MECHANISMS OF ABERRANT EGF RECEPTOR SORTING IN POLYCYSTIC KIDNEY DISEASE

多囊肾疾病中异常 EGF 受体分选的机制

• 批准号: 6651773
• 负责人: CATHLEEN R CARLIN
• 金额: $ 13.53万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-01 至 2003-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6651773
• 关键词: CHO cells; MDCK cell; apical membrane; basolateral membrane; biological signal transduction; cellular polarity; epidermal growth factor; gene mutation; genetic regulation; growth factor receptors; human tissue; intracellular transport; membrane permeability; nucleic acid sequence; phosphorylation; point mutation; polycystic kidney; polymerase chain reaction; protein kinase C; protein sequence; protein transport

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.

上皮细胞极性的形成是上皮细胞分化的基本过程。 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物理屏障 对体内平衡至关重要极化的上皮细胞保持明显的顶端 (Ap)和基底外侧（BL）膜结构域，通过主动调节 脂质和蛋白质的膜分布，以及亚 每个表面都有独特的膜状细胞骨架。还有许多问题 关于膜极性是如何实现的，其中最主要的是Ap和BL是如何实现的， 蛋白质被包装在不同的运输囊泡中， 网络（TGN）。BL蛋白的分选部分通过以下途径介导： 识别不同的细胞质分选信号， 调节内体中的极化分选。虽然许多BL分选信号 有共同的特征，一致的基序还没有出现， 所有BL信号是否通过相同或不同的 途径。我们建议通过研究一部小说来解决这个问题 我们已经在EGF受体（EGFR）中鉴定了自主B1信号。 该信号位于EGFR转录因子中的残基K652至A674之间。 膜结构域，并介导细胞质截短的BL转运 EGFR和蛋白质嵌合体含有管腔。此BL信号临界 酪氨酸和亮氨酸残基，并且不与任何已知的EGFR 内吞信号，这些特征使它区别于许多其他的 其特征在于BL分选信号。电脑模拟显示 与其他BL相比，该区域形成两亲性螺旋， 信号的关键结构表明该地区倾向于 形成两亲性螺旋，与其他BL信号相反， 在一个beta turn。该区域还诱导T654，一种已知的底物 蛋白激酶C，提高了其活性的可能性 由磷酸化调节。近期目标是了解 该信号建立和维持EGFR极性的机制。 一个长期的目标是了解导致多囊肾的基因 疾病改变了这一过程，因为非极性EGFR表达是常见的 发现肾囊肿在人类和动物疾病模型中均存在。的 具体的目标将：测试假设，BL分选的细胞质 截短的EGFR严重依赖于特定的氨基酸， BL信号;检验残基K652至A674调节BL的假设 全长EGFR的转运;检验残基K652至 A674调节核内体中的极化分选;并表征 通过鉴定与EGFR相互作用的蛋白质， 残基K652至A674。