AuthorNovak, Stefanie Whalen Mares
AdvisorGregorio, Carol C.
MetadataShow full item record
PublisherThe University of Arizona.
RightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
AbstractIn striated muscle, contractile activity is dependent on the coordination between the basic contractile unit called the sarcomere and complex cytoskeletal networks. For efficient contractile function, each component is highly regulated to ensure proper expression, assembly and localization within the cell. The molecular mechanisms that govern regulation in muscle cells are still being investigated. In this dissertation, two areas of regulation were investigated: 1) regulation of the heart's conduction system by the RNA-binding protein Fragile X (Chapter 2); and 2) regulation of the sarcomere’s thin filament system by the actin-binding proteins tropomodulin and leiomodin (Chapter 3). The function of Fragile X protein (FraX) in the heart is not well understood. In Drosophila, there is one functionally conserved FraX termed dFmr1, whereas in mammals there are three FraX members with predominate expression of FXR1 in striated muscle. We found that in Drosophila, dFmr1 is required cell autonomously in cardiac cells for regulating heart rate. In mice, cardiac specific loss of FXR1 results in enlarged ventricular lumens and a significant reduction in ejection fraction. Further analyses show FXR1 may influence cardiac membrane potential and calcium homeostasis. To better understand the role of FraX in disease, human and mouse models of dilated cardiomyopathy were examined. We show that FXR1 protein is upregulated and increased expression of FXR1 regulates gap junction remodeling contributing to ventricular tachycardia in mouse hearts. Overall our results support FraX’s essential role in regulating heart function. Another important factor in maintaining proper heart function is regulation of the basic contractile unit – the sarcomere. The actin-binding proteins tropomodulin (Tmod1-4) and leiomodin (Lmod1-3) are considered to be important regulators of the thin filament but their functional properties are still being studied. In striated muscle, Tmod1 and Lmod2 both assemble at the pointed ends of thin filaments but function differently – Tmod1 restricts while Lmod2 elongates thin filament lengths. Given slight differences in structure, we sought to determine the functional significance of their individual domains. For Tmod1, we verify that both tropomyosin-binding sites are necessary for pointed-end assembly and suggest another regulatory site is located within the C-terminal LRR domain. For Lmod2, we confirm the presence of only one functionally significant tropomyosin-binding site and the presence of an N-terminal actin-binding site that influences pointed-end assembly. We also show that tropomyosin-binding affinity of Tmod1 affects its localization, its actin-capping properties, and in skeletal muscle its ability to compete with Tmod3 and Tmod4 for pointed-end assembly. Moreover, we demonstrate endogenous phosphorylation of Tmod1 and Lmod2, suggesting a potential regulatory mechanism, as well as identify potential binding partners that may influence their function in the cell. In summary, the ability of the heart to function properly is dependent on its ability to create an electrical signal and transmit that signal between cells in order to generate muscle contraction. Taken together, these data indicate that FraX contributes to the regulation of membrane potential and gap junction properties, whereas Tmod and Lmod regulate the thin filament – both influencing muscle contraction.
Degree ProgramGraduate College
Cellular and Molecular Medicine