Date of Award

8-1-2016

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Field of Study

Cancer Biology

Department

Graduate School of Biomedical Sciences

First Advisor

Julian Borejdo

Second Advisor

Ignacy Gryczynski

Third Advisor

Abbot F. Clark

Abstract

Familial hypertrophic cardiomyopathy (FHC) is the most common cause of sudden cardiac death in young individuals. Molecular mechanisms underlying this disorder are largely unknown; this study aims at revealing how disruptions in actin-myosin interactions can play a role in the pathogenesis of this disorder. Cross-bridge (XB) kinetics and the degree of order were examined in contracting myofibrils from the ex vivo ventricles of transgenic (Tg) mice expressing FHC regulatory light chain (RLC) mutation K104E and Troponin I mutation, R21C. Because the degree of order and the kinetics are best studied when an individual XB makes a significant contribution to the overall signal, the number of observed XBs in an ex vivo ventricle was minimized to 20. Autofluorescence and photobleaching were minimized by using a relatively long-lived red-emitting dye.

In case of K104E, mutated XBs were significantly better ordered during steady-state contraction and during rigor, but the mutation had no effect on the degree of order in relaxed myofibrils. The K104E mutation increased the rate of XB binding to thin filaments and the rate of execution of the power stroke. In case of R21C, differences were investigated in the left (LV) and right ventricle (RV) mutant where it was found that the mutation imposed significant difference in the distribution of angles that actin makes with thin filament axis: during contraction, actin angles from LV were more tightly distributed compared to actin angles from RV. Collectively, the data indicates that the mutation-induced changes in the interaction of myosin with actin during the contraction- relaxation cycle may contribute to altered contractility and the development of FHC. Phenotypic differences of the R21C mutation in the left versus right mouse ventricles, even though both ventricles express the same isoform of the cardiac highlights the importance of functional differences between the two ventricles of cardiac disease.

Comments

Duggal, Divya., Effect of Phosphorylation on Muscle Physiology and Biophysical Characterization of Mutations Responsible for Familial Hypertrophic Cardiomyopathy. Doctor of Philosophy (Biochemistry and Cancer Biology), June, 2016, 186 pp, 17 tables, 43 illustrations. Available worldwide August 2017.

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