The role of Tomosyn-2 in insulin secretion and glucose tolerance

Tomosyn-2在胰岛素分泌和葡萄糖耐量中的作用

基本信息

批准号：
10348695
负责人：
Sushant Bhatnagar
金额：
$ 37.13万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-04-05 至 2024-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10348695
关键词：
Acute Adenosine Monophosphate Adult Affect B-Lymphocytes Beta Cell Binding Binding Proteins Biochemical Biological Assay Blood Glucose Cell Line Cell membrane Cell physiology Chronic Co-Immunoprecipitations Competence Complex Cues Data Defect Development Diabetes Mellitus Diglycerides Disease Disease model Epidemic Event Exocytosis Functional disorder Genes Glucose Goals High Fat Diet Human Hyperglycemia Impairment In Vitro Insulin Insulin Antagonists Insulin Resistance Islets of Langerhans Kinetics Knockout Mice Knowledge Mediating Methods Microscopic Mission Molecular Molecular Target Mus Mutation Non-Insulin-Dependent Diabetes Mellitus Nutritional Outcome Pathway interactions Periodicity Peripheral Persons Phase Phenotype Phosphorylation Phosphorylation Site Physiology Plasma Population Post-Translational Protein Processing Prediabetes syndrome Predisposition Prevention Proteins Public Health Regulation Role SNAP receptor Signal Pathway Structure of beta Cell of islet Testing Therapeutic Thinness Ubiquitin United States United States National Institutes of Health base confocal imaging diabetic forward genetics gain of function mutation genetic approach glucose tolerance hormonal signals hormone regulation impaired glucose tolerance improved innovation insight insulin granule insulin regulation insulin secretion islet knock-down novel nutrition related genetics prevent protein degradation recruit response secretion process soluble NSF attachment protein synaptotagmin syntaxin syntaxin A syntaxin-2 ubiquitin-protein ligase

项目摘要

Type 2 diabetes (T2D) has reached epidemic proportions, with ~9.4% of the US adult population being diabetic, and another 84.1 million have pre-diabetes. It is currently acknowledged that both insulin resistance and b-cell dysfunction are early and essential events in the development of T2D. The formation of the SNARE (Soluble NSF Attachment Protein Receptor) complex is rate limiting for insulin secretion. Our understanding of factors that regulate the formation of the SNARE complex and how they contribute to reduced insulin secretion from b- cells in impaired glucose tolerance is lacking. To this end, by using forward genetics approach, we have identified Tomosyn-2, which is an endogenous inhibitor of insulin secretion and functions by binding to syntaxin. Syntaxin is a key component of the SNARE complex that modulates the fusion of the insulin granules to the plasma membrane for insulin secretion from b-cells. We have discovered that a gain-of-function mutation in the Tomosyn-2 gene led to an increase in islet Tomosyn-2 protein abundance and formation of hypoinsulinemic/ hyperglycemic phenotypes in mice. Increased abundance and/or the functional activity of Tomosyn-2 causes reduction in insulin secretion from human and mouse islets. Thus, the long-term goal is to understand how Tomosyn-2 function in b-cells can be manipulated to improve insulin secretion in impaired glucose tolerance for the treatment and prevention of prediabetes and T2D. The objective of this application is to determine how Tomosyn-2 inhibits insulin secretion from b-cells in the pathophysiology, physiology, and at the molecular level, and how its inhibitory function in b-cells is regulated. Our data show that the improved glucose tolerance in Tomosyn-2-null mice is a direct result of enhanced insulin secretion from pancreatic islets. Further, reduced insulin secretion is observed in islets of mice on a high-fat diet that have elevated Tomosyn-2 protein levels. We have identified phosphorylation sites in response to major b-cell signaling pathways that modulate Tomosyn-2 inhibitory function. Also, E3-ubiquitin ligase, Hrd1 and an insulin granule protein, Syt9 bind and regulate the protein abundance of Tomosyn-2. Our hypothesis is that Tomosyn-2 is a key protein in the exocytotic machinery that regulates SNARE complex-mediated insulin secretion in response to nutritional and genetic cues, and that specific post-translational modifications of Tomosyn-2 increase insulin secretion. To test this hypothesis, we propose three aims: 1) determine the sub-cellular mechanisms by which Tomosyn-2 inhibits insulin secretion, 2) determine Tomosyn-2 phosphorylation regulates its activity on downstream insulin secretion, and 3) determine the role of the Tomosyn-2-binding proteins, Syt9 and Hrd1, in regulating insulin secretion. Outcomes from this project will provide novel information on how b-cells prevent inappropriate insulin secretion, identify the molecular target for the early phase insulin secretion, and insights into the loss in fusion competency of insulin granules during impaired glucose tolerance. Our results will provide fundamental new knowledge of the nutritional and hormonal regulation of the SNARE complex, identifying steps that could be modulated therapeutically in T2D.

