Regulation of Actin Dynamics in Airway Smooth Muscle

最近的证据表明，收缩模拟启动了肌动蛋白的聚合，这是气道平滑肌组织张力发展所必需的；然而，新聚合的肌动蛋白的功能和肌动蛋白丝组装响应收缩刺激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响应外部信号的肌动蛋白丝组装的第一个已知激活因子。本实验的目的是评估气道平滑肌收缩刺激激活N-WASp的假设，N-WASp随后激活rp2/3复合体，启动气道平滑肌收缩张力发展所必需的新肌动蛋白细丝的形成。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得到解决，这些实验将利用我们在体外分离气道平滑肌组织中操纵内源性和重组蛋白表达的能力。本课题的具体目的是：1)评估N-WASp和Arp2/3复合物在气道平滑肌肌动蛋白聚合和收缩调控中的作用。2.） 评估调节气道平滑肌N-WASp和肌动蛋白聚合激活的信号机制。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时组织的力学特性。这些实验将为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提供新的信息。这一信息可能为哮喘等病理生理条件下气道平滑肌收缩调节的新治疗靶点的开发提供基础。

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.