BETA1 AND BETA2 ADRENOCEPTOR SIGNALING IN CARDIAC MYOCYTE

描述（由申请人提供）：β肾上腺素受体（β AR）信号传导被精细调节，以响应交感神经系统输入（去甲肾上腺素）和来自肾上腺的肾上腺素（肾上腺素）控制心血管功能。在动物心脏中，两种高度同源的β AR（1 β AR和2 β AR）被儿茶酚胺刺激以增强收缩性和心率。循环儿茶酚胺升高以及心脏1 β AR密度降低对心脏β AR的慢性刺激在临床上与心力衰竭相关。GPCR的主要功能是配体结合和G蛋白偶联。然而，GPCR在体内的特异性和多样化的功能特性涉及用于结合信号复合物中不同配偶体的动态受体构象变化，以及受体信号复合物以细胞类型特异性方式的定位。我们假设去甲肾上腺素和肾上腺素可以激活心肌细胞中1 β AR和2 β AR的不同信号通路。我们选择β-AR基因缺陷小鼠的新生心肌细胞作为模型系统，研究配体-受体相互作用在分化细胞中的功能作用。本提案的目的是（1）表征新生心肌细胞中不同激动剂的1 β AR和2 β AR亚型特异性信号传导，（2）定义心肌细胞中去甲肾上腺素和肾上腺素刺激下1 β AR和2 β AR的细胞和生化特性，（3）表征1 β AR和2 β AR的亚细胞分布，和其它信号分子，（4）鉴定受体相互作用结构域及其靶蛋白，并表征受体信号复合物的形成和激动剂刺激后的稳定性。更好地了解激动剂诱导的AR构象变化的信号复合物的组织可能有助于新的药物设计，并建议在治疗各种心血管疾病的新的临床应用。

DESCRIPTION (provided by applicant): Beta Adrenoceptors (Beta ARs) signaling is finely regulated to control cardiovascular function in response to sympathetic nervous system input (norepinephrine) and to adrenaline (epinephrine) from the adrenal gland. In animal hearts, two highly homologous Beta ARs (1Beta AR and 2Beta AR) are stimulated by catecholamines to enhance contractility and heart rate. Chronic stimulation of cardiac Beta ARs by elevated circulating catecholamines as well as decreased cardiac 1Beta AR density is clinically associated with heart failure. The primary functions attributed to GPCRs are ligand binding and G-protein coupling. However, the specific and diversified functional properties of GPCRs in vivo involve kinetic receptor conformational changes for binding different partners in signaling complexes, and the localization of receptor signaling complexes in cell type- specific manner. We hypothesize that norepinephrine and epinephrine can activate distinct signaling pathways for both 1Beta AR and 2Beta AR in cardiac myocytes. We have chosen the neonatal cardiac myocyte isolated from the Beta AR gene deficienct mice as a model system to study the functional roles of ligand- receptor interactions in differentiated cells. The goals of this proposal are (1) to characterize 1Beta AR and 2 Beta AR subtype-specific signaling by different agonists in neonatal myocytes, (2) define the cellular and biochemical properties of 1Beta AR and 2Beta AR under norepinephrine and epinephrine stimulation in cardiac myocytes, (3) to characterize the subcellular distribution of 1Beta AR and 2Beta AR, and other signaling molecules under norepinephrine and epinephrine stimulation in cardiac myocytes, (4) to identify receptor interaction domains and their targeted proteins, and to characterize the receptor signaling complex formation and stability upon agonist stimulation. A better understanding of agonist-induced AR conformation change for the organization of signaling complexes may facilitate new drug designs and suggest new clinical applications in treating various cardiovascular diseases.