Mechanism of Protein Accumulation in Renal Hypertrophy

肾脏肥大中蛋白质积累的机制

• 批准号: 8068186
• 负责人: HAROLD A FRANCH
• 金额: $ 32.62万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-05-01 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8068186
• 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; 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

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)：肾脏肥大是糖尿病最早的反应之一，并预测微量白蛋白尿的发展。一个关键领域是肾脏细胞如何积累生长所需的蛋白质。糖尿病肾肥厚包括刺激蛋白质合成和抑制蛋白质降解(PD)。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，这些蛋白质是伴侣介导的自噬(CMA)破坏溶酶体的底物。CMA破坏对肾小管细胞生长至关重要的特定蛋白质。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)信号降低CMA，而H_2O_2则刺激CMA。然而，EGF和H_2O_2都通过Akt蛋白激酶激活信号 这会减少体内的CMA。氧化和生长因子对CMA的影响之间的冲突提出了一些基本问题。1)是什么机制解释了两种相反作用于CMA的刺激在调节CMA的信号上的作用方式？2)氧化和生长因子如何在糖尿病肥厚中控制CMA？3)当存在氧化应激时，蛋白质氧化抑制CMA的结果是什么？我们推测，生长因子和氧化应激对CMA具有拮抗作用，部分是通过与Akt下游调节因子相互作用，传递O类F盒转录因子(FOXO)。在这一假设下，我们的目标是1)研究FoxO信号对H_2O_2和EGF的响应；2)研究H_2O_2和EGF信号在CMA和生长中的相互作用。我们的第二个假设是，在肥厚过程中，氧化蛋白随着CMA被抑制而积累。在这一假设下，我们的目标是3)研究改变CMA如何影响细胞培养中氧化蛋白的生长和积累，以及4)检测通过饮食操作改变CMA时糖尿病肾皮质中的蛋白质分解和蛋白质氧化。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蛋白分解系统组件的技术。我们将测量FOXO信号、蛋白质周转、细胞生长、活性氧的产生和氧化蛋白的积累。最后，我们将测试减少肾脏肥大和糖尿病肾脏损害的改良饮食是否激活CMA并减少氧化蛋白的积累。公共卫生相关性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至关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被称为伴侣介导的自噬过程中被破坏的蛋白质的数量。这项建议研究了同样的过程对于破坏糖尿病造成的导致肾脏疾病的蛋白质可能是如何重要的。