进入管腔是至关重要的，反应产物（如果允许其积累将抑制反应）从反应部位流出也是至关重要的。通过关注一种特殊的核苷酸糖（udp -半乳糖）的运输和区隔利用，他的研究将增加我们对这种微妙而关键的区隔如何在活细胞中起作用的理解。

The overall aim of this research is to understand the molecular mechanisms by which substrates gain access to the glycosyltransferases during terminal glycosylation of secreted glycoproteins in the Golgi apparatus. Studies from several laboratories, including this one, have demonstrated that specific carrier proteins are involved in transporting nucleotide sugars into the Golgi lumen and nucleotide products of the transferases out of the lumen. The specific goals of the project are to: (1) identify the proteins responsible for UDP-galactose and UMP transport across Golgi membranes; (2) isolate and characterize the transport proteins; and (3) determine how the transporters are oriented in the membrane, and how they are distributed relative to galactosyltransferases. Perhaps the most striking thing about living cells is the very high degree of order and organization which is required for life processes to occur. The cell is compartmentalized into specific regions which carry out specific functions, and this compartmentalization allows for a high degree of regulation of those functions with minimal interference from other cellular functions. Some of these "compartments" consist of membrane- enclosed intracellular organelles, within which specific biochemical reactions occur. In the case of post-translational processing of "secretory" proteins (including membrane proteins and lysosomal proteins, the biosyntheses of which also utilize the "secretory pathway"), late post-translational modification by the addition of terminal sugars (and other substituents) takes place within the lumen of the Golgi apparatus, a complex collection of membrane-bound biosynthetic compartments to which the nascent proteins are transported. The sugar transferase reactions require the participation of the membrane-bound transferase enzymes, the nascent glycoprotein, and the nucleotide-linked "activated" sugar substrates. The activated nucleotide sugars are not biosynthesized within the Golgi lumen, but elsewhere in the cell (usually in the cytoplasm). These compounds do not readily cross biological membranes. Similarly, the resulting reaction product of the transferase reaction, free nucleotide, also does not cross membranes readily. Yet, it is critical for the nucleotide sugars to enter the lumen, and also critical for the reaction products (which would inhibit the reaction if allowed to acumulate) to exit from the site of the reaction. By focusing on the transport and compartmentalized utilization of a particular nucleotide sugar, UDP-galactose, his research will increase our understanding of how this delicate, yet critical, compartmentalization works in the living cell.