Acute narrowing of the airway lumen is initiated by crossbridge cycling that is mediated by a mechanism of intracellular and intercellular force transmission, propagating contractile force through the interconnected contractile units and smooth muscle cells. However, the molecular mechanisms that mediate force transmission in smooth muscle cells and tissues are largely unknown. Pilot studies indicate that intermediate filament proteins may play a role in mediating force transmission and contraction in airway smooth muscle. The goals of the proposed experiments are to test the hypothesis that contractile stimulation of airway smooth muscle may regulate the spatial organization and the connection of intermediate filaments to the membrane, which may mediate intercellular (direct cell-cell connection) and intracellular force transmission and regulate contractile force in airway smooth muscle.
The Specific Aims of this proposal are: 1.) Evaluate the role of the intermediate filament proteins vimentin and desmin in the functional properties of smooth muscle, and determine depolymerization/polymerization, spatial orientation and phosphorylation of vimentin/desmin in airway smooth muscle cells and tissues in response to contractile stimulation. 2.) Assess the role of PAK (p21-activated protein kinase) in the regulation of vimentin and/or desmin phosphorylation, and evaluate the activation of PAK in response to stimulation with various contractile stimuli. 3.) Determine the association of the desmosome-associated protein plakoglobin with vimentin/desmin and evaluate desmosomal formation in tracheal smooth muscle cells under contractile stimulation. These studies will provide insight into the mechanisms of intercellular and intracellular force transmission and the role of intermediate filament proteins in cytoskeletal organization. The knowledge obtained from these studies may disclose new biological targets for the development of more effective pharmacological treatment of pulmonary diseases such as asthma.
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