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B 12）是维持正常代谢的必要微量营养素。人类和其他动物已经发展出一种高度特异性的机制，可以将饮食中钴胺素浓缩在远端小肠肠细胞的吸收表面上。钴胺素内吞作用是由Cubilin介导的，Cubilin是顶端膜的大型多物体受体。 Imerslund-Grasbeck综合征（I-GS）是选择性钴胺毒素吸收不良和蛋白尿的常染色体隐性特征，这是由于未在卵母细胞的顶膜中表达功能性纤维蛋白和肾近端肾小管细胞中的功能性cubilin。 Cubilin突变导致一个人群中的人类I-GS，但其他原因仍有待确定。在犬I-GS模型中已经描述了cubilin tocrickin的遗传缺陷，但cubilin被排除为疾病基因，表明未知的辅助活性对于肠道和肾上皮细胞中极化的cubilin表达至关重要。我们的目标是利用这种独特的，天然的动物模型，以更好地了解偏光性上皮中受体表达的细胞生物学。缺乏候选功能基因，我们启动了一种比较位置候选基因方法来确定I-GS疾病基因。发现了与基因标记的显着连接，定义了约9厘米的疾病基因座。该提案的具体目的是：1）通过开发和应用新的遗传标记物来最大程度地减少联系区域，以确定人类基因组同源区域中基因的一部分作为位置候选者； 2）克隆并分析位置候选突变； 3）开始对引起疾病的基因产物的功能分析。从显示与疾病基因座紧密联系的标志物的位置，直到发现与疾病基因座没有重新组合的基因标记，将选择基因进行新标记的发展。 I-GS基因将通过突变分析鉴定，并将生成抗体来研究细胞内定位并确定结合伴侣。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个窗口。