Acting Based Regulation of Smooth Muscle Contraction

描述（由申请人提供）：平滑肌收缩的调节在几个方面不同于其他肌肉类型。对这些差异的详细了解可能会导致平滑肌疾病的新疗法，包括高血压、哮喘、消化系统和泌尿生殖系统疾病。与横纹肌相比，平滑肌含有钙调素、钙钙蛋白和其他肌动蛋白结合蛋白，在模型系统中抑制atp酶活性和力的产生。我们已经确定了一种新的肌动蛋白结合蛋白，fessselin，刺激肌动蛋白聚合，抑制肌动蛋白激活肌球蛋白atp酶活性。各种平滑肌肌动蛋白结合蛋白可以通过改变(a)肌动蛋白-肌球蛋白结合，(b)两种肌动蛋白-肌球蛋白复合物之间的转换，(c)细胞信号传导，或(d)细胞骨架的可塑性来发挥作用。我们的第一个目标是确定caldesmon抑制平滑肌收缩的机制。caldesmon的关键问题是：(1)有多少收缩抑制是由肌球蛋白结合的竞争性抑制引起的？这个问题可以通过快速测量caldesmon和myosin与肌动蛋白结合时发生的荧光变化来回答。(2) caldesmon是否也影响过桥动力学？这将通过研究caldesmon对actin-S1荧光核苷酸衍生物释放的抑制来检验。caldesmon-myosin结合的功能是什么？我们将与Gabrielle fitzer博士合作，对缺乏caldesmon部分肌球蛋白结合区域的转基因小鼠进行表征。我们还将与Peter Knight博士合作研究caldesmon-myosin复合物的结构。我们的第二个目标是研究fessselin的性质和功能。将研究fessselin与肌动蛋白结合蛋白synaptopodin家族的关系。我们将研究肌动蛋白-肌球蛋白atp酶活性的抑制机制。fessselin最值得注意的活性是它对肌动蛋白聚合的加速作用。重点将放在这种活动和钙调蛋白的逆转。这些研究将在溶液和平滑肌纤维系统中进行。

DESCRIPTION (provided by applicant): The regulation of contraction of smooth muscle differs in several respects from other muscle types. A detailed understanding of these differences could lead to new therapies for disorders of smooth muscle including hypertension, asthma, and disorders of the digestive system and urogenital system. In contrast to striated muscle, smooth muscle contains caldesmon, calponin and other actin-binding proteins, that inhibit ATPase activity and force production in model systems. We have identified a novel actin binding protein, fesselin that stimulates actin polymerization and inhibits actin activation of myosin ATPase activity. The various smooth muscle actin-binding proteins may function by altering (a) actin-myosin binding, (b) a transition between two actin-myosin complexes, (c) cell signaling, or (d) the plasticity of the cytoskeleton. Our first goal is to finalize the mechanism by which caldesmon inhibits smooth muscle contraction. The key questions with caldesmon are: (1) How much of the inhibition of contraction results from competitive inhibition of myosin binding? This will be answered by using rapid measurements of the fluorescence changes that occur when caldesmon and myosin bind to actin. (2) Does caldesmon also affect cross-bridge kinetics? This will be examined by studying the inhibition of release of fluorescent nucleotide derivatives from actin-S1 by caldesmon. (3) What is the function of caldesmon-myosin binding? We will collaborate with Dr. Gabrielle Pfitzer on the characterization of transgenic mice lacking part of the myosin-binding region of caldesmon. We will also study the structure of the caldesmon-myosin complex in collaboration with Dr. Peter Knight. Our second goal is to investigate the properties and function of fesselin. The relationship of fesselin to the synaptopodin family of actin-binding proteins will be investigated. We will study the mechanism of inhibition of actin-myosin ATPase activity. The most noteworthy activity of fesselin is its large acceleration of actin polymerization. Heavy emphasis will be placed on this activity and its reversal by Cacalmodulin. These studies will be conducted both in solution and in smooth muscle fiber systems.