Regulation of Actin Dynamics in Airway Smooth Muscle

最近的证据表明，收缩模拟启动了气道平滑肌组织中张力发展所需的肌动蛋白的聚合，然而，新聚合的肌动蛋白的功能和收缩刺激反应中肌动蛋白细丝组装的调节机制尚不清楚。Arp2/3复合体和WASP家族蛋白N-Wasp(神经元Wiskott-Aldrich综合征蛋白)最近被确定为真核细胞中第一个已知的肌动蛋白细丝组装的激活剂，以响应外部信号。本实验的目的是评估一种假说，即对气道平滑肌的收缩刺激会激活N-WASP，进而激活rp2/3复合体，从而启动新的肌动蛋白细丝的形成，这是发展气道平滑肌收缩张力所必需的。此外，肌动蛋白结合蛋白profilin和cofilin被认为是收缩-松弛周期中肌动蛋白动力学调节的重要参与者。调节肌动蛋白聚合量对气道平滑肌组织收缩力学的影响将被确定。这些目标将在一系列实验中解决，这些实验利用我们在体外操纵分离的呼吸道平滑肌组织中内源性和重组蛋白的表达的能力。本研究的具体目的是：1)评价N-WASP和Arp2/3复合体在调节气道平滑肌肌动蛋白聚合和收缩中的作用。2.)评估调节N-WASP激活和气道平滑肌肌动蛋白聚合的信号机制。3)。评估调节呼吸道平滑肌中单体球状(G)肌动蛋白和丝状(F)肌动蛋白平衡的机制，并确定在收缩刺激过程中肌动蛋白动力学的调节如何影响组织的机械性能。这些实验将为肌动蛋白聚合的机制和功能以及它们在调节平滑肌组织收缩特性中的作用提供新的信息。这一信息可能为开发新的治疗靶点以调节哮喘等病理生理条件下的气道平滑肌收缩提供基础。

Recent evidence has shown that contractile simulation initiates the polymerization of actin that is required for tension development in airway smooth muscle tissues; however the function of the newly polymerized actin and the mechanisms for the regulation of actin filament assembly in response to a contractile stimulus are not known. The Arp2/3 complex and that the WASP family protein N-WASp (neuronal Wiskott-Aldrich Syndrome protein), have been recently identified as the first known activators of actin filament assembly in response to external signals in eukaryotic cells. The objectives of the proposed experiments are to evaluate the hypothesis that the contractile stimulation of airway smooth muscle activates N-WASp, which then activates the rp2/3 complex to initiate the formation of new actin filaments necessary for the development of contractile tension in airway smooth muscle. Furthermore, the actin-binding proteins profilin and cofilin are hypothesized to be essential participants in the regulation of actin dynamics during the contraction-relaxation cycle. The effects of modulating the amount of actin polymerization on the contractile mechanics of airway smooth muscle tissues will be determined. These objectives will be addressed in a series of experiments that take advantage of our ability to manipulate the expression of endogenous and recombinant proteins in isolated airway smooth muscle tissues in vitro. The Specific Aims of this proposal are: 1) Evaluate the roles of N-WASp and the Arp2/3 complex in the regulation of actin polymerization and contraction in airway smooth muscle. 2.) Evaluate the signaling mechanisms that regulate the activation of N-WASp and actin polymerization in airway smooth muscle. 3). Evaluate the mechanisms for regulating the balance of monomeric globular (G) actin and filamentous (F) actin in airway smooth muscle, and determine how the modulation of actin dynamics affects the mechanical properties of the tissue during contractile stimulation. These experiments will provide new information regarding the mechanisms and function of actin polymerization and their role in regulating the contractile properties of smooth muscle tissues. This information may provide a foundation for the development of new therapeutic targets for the regulation of airway smooth muscle contraction in pathophysiologic conditions such as asthma.