Investigating the role of SIRT3 in metabolic flexibility and proteostasis in the aging heart

研究 SIRT3 在衰老心脏代谢灵活性和蛋白质稳态中的作用

• 批准号: 10625412
• 负责人: Kenneth M Humphries
• 金额: $ 21.85万
• 依托单位: OKLAHOMA MEDICAL RESEARCH FOUNDATION
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10625412
• 关键词: Acceleration; Acetylation; Adult; Affect; Age; Aging; Carbon; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; Cause of Death; Cell Culture Techniques; Data; Deacetylase; Deuterium Oxide; Enzymes; Event; Exhibits; Fibrosis; Future; Global Change; Glucose; Goals; Half-Life; Heart; Heart Hypertrophy; Homeostasis; Hypertrophy; Individual; Intervention; Knockout Mice; Label; Longevity; Mass Spectrum Analysis; Measures; Metabolic; Metabolism; Mitochondria; Mitochondrial Proteins; Modeling; Modification; Mus; Pathology; Phenotype; Post-Translational Protein Processing; Predisposition; Process; Protein Acetylation; Protein Biosynthesis; Proteins; Proteomics; Role; Sirtuins; Stress; Structure; Testing; Tissues; United States; age related; aged; coronary fibrosis; experimental study; flexibility; healthspan; heart function; heart metabolism; improved; in vivo; mitochondrial dysfunction; model organism; novel; premature; prevent; protein degradation; proteostasis; restoration

Cardiovascular disease is the leading cause of death in the United States and its occurrence dramatically increases with age. Preventing or delaying cardiac aging can therefore have a significant effect on longevity and healthspan. A central goal of this proposal is to explore a novel mechanism by which mitochondria decline with age and contribute to loss of cardiac function. We are focusing on the post-translational modification, acetylation, because it increases with age and the enzyme that reverses this modification, sirtuin-3 (SIRT3), is a known longevity factor that decreases with age. However, if mitochondrial acetylation accelerates cardiac aging remains unresolved and controversial. Our overarching hypothesis is that an increase in acetylation has two effects in the aged heart. First, it causes metabolic inflexibility by directly affecting the activity of key regulatory enzymes. Second, the increase in acetylation affects proteostasis (protein synthesis and degradation homeostasis). Dysfunctional proteostasis in turn contributes to metabolic inflexibility by causing improper synthesis and degradation of modified mitochondrial metabolic enzymes. For this proposal, we generated a cardiomyocyte (CM) specific SIRT3 knockout mice (SIRT3CM-/-). Our preliminary data show that SIRT3CM-/- mice exhibit dramatic hypertrophy, loss of contractile function, fibrosis, metabolic abnormalities, and dysfunctional proteostasis by 10- months. This accelerated model of aging and hyperacetylation will be used to test our hypothesis through the following aims. Aim 1 will determine if the increase in acetylation caused by the loss of SIRT3 affects metabolic flexibility with age. We hypothesize that the loss of SIRT3 causes metabolic inflexibility by affecting the activity of discrete mitochondrial regulatory enzymes that result in the increased reliance on glucose. This aim will determine if the loss of SIRT3 affects metabolic flexibility by measuring cardiac and mitochondrial functions, enzymatic activities, and metabolic flexibility longitudinally in SIRT3CM-/- mice and isolated adult cardiomyocytes. Global acetylation and the acetylation of specific metabolic enzymes will be measured by mass spectrometry. The results of these studies will determine if there is a direct consequence of hyperacetylation on cardiac pathology in the absence of SIRT3. Aim 2 will determine how the loss of SIRT3 affects mitochondrial proteostasis. Acetylation of proteins can affect structure and function, yet little is known regarding its role in global changes in mitochondrial protein quality and turnover. This aim will use deuterium oxide (D2O) labeling and proteomics to determine if the loss of SIRT3 affects mitochondrial protein synthesis and the relative turnover rates of specific metabolic regulatory enzymes. Rescue experiments will be performed by AAV delivery of SIRT3. Mechanistic studies in cell culture will be performed to demonstrate how acetylation affects proteostasis. These will be the first studies to employ in vivo labeling to determine how acetylation affects protein synthesis and turnover. The results will provide an impetus for future projects further defining how acetylation and other carbon stresses affect cardiac aging.

在美国，心血管疾病是导致死亡的主要原因，而且发病率很高