Spatiotemporal regulation of beta adrenoceptor signaling in cardiacmyocytes

心肌细胞中β肾上腺素受体信号传导的时空调节

基本信息

批准号：
8204912
负责人：
YANG K XIANG
金额：
$ 17.43万
依托单位：
UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-02-01 至 2012-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8204912
关键词：
A kinase anchoring protein Adenoviruses Adrenergic Agents Adrenergic Receptor Agonist Animal Model Animals Arrestins Binding Biological Models Biosensor Calcium Signaling Cardiac Cardiac Myocytes Cell membrane Cell surface Code Complex Cyclic AMP Cyclic AMP-Dependent Protein Kinases Data Depressed mood Diffusion Disease Dissociation Down-Regulation Fluorescence Resonance Energy Transfer GRK Gene Transfer Genes Goals Health Heart Heart Hypertrophy Heart failure Measurement Mus Muscle Cells Myocardial Myocardium PDE4D3 Pathway interactions Patients Performance Phosphorylation Physiological Protein Isoforms Proteins Rattus Regulation Role Second Messenger Systems Series Signal Pathway Signal Transduction Staging Stress System Therapeutic Time United States adrenergic arrestin 2 base constriction in vivo mutant novel phosphoric diester hydrolase receptor receptor coupling response second messenger spatiotemporal therapeutic target

项目摘要

Summary Our long term goal is to understand mechanisms that govern spatiotemporal regulation of cAMP/PKA signaling in cardiac myocytes under physiological and pathophysiological conditions, and their implication in cardiac therapy. During heart failure, down-regulation of the b adrenergic receptor (AR) takes place; however downstream alterations in the pathway, i.e. regulation of substrate phosphorylation by cAMP- dependent protein kinase A (PKA), may precede the down-regulation. The major hypothesis here is that spatiotemporal propagation of cAMP/PKA signaling controlled by bAR subtype-associated phosphodiesterase (PDE) isoforms and arrestin-dependent sequestration of PDEs provides novel mechanism on regulating cardiac response to adrenergic stimulation. We will use both functional measurement of myocyte contraction and real-time measurement of cAMP/PKA activities under bAR subtype regulation to uncover the mechanism underlying functional regulation of intracellular propagation of bAR signaling from the cell surface to intracellular compartments in cardiac myocytes. Aim 1. To characterize the effects of subtype-specific bAR/PDE4D complexes on spatiotemporal cAMP/PKA signaling propagation in cardiac myocytes. We will characterize the association of PDE4Ds with bAR subtypes in different complexes, mechanisms of agonist-dependent dissociation of PDE4Ds from the complexes (via phosphorylation by PKA and/or GRK), and the effects of PDE4Ds on cAMP/PKA activities in different cellular compartments, substrate phosphorylation, calcium signaling, and cardiac myocyte contraction. Aim 2. To characterize the mechanism(s) by which barrestin 1 and 2 control spatiotemporal cAMP/PKA propagation during bAR stimulation in cardiac myocytes. We will examine the association of PDE4D isoforms with bAR subtype-activated arrestins and PKA in cardiac myocytes, the differential roles of arrestin 2 and 3 in controlling bAR subtype-induced cAMP/PKA activities in different cellular compartments, substrate phosphorylation, calcium signaling, and cardiac myocyte contraction. We will also examine the interactive effects of b1 and b2AR on the spatiotemporal propagation of cAMP signaling. Aim 3. To investigate the alteration(s) of bAR/PDE4D complexes in controlling spatiotemporal cAMP/PKA signaling propagation in cardiac myocytes from failing hearts. We will examine the integrity of bAR/PDE4D complexes in myocytes isolated from transverse aortic constriction (TAC)-induced cardiac hypertrophy and heart failure rats, and whether altered bAR association with PDE4D isoforms contributes to depressed spatiotemporal cAMP signal propagation and decreased cardiac contractile responses. We will attempt to selectively inhibit a specific PDE4D isoform by adenovirus gene transfer of PDE4D mutants into myocardium of TAC-treated animals, and examine cardiac performance in vivo.

概括我们的长期目标是了解控制 cAMP/PKA 时空调节的机制生理和病理生理条件下心肌细胞的信号传导及其在心肌细胞中的意义心脏治疗。心力衰竭期间，b 肾上腺素受体 (AR) 会发生下调；然而，该途径的下游变化，即 cAMP- 调节底物磷酸化依赖性蛋白激酶 A (PKA) 可能先于下调。这里的主要假设是 bAR 亚型相关控制的 cAMP/PKA 信号的时空传播磷酸二酯酶 (PDE) 亚型和抑制蛋白依赖性 PDE 隔离提供了新的调节心脏对肾上腺素能刺激反应的机制。我们将使用这两种功能 bAR下肌细胞收缩的测量和cAMP/PKA活性的实时测量亚型调节揭示细胞内增殖功能调节的潜在机制 bAR 信号从细胞表面到心肌细胞内的细胞内区室。目标 1. 至表征亚型特异性 bAR/PDE4D 复合物对时空 cAMP/PKA 信号传导的影响在心肌细胞中传播。我们将描述 PDE4D 与 bAR 亚型的关联不同的复合物，PDE4Ds从复合物中激动剂依赖性解离的机制（通过 PDE4Ds 对不同细胞中 cAMP/PKA 活性的影响细胞区室、底物磷酸化、钙信号传导和心肌细胞收缩。目的 2. 表征 barrestin 1 和 2 控制时空 cAMP/PKA 的机制 bAR 刺激心肌细胞期间的传播。我们将检查 PDE4D 的关联心肌细胞中 bAR 亚型激活的抑制蛋白和 PKA 的异构体，抑制蛋白的不同作用 2和3在不同细胞区室中控制bAR亚型诱导的cAMP/PKA活性，底物磷酸化、钙信号传导和心肌细胞收缩。我们还将检查 b1 和 b2AR 对 cAMP 信号传导时空传播的相互作用。目标 3. 至研究 bAR/PDE4D 复合物在控制时空 cAMP/PKA 信号传导中的改变来自衰竭心脏的心肌细胞的增殖。我们将检查 bAR/PDE4D 的完整性从横主动脉缩窄 (TAC) 诱导的心脏肥大中分离出的心肌细胞中的复合物心力衰竭大鼠，以及改变 bAR 与 PDE4D 亚型的关联是否会导致抑郁时空 cAMP 信号传播和心脏收缩反应降低。我们将尝试通过将 PDE4D 突变体的腺病毒基因转移到 TAC 处理的动物的心肌，并检查体内心脏性能。