MOLECULAR CHARACTERIZATION OF PC-1 IN TYPE 2 DIABETES IN MEXICAN-AMERICANS

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。目的：本研究的主要目的是验证PC-1基因参与胰岛素抵抗，从而导致墨西哥裔美国人2型糖尿病(T2 DM)发生的假说。如果观察到PC-1基因内部或附近的多态与胰岛素抵抗之间存在关联，并且可以排除附近的基因，则将继续进行进一步的功能研究。这些包括检测PC-1mRNA的稳定性和表达水平，以及直接测量共表达胰岛素受体和PC-1变体的CHO/IR细胞中的胰岛素受体酪氨酸磷酸化水平。研究计划和方法：将使用最先进的方法对胰岛素受体酪氨酸磷酸化进行定量，从而在分子水平上确定胰岛素抵抗的表型。在骨骼肌胰岛素受体磷酸化正常(N=10)和受损(N=10)的个体中，将通过PC-1基因的直接DNA测序来识别单核苷酸多态(SNPs)。高通量分析将被设计用于筛查等位基因频率为10%的SNPs。选定的SNPs将在来自SAFADS和GENNID研究的更大的墨西哥裔美国人(N=1137)样本中进行筛查。将统计分析SNP基因与胰岛素受体酪氨酸磷酸化、胰岛素抵抗和其他T2 DM表型的相关性。连锁数据将被用于对这些SNP进行尖端的连锁/连锁不平衡分析，以检测可能构成观察到的连锁高峰的功能变异。临床意义：墨西哥裔美国人约占圣安东尼奥人口的52%。这一人群的T2 DM患病率极高，对T2 DM潜在遗传机制的了解有所改善，因此，很好地满足了这一人群的需求。在这个民族中，胰岛素抵抗是严重的，并且在显性糖尿病发作之前就已经确定了。功能研究已经证实，PC-1基因突变会导致胰岛素信号转导受损。这些研究为PC-1参与胰岛素抵抗的直接机制提供了令人信服的证据。在圣安东尼奥家族糖尿病研究(SAFADS)中，该基因位于一个染色体区域，该区域与墨西哥裔美国人的胰岛素抵抗有关。位于6号染色体长臂上的连锁区域产生了与双变量胰岛素抵抗/肥胖表型(瘦素+HOMA IR)连锁的对数赔率(LOD)分数5.8。这是有记录以来针对常见多因素疾病的最强遗传信号之一。在这个高度显著的区域，PC-1是最强的位置候选基因，很可能是该高危人群中胰岛素抵抗/T2 DM的重要易感基因。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. OBJECTIVE: The primary objective of this study is to test the hypothesis that the PC-1 gene is involved in the insulin resistance, and therefore contributes to the development of type 2 diabetes mellitus (T2DM) in Mexican Americans. If an association is observed between polymorphisms within, or near, the PC-1 gene and insulin resistance, and nearby genes can be excluded, further functional studies will be pursued. These include examination of PC-1 mRNA stability and expression levels, and direct measurement of insulin receptor tyrosine phosphorylation levels in CHO/IR cells co-expressing the insulin receptor and PC-1 variants. RESEARCH PLAN AND METHODS: State-of-the-art methods will be used to quantitate insulin receptor tyrosine phosphorylation, thereby defining the insulin resistance phenotype at the molecular level. Single nucleotide polymorphisms (SNPs) will be identified by direct DNA sequencing of the PC-1 gene in individuals with normal (N=10) and impaired (N=10) skeletal muscle insulin receptor phosphorylation. High-throughput assays will be designed to screen SNPs with allele frequencies 10%. Selected SNPs will be screened within a larger population sample of Mexican Americans (N=1137) derived from the SAFADS and GENNID studies. SNP genotypes will be statistically analyzed for association with insulin receptor tyrosine phosphorylation, insulin resistance, and other T2DM phenotypes. Linkage data will be utilized in cutting-edge combined linkage/linkage disequilibrium analyses with these SNPs to detect functional variants, which may underlie the observed linkage peak. CLINICAL RELEVANCE: Mexican Americans represent approximately 52% of the San Antonio population. This population has an extremely high prevalence of T2DM, and an improved understanding of the underlying genetic mechanisms of T2DM, therefore, well serves the needs of this population. In this ethnic group insulin resistance is severe and is well established before the onset of overt diabetes. Functional studies have established that mutations within the PC-1 gene lead to impaired insulin signaling. These studies provide convincing evidence of a direct mechanism for PC-1 involvement in insulin resistance. The gene is located within a chromosomal region that has been genetically linked to insulin resistance in Mexican Americans in the San Antonio Family Diabetes Study (SAFADS). The linked region, on the long arm of chromosome 6, produced a logarithm of the odds (LOD) score of 5.8 for linkage with a bivariate insulin resistance/obesity phenotype (leptin + HOMA IR). This is one of the strongest genetic signals ever recorded for a common multifactorial disease. Within this highly significant region, PC-1 is the strongest positional candidate gene and highly likely to be an important susceptibility gene for insulin resistance/T2DM in this at-risk population.