Regulation of Actin Dynamics in Airway Smooth Muscle

最近的证据表明，收缩刺激启动了肌动蛋白的聚合，这是气道平滑肌组织中张力发展所必需的;然而，新聚合的肌动蛋白的功能和响应于收缩刺激的肌动蛋白丝组装的调节机制尚不清楚。Arp 2/3复合物和WASP家族蛋白N-WASp（神经元Wiskott-Aldrich综合征蛋白）最近被鉴定为真核细胞中响应于外部信号的肌动蛋白丝组装的第一个已知激活剂。所提出的实验的目的是评估以下假设：气道平滑肌的收缩刺激激活N-WASp，N-WASp然后激活rp 2/3复合物以启动形成气道平滑肌中收缩张力发展所必需的新肌动蛋白丝。此外，肌动蛋白结合蛋白profilin和cofilin被假设为是必不可少的参与者在收缩-舒张周期的肌动蛋白动力学的调节。将确定调节肌动蛋白聚合的量对气道平滑肌组织的收缩力学的影响。这些目标将在一系列实验中解决，利用我们的能力，操纵内源性和重组蛋白在离体气道平滑肌组织中的表达在体外。本研究的具体目的是：1）研究N-WASp和Arp 2/3复合物在调节气道平滑肌肌动蛋白聚合和收缩中的作用。2.）评估调节气道平滑肌中N-WASp活化和肌动蛋白聚合的信号传导机制。3）。评估调节气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，并确定肌动蛋白动力学的调节如何影响收缩刺激期间组织的机械特性。这些实验将提供有关肌动蛋白聚合的机制和功能及其在调节平滑肌组织收缩特性中的作用的新信息。这些信息可能为开发新的治疗靶点提供基础，用于调节哮喘等病理生理条件下的气道平滑肌收缩。

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.