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Mechanism of Protein Accumulation in Renal Hypertrophy

肾脏肥大中蛋白质积累的机制

基本信息

批准号：
8306990
负责人：
HAROLD A FRANCH
金额：
$ 32.66万
依托单位：
EMORY UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-05-01 至 2014-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8306990
关键词：
Affect Aging Area Autophagocytosis Blood Vessels Caloric Restriction Cell Culture Techniques Cell Growth Processes Cells Conflict (Psychology)Data Development Diabetes Mellitus Diabetic Nephropathy Diet Disease Epidermal Growth Factor F Box Domain Family Growth Growth Factor Health Hydrogen Peroxide Hypertrophy In Vitro Kidney Kidney Diseases Laboratories Lysosomes Measures Mediating Mediator of activation protein Microalbuminuria Modification Molecular Chaperones Muscle Nutritional Organ Oxidative Stress Phosphorylation Phosphotransferases Process Production Protein Biosynthesis Protein Isoforms Protein Kinase Proteins Proteolysis Proto-Oncogene Proteins c-akt Reactive Oxygen Species Reagent Reducing diet Regulation Renal function Rodent Model Signal Pathway Signal Transduction Signaling Molecule Signaling Protein Skeletal Muscle Small Interfering RNA Stimulus Stream System Techniques Testing Tissues Tubular formation Ubiquitin Uremia Work base cell growth cell type diabetic expression vector in vivo kidney cell kidney cortex lysosomal proteins multicatalytic endopeptidase complex novel oxidation prevent protein degradation response transcription factor

项目摘要

DESCRIPTION (provided by applicant): Renal hypertrophy is among the earliest responses to diabetes and predicts the development of microalbuminuria. One critical area is how kidney cells accumulate the protein needed for growth. Diabetic renal hypertrophy involves both stimulation in protein synthesis and blockade of protein degradation (PD). In diabetes, our work has shown that the kidney suppresses PD through a lysosomal system leading to the accumulation of proteins that are substrates for destruction in lysosomes by chaperone-mediated autophagy (CMA). CMA destroys particular proteins important for renal tubular cell growth. However, CMA also destroys proteins that have undergone oxidative modification. In vitro, epidermal growth factor (EGF) signaling reduces CMA, while H2O2 stimulates it. However, both EGF and H2O2 activate signaling through the Akt protein kinase which reduces CMA in vivo. These conflicts between oxidative and growth factor effects on CMA raise some fundamental questions. 1) What mechanism explains two stimuli that act oppositely on CMA acting the same way on the signaling that regulates CMA? 2) How do oxidation and growth factors interact in controlling CMA in diabetic hypertrophy? 3) What is the consequence of protein oxidation on suppressing CMA when there is oxidative stress? We hypothesize that growth factors and oxidative stress have antagonistic effects on CMA in part through interactions on a down stream mediator of Akt signaling the class O F box transcription factors (FoxOs). Our Aims under this hypothesis are 1) To examine FoxO signaling in response to H2O2 and EGF and 2) To examine the interaction between H2O2 and EGF signaling on CMA and growth. Our second hypothesis is that oxidized proteins accumulate as CMA is suppressed during hypertrophy. Our Aims under this hypothesis are 3) To examine how altering CMA affects growth and accumulation of oxidized proteins in cell culture and 4) To examine proteolysis and protein oxidation in diabetic renal cortex when CMA is altered by dietary manipulation. To accomplish these objectives, we will use adenoviral expression vectors and siRNA techniques to manipulate molecules in the FoxO signaling pathway and components of the CMA proteolytic system. We will measure FoxO signaling, protein turnover, cell growth, reactive oxygen species production and the accumulation of oxidized proteins. Finally, we will test whether modified diets that reduce renal hypertrophy and diabetic kidney damage activate CMA and reduce accumulation of oxidized proteins. PUBLIC HEALTH RELEVANCE Our laboratory has observed that when kidneys grow in diabetes (hypertrophy), they decrease the breakdown of certain proteins important for growth. Certain signaling molecules in the diabetic kidney reduce the amount of protein destroyed in a process called chaperone-mediated autophagy. This proposal studies how this same process may be important for destroying proteins damaged by diabetes that contribute to kidney disease.

描述（由申请人提供）：肾肥大是糖尿病的最早反应之一，可预测微量白蛋白尿的发生。一个关键领域是肾细胞如何积累生长所需的蛋白质。糖尿病肾肥大涉及蛋白质合成的刺激和蛋白质降解（PD）的阻断。在糖尿病中，我们的工作表明，肾脏通过溶酶体系统抑制PD，导致蛋白质的积累，这些蛋白质是通过伴侣介导的自噬（CMA）在溶酶体中破坏的底物。CMA破坏对肾小管细胞生长重要的特定蛋白质。然而，CMA也会破坏经过氧化修饰的蛋白质。在体外，表皮生长因子（EGF）信号减少CMA，而H2O2刺激CMA。然而，EGF和H2O2都通过Akt蛋白激酶激活信号其在体内减少CMA。氧化和生长因子对CMA的影响之间的这些冲突提出了一些基本问题。1)什么机制解释了两种刺激对CMA的作用相反，对调节CMA的信号作用相同？2)氧化和生长因子在控制糖尿病性肥大的CMA中如何相互作用？3)当存在氧化应激时，蛋白质氧化对抑制CMA的影响是什么？我们假设生长因子和氧化应激对CMA具有拮抗作用，部分是通过Akt信号传导的下游介体F盒类转录因子（FoxOs）的相互作用。我们的目的是1）检测FoxO信号对H2O2和EGF的反应，2）检测H2O2和EGF信号对CMA和生长的相互作用。我们的第二个假设是，氧化蛋白质积累的CMA是在肥大过程中受到抑制。我们的目的是在这个假设下3）检查如何改变CMA影响细胞培养物中氧化蛋白的生长和积累和4）检查当CMA通过饮食操作改变时糖尿病肾皮质中的蛋白水解和蛋白氧化。为了实现这些目标，我们将使用腺病毒表达载体和siRNA，操纵FoxO信号通路中的分子和CMA蛋白水解系统的组分的技术。我们将测量FoxO信号，蛋白质周转，细胞生长，活性氧产生和氧化蛋白质的积累。最后，我们将测试减少肾脏肥大和糖尿病肾损伤的改良饮食是否激活CMA并减少氧化蛋白的积累。我们的实验室观察到，当肾脏在糖尿病（肥大）中生长时，它们会减少某些对生长重要的蛋白质的分解。糖尿病肾脏中的某些信号分子减少了蛋白质在分子伴侣介导的自噬过程中被破坏的数量。这项提案研究了同样的过程如何对破坏糖尿病损害的蛋白质起重要作用，这些蛋白质会导致肾脏疾病。