MECHANISMS OF ABERRANT EGF RECEPTOR SORTING IN POLYCYSTIC KIDNEY DISEASE

上皮细胞极性的形成是一个基本的过程 胚胎发育和器官发生。成人极化上皮 动物在宿主和外部环境之间形成了物理屏障 对于动态平衡是必不可少的。极化上皮保持明显的顶端 (AP)和(BL)膜结构域，通过主动调节 脂质和蛋白质的膜分布以及亚细胞的分布 每个表面都有独特的膜状细胞骨架。许多问题仍然存在 关于膜极性是如何实现的，其中最主要的是AP和BL如何实现 蛋白质被包装在不同的运输小泡中，在跨高尔基体- 网络(TGN)。BL蛋白的分选部分是通过 识别不同的细胞质分选信号，似乎也 规范内小体的极化分选。尽管许多BL分选信号 有共同的特征，共识的主题还没有出现，这使得它变得不清楚 所有的BL信号是否通过相同或不同的方式来调节运输 小路。我们建议通过研究一本小说来解决这个问题 我们已经在EGF受体(EGFR)中发现了自主的B1信号。 该信号位于EGFR中的残基K652至A674之间。 膜结构域，并介导胞质截短的BL转运 含有管腔的EGFR和蛋白质嵌合体。这个BL信号很关键 酪氨酸和亮氨酸残基，不与任何已知的EGFR重叠 内吞信号，区别于许多其他井的特征- 表征了BL对信号的排序。计算机模拟显示出一种倾向 这一区域形成两亲性螺旋，与其他BL相反 其关键结构表明该区域倾向于 形成两亲性螺旋，而不是其他BL信号，其关键 结构处于贝塔转弯阶段。该区域还诱导已知底物T654 对于蛋白激酶C，增加了它的活性是 受磷酸化调节。眼下的目标是理解 该信号建立和维持EGFR极性的机制。 一个长期目标是了解导致多囊肾的基因是如何 疾病改变了这一过程，因为非极性EGFR表达是一种常见的 在人类和动物的疾病模型中都发现了肾囊肿。这个 具体目标将：检验细胞质BL分选的假设 截短的EGFR严重依赖于体内的特定氨基酸 BL信号；检验K652至A674残基调节BL的假设 全长EGFR的运输；检验残基K652到 A674调节内体的极化分选；并表征了 通过识别与EGFR相互作用的蛋白质来实现EGFR分选机制 残基K652至A674。

Formation of epithelial cell polarity is a fundamental process in embryonic development and organogenesis. Polarized epithelia in adult animals form a physical barrier between the host and external environment essential for homeostasis. Polarized epithelia maintain distinct apical (Ap) and basolateral (BL) membrane domains, by actively regulating membrane distribution of lipids and proteins as well as the sub- membranous cytoskeleton unique to each surface. Many questions remain about how membrane polarity is achieved, chief among them how Ap and BL proteins are packaged in distinct transport vesicles at the trans-Golgi- network (TGN). Sorting of BL proteins is mediated in part through recognition of distinct cytoplasmic sorting signals that also appear to regulate polarized sorting in endosomes. Although many BL sorting signals have common features, consensus motifs have not emerged, making it unclear whether all BL signals mediate transport by the same or different pathways. We propose to address this question by studying a novel autonomous Bl signal which we have identified in the EGF receptor (EGFR). This signal is located between residues K652 to A674 in the EGFR juxta- membrane domain, and mediates BL transport of cytoplasmically truncated EGFRs and protein chimeras containing a luminal. This BL signal critical tyrosine and leucine residues and do not overlap any of the known EGFR endocytic signals, features that distinguish it from many other well- characterized BL sorting signals. Computer modeling suggests a propensity for this region to form an amphipathic helix, in contrast to other BL signals whose critical structure suggests a propensity for this region to form a amphipathic helix, in contrast to other BL signals whose critical structure in a beta-turn. This region also induce T654, a known substrate for protein kinase C, raising the possibility that its activity is regulated by phosphorylation. Immediate goals are to understand the mechanism by which this signal establishes and maintains EGFR' polarity. A long-term goal is to understand how genes which cause polycystic kidney disease alter this process, since non-polar EGFR expression is a common finding that renal cysts both in humans and animals disease models. The specific aims will: test the hypothesis that BL sorting of cytoplasmically truncated EGFRs is critically dependent on particular amino acids in the BL signal; test the hypothesis that residues K652 to A674 regulate BL transport of full-length EGFRs; test the hypothesis that residues K652 to A674 regulate polarized sorting in endosomes; and characterize elements of the EGFR sorting machinery by identifying proteins that interact with EGFR residues K652 to A674.