Calcium desensitization in Smooth Muscle

平滑肌在多种生理过程中起着至关重要的作用，尽管 每条平滑肌的基本功能是相同的，收缩和松弛，机械性能和 不同类型的血管对激素、神经递质和药物的反应差异很大。 决定平滑肌收缩特性的因素包括质膜特性， 信号转导蛋白的比例和互补，收缩装置本身的组成。 这些组分的正常混合中的变化被认为是几个 涉及平滑肌的人类疾病，包括高血压、支气管痉挛、性功能障碍、 胃肠道疾病和青光眼。我们的假设是，通过研究分子过程 哪些个体的平滑肌肉通常对刺激有反应将导致更多的选择性疗法来治疗 这些障碍。最近完成的人类和小鼠基因组与先进的 质谱学技术为探索细胞内的信号转导途径提供了新的机会。 在这项提案中，我们将采用蛋白质组学、肌肉生理学、分子生物学、 免疫组织化学和小鼠遗传学研究cGMP的分子机制 依赖于蛋白的蛋白激酶(PKG)通过激活环状GMP来调节平滑肌的松弛。 对不同平滑肌的磷酸蛋白质组的检查确定了早期蛋白质的一个独特的亚群 PKG的目标。在老鼠和人类的基因组中发现了几种，包括一种新的 含有先前未知基序的蛋白质，该基序在平滑蛋白家族中高度保守 肌肉特有的蛋白质。当添加到通透性的平滑肌肉中时，裂缝会产生钙 以磷酸化依赖的方式脱敏和松弛。层序发散度 裂隙基序区域内相互作用的非保守氨基酸表明裂隙和 平滑蛋白可能是一个更大的平滑肌特异蛋白家族的一部分，这些蛋白在 介导cGMP/PKG的作用。为了直接测试这一假设，我们删除了裂隙基因，并 获得无裂隙小鼠。

Smooth muscle plays an essential role in a wide variety of physiological processes, and although the basic function of every smooth muscle is the same, to contract and relax, the mechanical properties and responsiveness to hormones, neurotransmitters and drugs varies greatly between smooth muscle types. Factors that dictate the contractile characteristics of smooth muscle include plasma membrane properties, ratio and compliment of signal transducing proteins, the composition of the contractile apparatus itself. Alteration in the normal blend of these components is thought to underlie the molecular basis of several human diseases that involve smooth muscle including hypertension, bronco spasm, sexual dysfunction, gastrointestinal disorders and glaucoma. It is our hypothesis that by studying the molecular processes by which individual smooth muscles normally respond to stimulation will lead to more selective therapies to treat these disorders. The recent completion of the human and mouse genomes in combination with advanced techniques in mass spectrometry affords new opportunities for probing signal transduction pathways in cells. In this proposal we will employ a unique combination of proteomics, muscle physiology, molecular biology, immuno-histochemistry and mouse genetics to determine the molecular mechanisms by which cGMP through the activation of cyclic GMP dependant protein kinase (PKG) regulates smooth muscle relaxation. Examination of phosphoproteomes of various smooth muscles identified a distinct subset of early protein targets for PKG. Several were identified in the mouse and human genome, including CHASM, a novel protein containing a previously unidentified motif that is highly conserved in the smoothelin family of smooth muscle specific proteins. When added to permeabilized smooth muscles, CHASM causes calcium desensitization and relaxation in a phosphorylation dependant manner. The degree of sequence divergence of the intervening non-conserved amino acids within the CHASM motif region suggests that CHASM and the smoothelins may be part of a larger family of smooth muscle specific proteins that are important in the mediating the actions of cGMP/PKG. To directly test this hypothesis we have deleted the CHASM gene and obtained CHASM null mice.