MAL proteolipids in apical protein delivery in epithelia

描述(申请人提供)：极化上皮细胞的质膜(PM)在物理上是连续的，但在功能和成分上分为两个独立的区域：顶端和基底外侧。新合成的PM蛋白实现其特定而不对称分布的分子分选机制和途径正在许多极化上皮细胞类型中被活跃地研究。新合成的顶端蛋白通过两条途径传递到顶端PM：直接途径或间接途径。使用直接途径的蛋白质直接从跨高尔基网络(TGN)输送到根尖表面，而使用间接途径的蛋白质则采用更迂回的途径。它们首先从TGN传递到基底侧域，在那里它们通过内吞作用被取回，并被跨细胞传递到根尖表面。本研究的重点是MAL、L和MAL 2蛋白脂在调节根尖传递中的作用。MAL-L参与了蛋白质的直接分选，而MAL2在间接分选中发挥了重要作用。我们的长期目标是了解调节肝细胞顶端蛋白递送的机制，肝细胞是肝脏的主要上皮细胞。与大多数简单的上皮细胞不同，肝细胞使用间接途径运送心尖蛋白。我们之前已经证明，肝细胞的间接分选需要胆固醇和神经鞘糖脂。由于MAL L和MAL 2在两条途径中都被认为是顶端PM递送的重要调节因子，而且它们的活动需要胆固醇和神经鞘糖脂，我们建议在AIM 1的极化肝细胞和AIM 3的非极化肝细胞中研究MAL蛋白脂在顶端PM分选中的作用机制。有趣的是，MAL L和MAL 2的组织表达模式通常是不重叠的，这表明它们参与了细胞特异性的转运途径。MAL2是否赋予和调节肝细胞顶端蛋白的间接分类？同样，在肾脏来源的细胞中，Mal L是否直接从TGN进行顶端靶向？我们打算在目标2中验证这一假说。识别调控新合成的根尖驻留物通过囊泡传递到根尖PM的机制，对于理解这一表面是如何建立和维持至关重要。这些信息将提供对人类肝病和其他上皮细胞疾病的分子基础的洞察。

DESCRIPTION (provided by applicant): The plasma membrane (PM) of polarized epithelial cells is physically continuous, but functionally and compositionally divided into two separate domains: the apical and basolateral. The molecular sorting mechanisms and pathways by which newly synthesized PM proteins achieve their specific yet asymmetric distributions are actively being examined in many polarized epithelial cell types. Newly synthesized apical proteins are delivered to the apical PM by two pathways: the direct or indirect. Proteins using the direct pathway are delivered directly form the trans-Golgi network (TGN) to the apical surface whereas proteins using the indirect pathway take a more circuitous route. They are first delivered from the TGN to the basolateral domain where they are retrieved by endocytosis and transcytosed to the apical surface. This proposal focuses on the role of MAL l and MAL2 proteolipids in regulating apical delivery. MAL l has been implicated in sorting proteins in the direct pathway and MAL2 has a presumed role in indirect sorting. Our long-term goal is to understand the mechanisms regulating apical protein delivery in hepatocytes, the major epithelial cell of the liver. Unlike most simple epithelial cells, hepatocytes use the indirect pathway for apical protein delivery. We have previously shown that indirect sorting in hepatocytes requires cholesterol and glycosphingolipids. Because MAL l and MAL 2 have been identified as important regulators of apical PM delivery in both pathways and because their activity requires cholesterol and glycosphingolipids, we propose to characterize the mechanism by which the MAL proteolipids function in apical PM sorting in polarized hepatic cells in Aim 1 and in non-polarized hepatic cells in Aim 3. Interestingly, the tissue expression patterns of MAL l and MAL 2 are generally non-overlapping suggesting they participate in cell-specific transport pathways. Does MAL2 confer and regulate indirect sorting of apical proteins in hepatocytes? Likewise, does MAL l confer direct apical targeting from the TGN in kidney-derived cells? We intend to test this hypothesis in Aim 2. Identifying the mechanisms regulating the vesicle-mediated delivery of newly synthesized apical residents to the apical PM is imperative to understanding how this surface is established and maintained. Such information will provide insight into the molecular basis of human liver disease and other diseases of epithelial cells.