Transcriptional and Post-translational Dysregulation of SERCA in Type 2 Diabetes

描述(由申请人提供)： 在诊断时，2型糖尿病(T2 DM)患者估计已经丧失了近50%的2细胞功能。此外，像UKPDS这样的大型前瞻性研究表明，胰岛功能在整个疾病过程中不断下降。本申请者的长期目标是确定控制胰腺2细胞中基因表达和细胞功能的生化和表观遗传途径，以确定T2 DM病理状态中调节失调的关键部位。2细胞的胰岛素释放是通过细胞去极化、电压门控钙通道的细胞内钙离子内流和胰岛素颗粒胞吐相结合的钙依赖机制实现的。细胞内钙动态平衡的维持有赖于足够的肌浆网钙ATPase(SERCA)基因表达、蛋白水平和蛋白活性。初步和已发表的研究表明，糖尿病小鼠的胰岛在SERCA mRNA和蛋白质水平上显示出显著的变化。此外，来自心血管文献的数据表明，SERCA在炎症条件下非常容易发生硝化、氧化和降解。这一假设认为，糖尿病的高血糖和细胞因子丰富的环境导致SERCA2基因表达和蛋白活性的双重改变，从而对胰岛钙稳态和胰岛分泌功能产生深远的下游影响。为了验证这一假设，本文提出了两个具体目标。在目标1中，将描述SERCA2基因在2型糖尿病中的转录调控。将进行实验，以确定SERCA2基因反式激活的转录辅助因子，并确定控制SERCA2在胰岛表达的表观遗传途径。在目标2中，将通过设计实验来系统地表征T2 DM中SERCA2活性的翻译后调节，该实验旨在将SERCA2蛋白翻译后不可逆失活修饰的发展与内质网钙水平和胰岛分泌功能相关联。 公共卫生相关性： 2型糖尿病是一种由胰岛素抵抗和胰腺胰岛素分泌细胞进行性功能障碍引起的血糖稳态紊乱。在我们的社会中，2型糖尿病的发病率正在急剧增加。本申请中提出的研究将提高我们对2型糖尿病发病机制的理解，并为该疾病以保护2细胞功能为重点的新治疗策略的开发提供信息。

DESCRIPTION (provided by applicant): At the time of diagnosis, a patient with Type 2 Diabetes Mellitus (T2DM) is estimated to have lost nearly 50% of his or her 2 cell function. Moreover, large prospective studies like the UKPDS, demonstrate that islet function declines continuously throughout the course of this disease. The long-range objective of this applicant is to define the biochemical and epigenetic pathways that govern gene expression and cellular function in the pancreatic 2 cell in order to identify key sites of dysregulation in the pathological state of T2DM. Insulin release from the 2 cell occurs through a Ca2+ dependent mechanism that couples cellular depolarization with intracellular Ca2+ influx from voltage-gated Ca2+ channels, and insulin granule exocytosis. The maintenance of intracellular calcium homeostasis is dependent on adequate sarco-endoplasmic reticulum Ca2+ ATPAse (SERCA) gene expression, protein levels, and protein activity. Preliminary and published studies show that islets from diabetic mice manifest striking changes in SERCA mRNA and protein levels. Further, data from the cardiovascular literature suggests that SERCA is highly susceptible to nitration, oxidation, and degradation under inflammatory conditions. The hypothesis underlying this proposal is that the hyperglycemic and cytokine enriched milieu of diabetes leads to dual insults that alter SERCA2 gene expression and protein activity, which have profound downstream effects on islet calcium homeostasis and islet secretory function. To test this hypothesis, two specific aims are proposed. In Aim 1, the transcriptional regulation of the SERCA2 gene in Type 2 diabetes will be characterized. Experiments will be performed to identify transcriptional cofactors for SERCA2 gene transactivation and to define the epigenetic pathways that govern SERCA2 expression in the islet. In Aim 2, the post-translational regulation of SERCA2 activity in T2DM will be systematically characterized through experiments designed to correlate the development of irreversible inactivating post- translational modifications of SERCA2 protein with endoplasmic reticulum Ca2+ levels and islet secretory function. PUBLIC HEALTH RELEVANCE: Type 2 diabetes mellitus is a disorder of glucose homeostasis caused by a combination of insulin resistance and progressive dysfunction of the insulin producing 2 cells of the pancreas. The incidence of Type 2 diabetes mellitus is increasing dramatically in our society. The research proposed in this application will improve our understanding of the pathogenesis of Type 2 diabetes and inform the development of new treatment strategies for this disease that focus on the preservation of 2 cell function.