Basolateral targeting in polarized epithelial cells

描述（由申请方提供）：上皮细胞极化为不同的顶侧和基底侧质膜结构域。顶膜面向身体外部，基底外侧膜与结缔组织接触。在任何给定的上皮细胞的生命周期中，维持这种既定的极性对于器官功能至关重要。例如，极性的丧失使细胞能够从单层崩解，这是转移性癌症的早期步骤。 顶端和基底侧质膜域具有不同的脂质和质膜受体组。为了保持这种独特的分布，细胞需要不断地将新合成的蛋白质沿着生物合成途径分类，并将内化的受体再循环到正确的膜上。基底外侧靶向通常取决于跨膜蛋白质的胞质尾区中的靶向决定簇和识别它们并介导分选的胞质衔接分子。我们鉴定了一种主要的胞质组分，网格蛋白-衔接子复合物AP-1B（Folsch等人，1999年）。网格蛋白衔接子复合物是四异聚体。典型地，中等亚单元直接与分选信号相互作用，因此特别感兴趣。AP-1B的中亚基mu 1B仅在上皮组织中表达，并直接与基底外侧货物分子相互作用（Folsch et al.，2001年）。 我们的工作将集中在定义AP-1B功能的分子机制及其作用位点。此外，我们将分析AP-1B的膜募集是如何调节的。此外，我们将研究导致AP-1B介导的基底外侧靶向的分子相互作用。我的实验室将联合收割机结合细胞生物学、生物化学和遗传学方法，以解决以下具体目标： 1)AP-1B在细胞内的哪个位点完成其分选功能？ 2)AP-1B膜募集和囊泡形成是如何调节的？ 3)AP-1B特异性分选事件的分子基础是什么？ 更好地理解上皮细胞保持极性的过程是必不可少的，如果我们想知道分子分选中的哪些缺陷可能将上皮细胞转化为癌细胞，仅举一个与细胞极性丧失相关的疾病的例子。

DESCRIPTION (provided by applicant): Epithelial cells are polarized into distinct apical and basolateral plasma membrane domains. The apical membrane faces the outside of the body and the basolateral membrane is in contact with connective tissues. During the lifetime of any given epithelial cell it is crucial for organ function that this established polarity is maintained. For example, loss of polarity enables a cell to disintegrate from a monolayer, which is an early step in metastatic cancer. Apical and basolateral plasma membrane domains have distinct sets of lipids and plasma membrane receptors. To maintain this distinct distribution, the cells continuously need to sort newly synthesized proteins along the biosynthetic pathway and recycle internalized receptors to the correct membrane. Basolateral targeting often depends on targeting determinants in the cytoplasmic tail of a transmembrane protein and cytosolic adaptor molecules that recognize them and mediate sorting. We identified one major cytosolic component, the clathrin-adaptor complex AP-1B (Folsch et al., 1999). Clathrin adaptor complexes are tetraheteromers. Typically the medium subunit interacts directly with the sorting signals and is therefore of particular interest. The medium subunit of AP-1B, mu1B is exclusively expressed in epithelial tissues and directly interacts with basolateral cargo molecules (Folsch et al., 2001). Our work will focus on defining the molecular mechanisms of AP-1B function and its site(s) of action. Moreover, we will analyze how the membrane recruitment of AP-1B is regulated. Furthermore, we will investigate the molecular interactions leading to AP-1B mediated basolateral targeting. My laboratory will combine cell biological, biochemical and genetic approaches to resolve the following specific aims: 1) At which intracellular site(s) does AP-1B fulfill its sorting function? 2) How is AP-1B membrane recruitment and vesicle formation regulated? 3) What is the molecular basis for AP-1B specific sorting events? A better understanding of the processes by which epithelial cells maintain polarity is essential, if we want to learn which defects in molecular sorting may transform epithelial cells into cancer cells, to name only one example of diseases associated with the loss of cell polarity.