OKHSC COBRE: THE ROLE OF THE UBIQUITIN SYSTEM IN RETINAL DEGENERATION

OKHSC COBRE：泛素系统在视网膜变性中的作用

基本信息

批准号：
8360280
负责人：
Scott M Plafker
金额：
$ 16.48万
依托单位：
UNIVERSITY OF OKLAHOMA HLTH SCIENCES CTR
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-07-01 至 2012-09-09
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8360280
关键词：
Angiotensin Type 1a Receptor Animal Model Animals Binding Biological Models Cell Cycle Cell model Centers of Research Excellence Complement Complex Complications of Diabetes Mellitus Diabetes Mellitus Diabetic Nephropathy Disease End stage renal failure Event Funding Genetic Glucose Grant Histologic Hyperglycemia Hypertrophy Incidence Kidney Kidney Failure Knockout Mice Learning Maps Mentors Methods National Center for Research Resources Oklahoma Physiological Principal Investigator Rattus Research Research Infrastructure Resources Retinal Degeneration Rodent Model Role S Phase Signal Transduction Source Streptozocin System Tubular formation Ubiquitin Ubiquitin-Conjugating Enzymes United States National Institutes of Health cost design diabetic hemodynamics inhibitor/antagonist mesangial cell response tissue/cell culture tool

项目摘要

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Diabetic nephropathy is a major complication of diabetes and the leading cause of end-stage renal disease. The earliest morphological changes accompanying diabetic nephropathy are hemodynamic and structural changes in the renal glomerular and tubular compartments. At the cellular level, hypertrophy of the mesangial cells is a hallmark of these early changes. Multiple lines of experimental evidence have revealed a prominent role for the interplay of the angiotensin II receptor type 1A (AT1 A) and p27Kip1 in diabetic mesangial cell hypertrophy. p27Kip1 is a cell cycle inhibitor that normally must be degraded to enable progression from G1 to S phase. During hyperglycemia, p27Kip1 is inappropriately stabilized through signaling events from the AT1 A. An effector of the AT1A called RCBTB1 is required for the induction of cellular hypertrophy, but the function of RCBTB1 and how it contributes to hypertrophy following hyperglycemia are unknown. We have discovered that RCBTB1 is a component of the ubiquitin system and that the same domain of RCBTB1 that binds to the AT1A also interacts with UBE2E3, a ubiquitin conjugating enzyme. Our hypothesis is that UBE2E3 and RCBTB1 modulate the AT1A in a ubiquitin-dependent fashion to contribute to p27Kip1-dependent, mesangial cell hypertrophy. Our model system is rat mesangial cells, a well-characterized glomerular cell model that recapitulates the hypertrophy observed in diabetic nephropathy. Our specific aims are: (1) to biochemically map the complexes formed between the AT1 A, RCBTB1, and UBE2E3, and (2) to determine if disrupting the expression/function of RCBTB1 and UBE2E3 alters p27Kip stabilization and cellular hypertrophy in response to high glucose. These studies will be done in tissue culture cells and complemented by studies in an STZ-rodent model of diabetic nephropathy. Kidneys from these animals will be analyzed histologically, immunohistologically, and biochemically for hypertrophy and p27Kip1 accumulation. To complement these aims, we will generate mice null for UBE2E3 or RCBTB1. These animals will provide us with powerful tools to study UBE2E3 and RCBTB1 by genetic and physiological methods in validated animal models of diabetic disease. Collectively, the information learned from these studies may contribute to the rational design of new treatment options for diabetic nephropathy. Such therapies are desperately needed as the incidence of diabetes and renal failure escalate

该子项目是利用资源的众多研究子项目之一由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持并且子项目的首席研究员可能是由其他来源提供的，包括其他 NIH 来源。子项目可能列出的总成本代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。糖尿病肾病是糖尿病的主要并发症，也是导致终末期肾病的主要原因。糖尿病肾病伴随的最早的形态学变化是肾小球和肾小管室的血流动力学和结构变化。在细胞水平上，肥大系膜细胞是这些早期变化的标志。多项实验证据揭示了 1A 型血管紧张素 II 受体 (AT1 A) 和 p27Kip1 的相互作用在糖尿病肾小球系膜细胞肥大中发挥着重要作用。 p27Kip1 是一种细胞周期抑制剂，通常必须被降解才能启用从G1期进展到S期。在高血糖期间，p27Kip1 通过来自 AT1 A 的信号事件而异常稳定。诱导细胞肥大需要 AT1A 的效应子 RCBTB1，但 RCBTB1 的功能以及它如何促进高血糖后细胞肥大尚不清楚。我们发现 RCBTB1 是泛素系统的一个组成部分，RCBTB1 与 AT1A 结合的相同结构域也与泛素结合酶 UBE2E3 相互作用。我们的假设是，UBE2E3 和 RCBTB1 以泛素依赖的方式调节 AT1A，从而导致 p27Kip1 依赖的系膜细胞肥大。我们的模型系统是大鼠系膜细胞，这是一种特征良好的肾小球细胞模型，可概括在糖尿病患者中观察到的肥大肾病。我们的具体目标是：(1) 对 AT1 A、RCBTB1 和 UBE2E3 之间形成的复合物进行生化图谱，以及 (2) 确定破坏 RCBTB1 和 UBE2E3 的表达/功能是否会改变 p27Kip 稳定性和细胞肥大，以响应高葡萄糖。这些研究将在组织培养细胞中进行，并辅之以糖尿病肾病 STZ 啮齿动物模型的研究。对这些动物的肾脏进行组织学、免疫组织学和生化分析，以了解肥大和 p27Kip1 积累情况。为了实现这些目标，我们将生成 UBE2E3 无效的小鼠或 RCBTB1。这些动物将为我们提供强大的工具，通过遗传和生理方法在经过验证的糖尿病动物模型中研究 UBE2E3 和 RCBTB1。总的来说，从这些研究中获得的信息可能有助于合理设计糖尿病肾病新的治疗方案。随着糖尿病和肾衰竭发病率的上升，迫切需要此类疗法