Smooth muscles are essential for maintaining homeostasis in all chordate animals. In humans, aberrations in smooth muscle contractile responses have been implicated in hypertension, cardiovascular disease and asthma. Elucidation of the cellular and molecular basis of smooth muscle function is, therefore, crucial for the therapy of a number pathophysiological processes. It is now generally accepted that the phosphorylation of smooth muscle myosin by the Ca 2+/calmodulin dependent enzyme myosin light chain kinase is a key regulatory event in the contractile process. Consequently, the experiments described in this proposal are designed to obtain a detailed understanding of the role of myosin phosphorylation/dephosphorylation n regulating smooth muscles. Three series of experiments are proposed in order to do so. In the first two, we will use a combination of physiological, pharmacological and biochemical approaches to investigate smooth muscle contraction. We have used these techniques extensively in the past and this approach has resulted in a number of important papers. The third series of experiments represents a substantial departure for PI from his previous experience. In this experiment we propose to use molecular biology techniques to study smooth muscles. Therefore, funds provided by the RCDA will be used to curtail the PI's teaching and other departmental responsibilities so that he can learn molecular biology by developing an ongoing collaboration with Dr. Rex Chisholm, an expert in molecular biology, on the structure and function of myosin light chains. As his experience with molecular biology techniques increases, the PI will transfer this technology to his own laboratory and apply them to the study of smooth muscle. Thus, the PI's background and expertise will increase in an important discipline during the funding period. Moreover, the application of molecular biology techniques to study the physiology of smooth muscles will result in a detailed understanding of the role of myosin phosphorylation in smooth muscles and, hopefully, improved therapy for smooth muscle-associated diseases.
