Mechanisms of Transport in the Golgi Apparatus

这项研究的总体目的是了解底物在高尔基体中分泌的糖蛋白的末端糖基化过程中获得糖基转移酶的分子机制。包括这个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将核苷酸糖输送到高尔基体腔内，并将转移酶的核苷酸产物输送出腔外。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)分离和鉴定运输蛋白；(3)确定转运蛋白在膜中的定位，以及它们相对于半乳糖转移酶的分布方式。也许关于活细胞最引人注目的事情是生命过程发生所需要的高度的秩序和组织。细胞被划分成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”由被膜包裹的细胞内细胞器组成，在这些细胞器内发生特定的生化反应。在“分泌型”蛋白质(包括膜蛋白和溶酶体蛋白，其生物合成也利用“分泌型途径”)的翻译后加工的情况下，通过添加末端糖(和其他取代基)的晚期翻译后修饰在高尔基体的管腔内发生，膜结合的生物合成隔间的复杂集合，新生蛋白质被输送到这些隔间。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。被激活的核苷酸糖不是在高尔基体腔内生物合成的，而是在细胞的其他地方(通常在细胞质中)合成的。这些化合物不容易穿过生物膜。同样，转移酶反应产生的反应产物--游离核苷酸也不容易跨膜。然而，核苷酸糖进入管腔是关键的，反应产物(如果允许累积将抑制反应)离开反应部位也是关键的。通过专注于特定核苷酸糖UDP-半乳糖的运输和分区利用，他的研究将增加我们对这种微妙但关键的分区在活细胞中是如何工作的理解。

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.