Molecular Mechanism of Polarized Cubilin Expression

极化Cubilin表达的分子机制

• 批准号: 6598635
• 负责人: JOHN C FYFE
• 金额: $ 14.54万
• 依托单位: MICHIGAN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-05-01 至 2005-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6598635
• 关键词: apical membrane; biotechnology; cell surface receptors; cobalamin; disease /disorder classification; disease /disorder etiology; disease /disorder model; dogs; functional /structural genomics; gastrointestinal nutrient absorption; gene mutation; genetic disorder; genetic markers; linkage mapping; malabsorption; molecular cloning; molecular genetics; nutrition related tag; protein transport; proteinuria; receptor expression; single nucleotide polymorphism; small intestines; southern blotting; syndrome

DESCRIPTION (provided by applicant): Cobalamin (vitamin B 12) is an essential micronutrient for the maintenance of normal metabolism. Humans and other animals have evolved a highly specific mechanism for concentrating dietary cobalamin on the absorptive surface of distal small intestinal enterocytes. Cobalamin endocytosis is mediated by cubilin, a large multiligand receptor of the apical membrane. Imerslund-Grasbeck syndrome (I-GS) is an autosomal recessive trait of selective cobalamin malabsorption and proteinuria due to failure to express functional cubilin in the apical membrane of ileal enterocytes and renal proximal tubule cells. Cubilin mutations cause human I-GS in one population, but other causes remain to be determined. An inherited defect of cubilin trafficking to the apical membrane has been described in a canine I-GS model, but cubilin was excluded as the disease gene, indicating that an unknown accessory activity is essential for polarized cubilin expression in intestinal and renal epithelial cells. Our goal is to take advantage of this unique, naturally-occurring animal model to better understand the cell biology of receptor expression in polarized epithelia. Lacking functional gene candidates, we initiated a comparative positional-candidate gene approach to determine the I-GS disease gene. Significant linkage to a gene marker was found, defining an approximately 9 cM region harboring the disease locus. The specific aims of this proposal are: 1) to minimize the region of linkage by developing and applying new genetic markers to a large outbred linkage pedigree in order to identify a subset of genes in the homologous region of the human genome as positional candidates; 2) to clone and analyze positional candidates for mutations; and 3) to initiate functional analyses of the disease-causing gene product. Genes will be chosen for new marker development in an interative process from the locality of markers showing close linkage to the disease locus until a gene marker is found which exhibits no recombination with the disease locus. The I-GS gene will be identified by mutation analysis, and antibody will be generated to study intracellular localization and determine binding partners. Results of these efforts are expected to open a window onto a new aspect of nutrient absorption and membrane receptor biology.

描述（由申请人提供）：钴胺素（维生素b12）是维持正常新陈代谢所必需的微量营养素。人类和其他动物已经进化出一种高度特异性的机制，将膳食钴胺素集中在远端小肠肠细胞的吸收表面。钴胺素的内吞作用是由cubilin介导的，cubilin是顶膜上的一个大的多配体受体。imerslind - grasbeck综合征（I-GS）是一种常染色体隐性性状，由于回肠肠细胞和肾近端小管细胞的顶膜不能表达功能性立方素而导致选择性钴胺素吸收不良和蛋白尿。Cubilin突变在一个人群中导致人类I-GS，但其他原因仍有待确定。在犬I-GS模型中描述了cubilin向根尖膜运输的遗传缺陷，但cubilin被排除为疾病基因，这表明一种未知的辅助活性对肠和肾上皮细胞中cubilin极化表达至关重要。我们的目标是利用这种独特的、自然发生的动物模型来更好地理解极化上皮中受体表达的细胞生物学。由于缺乏功能性候选基因，我们启动了一种比较位置候选基因的方法来确定I-GS疾病基因。发现与基因标记的显著联系，定义了一个大约9厘米的区域，其中包含疾病位点。本提案的具体目标是：1)通过开发和应用新的遗传标记来最小化连锁区域，以便在人类基因组的同源区域中确定一个基因子集作为位置候选；2)克隆和分析候选突变位点；3)启动致病基因产物的功能分析。从与疾病位点有密切联系的标记开始，在一个相互作用的过程中，基因将被选择用于新的标记开发，直到发现一个与疾病位点没有重组的基因标记。通过突变分析鉴定I-GS基因，并产生抗体研究细胞内定位和确定结合伙伴。这些努力的结果有望为营养吸收和膜受体生物学的新方面打开一扇窗。