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）是维持正常新陈代谢的必需微量营养素。人类和其他动物已经进化出一种高度特异性的机制，用于将膳食钴胺素浓缩在远端小肠肠细胞的吸收表面上。钴胺素内吞作用由 cubilin 介导，cubilin 是顶膜的一种大型多配体受体。伊默斯伦德-格拉斯贝克综合征 (I-GS) 是一种常染色体隐性遗传特征，由于回肠肠上皮细胞和肾近曲小管细胞顶膜未能表达功能性 cubilin，导致选择性钴胺素吸收不良和蛋白尿。 Cubilin 突变会在一个人群中导致人类 I-GS，但其他原因仍有待确定。犬 I-GS 模型中描述了 cubilin 运输至顶膜的遗传缺陷，但 cubilin 被排除为疾病基因，表明未知的辅助活性对于肠和肾上皮细胞中的极化 cubilin 表达至关重要。我们的目标是利用这种独特的、天然存在的动物模型来更好地了解极化上皮细胞中受体表达的细胞生物学。由于缺乏功能候选基因，我们启动了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锁，定义了包含疾病位点的约 9 cM 区域。该提案的具体目标是：1）通过开发新的遗传标记并将其应用于大型远交连锁谱系，以最小化连锁区域，以便识别人类基因组同源区域中的基因子集作为位置候选； 2) 克隆和分析突变的候选位置； 3) 启动致病基因产物的功能分析。将在交互过程中选择用于新标记开发的基因，从显示与疾病基因座紧密连锁的标记的位置，直到发现不表现出与疾病基因座重组的基因标记。 I-GS 基因将通过突变分析进行鉴定，并生成抗体来研究细胞内定位并确定结合伴侣。这些努力的结果预计将为营养吸收和膜受体生物学的新方面打开一扇窗户。