Novel Regulation of Renal Function by S-Nitrosylation

S-亚硝基化对肾功能的新调节

基本信息

批准号：
10223283
负责人：
JONATHAN S. STAMLER
金额：
$ 43.38万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-25 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10223283
关键词：
Acute Acute Renal Failure with Renal Papillary Necrosis Animal Model Antioxidants Arginine Biological Models Cell model Coenzyme A Complement Crystallization Data Disease Drug Targeting Endothelium Enzymes Equilibrium Family Glycolysis Grant Injury to Kidney Ischemia Isoenzymes Kidney Knock-out Knockout Mice Mammals Mediating Metabolic Metabolism Modeling Modification Molecular Mus NADP NOS3 gene Nitric Oxide Nitric Oxide Synthase Operating System Oxidoreductase Pharmaceutical Preparations Physiological Post-Translational Protein Processing Protein S Proteins Proximal Kidney Tubules Pyruvate Kinase Refractory Regulation Renal function Renal tubule structure Reperfusion Injury Reperfusion Therapy Role S-Nitrosothiols SKIL gene Sepsis Signal Transduction Sterols Structure Sulfhydryl Compounds System Therapeutic Tubular formation Yeasts base design drug candidate in silico inhibitor/antagonist insight member nanomolar nephrotoxicity novel novel drug class novel therapeutics regenerative renal ischemia small molecule inhibitor small molecule therapeutics urinary tract obstruction

项目摘要

PROJECT SUMMARY/ABSTRACT Nitric oxide (NO), generated in the kidney by renal tubular NO synthase (eNOS), can exert renoprotective effects. However, the mechanisms by which NO does so remain poorly understood. Nitric oxide bioactivity is principally conveyed by S-nitrosylation of proteins, the oxidative modification of protein thiols by NO to form S-nitrosothiols (SNOs). Protein S-nitrosylation is reversibly regulated by enzymatic mechanisms including S-nitrosylases and denitrosylases and accumulating evidence implicates dysregulated S- nitrosylation in disease. Here, we identify a novel enzymatic machinery subserving protein S-nitrosylation in the kidney, including a novel metabolic intermediate S-nitroso-coenzyme A (SNO-CoA) that conveys NO bioactivity and its cognate reductase, SNO-CoA reductase (SCoAR), an enzyme of previously unknown function that is highly expressed in kidney proximal tubules. SCoAR mediates the breakdown of SNO-CoA thereby lowering steady-state levels of SNO-proteins (in equilibrium with SNO-CoA). Knockout of SCoAR in mice (SCoAR-/-) is shown to increase S-nitrosylation within the kidney and to protect against ischemia/reperfusion (I/R) injury, whereas deletion of eNOS (SCoAR-/-/eNOS-/-) abrogates this protection. Thus classic renoprotection by eNOS is identified with the SNO-CoA/SCoAR system. Using metabolic profiling and mass spec (MS)-based SNO-protein identification, we have found that renoprotection by the SNO-CoA/SCoAR system is likely mediated by metabolic reprogramming within the kidney. Mechanistically, our data suggest that pyruvate kinase (PKM2) is a major locus of regulation by the SNO-CoA/SCoAR system during acute kidney injury (AKI). Our grant thus explores the idea that S-nitrosylation of PKM2 is renoprotective through metabolic reprogramming that entails SCoAR-regulated coordination of fuel utilization and antioxidant/regenerative defenses. To lay the groundwork for new therapies, we have also screened for SCoAR inhibitors and successfully developed a new class of drug candidate with nanomolar potency, which will be studied herein. Thus, our discovery not only reveals the first physiological function of the SNO-CoA/SCoAR system in mammals, but promises to open a new chapter in our understanding of AKI with immediate therapeutic implications. !

项目概要/摘要一氧化氮 (NO)，由肾小管 NO 合酶 (eNOS) 在肾脏中产生，可以发挥肾脏保护作用影响。然而，NO 发挥作用的机制仍知之甚少。一氧化氮生物活性主要通过蛋白质的 S-亚硝基化传递，即 NO 对蛋白质硫醇的氧化修饰形成 S-亚硝基硫醇 (SNO)。蛋白质 S-亚硝基化通过酶机制可逆调节包括 S-亚硝基酶和脱亚硝基酶，并且越来越多的证据表明 S- 失调疾病中的亚硝基化。在这里，我们发现了一种新的酶机制，可促进蛋白质 S-亚硝基化肾脏，包括一种新型代谢中间体 S-亚硝基辅酶 A (SNO-CoA)，可传递 NO 生物活性及其同源还原酶，SNO-CoA 还原酶 (SCoAR)，一种以前未知的酶肾近曲小管中高度表达的功能。 SCoAR 介导 SNO-CoA 的分解从而降低 SNO-蛋白的稳态水平（与 SNO-CoA 平衡）。 SCoAR 的敲除小鼠 (SCoAR-/-) 被证明可以增加肾脏内的 S-亚硝基化并防止缺血/再灌注 (I/R) 损伤，而 eNOS (SCoAR-/-/eNOS-/-) 的缺失则消除了这种保护作用。因此，eNOS 的经典肾脏保护作用可通过 SNO-CoA/SCoAR 系统来识别。利用代谢通过分析和基于质谱 (MS) 的 SNO 蛋白鉴定，我们发现肾脏保护作用 SNO-CoA/SCoAR 系统可能是由肾脏内的代谢重编程介导的。从机械上来说，我们的数据表明丙酮酸激酶 (PKM2) 是 SNO-CoA/SCoAR 调节的主要位点急性肾损伤（AKI）期间的系统。因此，我们的资助探索了 PKM2 的 S-亚硝基化是通过代谢重编程来保护肾脏，这需要 SCoAR 调节的燃料协调利用和抗氧化/再生防御。为了为新疗法奠定基础，我们还筛选SCoAR抑制剂并成功开发一类纳摩尔级新候选药物效力，本文将对此进行研究。因此，我们的发现不仅揭示了第一个生理功能哺乳动物中的 SNO-CoA/SCoAR 系统，有望开启我们对 AKI 理解的新篇章具有直接的治疗意义。！