2 型糖尿病 (T2D) 已达到流行病的程度，约 9.4% 的美国成年人患有糖尿病，另有 8410 万人患有糖尿病前期。目前公认胰岛素抵抗和b细胞功能障碍是 T2D 发展的早期和重要事件。 SNARE（可溶性 NSF 附着蛋白受体）复合物限制胰岛素分泌的速率。我们对因素的理解调节 SNARE 复合体的形成以及它们如何有助于减少 b- 的胰岛素分泌缺乏糖耐量受损的细胞。为此，通过使用正向遗传学方法，我们确定了 Tomosyn-2 是一种胰岛素分泌的内源性抑制剂，通过与突触融合蛋白结合发挥作用。突触融合蛋白是 SNARE 复合体的关键成分，可调节胰岛素颗粒与血浆的融合 B 细胞分泌胰岛素的膜。我们发现，功能获得性突变 Tomosyn-2 基因导致胰岛 Tomosyn-2 蛋白丰度增加和低胰岛素/ 小鼠的高血糖表型。 Tomosyn-2 丰度和/或功能活性增加导致人类和小鼠胰岛的胰岛素分泌减少。因此，长期目标是了解如何可以操纵 B 细胞中的 Tomosyn-2 功能来改善糖耐量受损的胰岛素分泌糖尿病前期和 T2D 的治疗和预防。该应用程序的目的是确定如何 Tomosyn-2 在病理生理学、生理学和分子水平上抑制 B 细胞的胰岛素分泌，以及它在 b 细胞中的抑制功能是如何调节的。我们的数据表明，葡萄糖耐量的改善 Tomosyn-2缺失小鼠是胰岛胰岛素分泌增强的直接结果。进一步，减少了在高脂肪饮食且 Tomosyn-2 蛋白水平升高的小鼠胰岛中观察到胰岛素分泌。我们已经确定了响应调节 Tomosyn-2 的主要 b 细胞信号通路的磷酸化位点抑制功能。此外，E3-泛素连接酶 Hrd1 和胰岛素颗粒蛋白 Syt9 结合并调节 Tomosyn-2 的蛋白质丰度。我们的假设是 Tomosyn-2 是胞吐机制中的关键蛋白调节 SNARE 复合物介导的胰岛素分泌以响应营养和遗传线索，并且 Tomosyn-2 的特定翻译后修饰可增加胰岛素分泌。为了检验这个假设，我们提出三个目标：1) 确定 Tomosyn-2 抑制胰岛素分泌的亚细胞机制，2) 确定 Tomosyn-2 磷酸化调节其对下游胰岛素分泌的活性，以及 3) 确定 Tomosyn-2 结合蛋白 Syt9 和 Hrd1 在调节胰岛素分泌中的作用。由此产生的结果该项目将提供关于 B 细胞如何防止胰岛素分泌不当、识别分子早期胰岛素分泌的目标，以及对胰岛素颗粒融合能力丧失的见解糖耐量受损期间。我们的结果将提供营养和营养方面的基本新知识 SNARE 复合体的激素调节，确定可以在 T2D 中进行治疗调节的步骤。