Characterisation of the formin protein FHOD1 in striated muscle
Form in homology 2 domain containing protein 1 (FHOD 1) is a diaphanous related formin of the FHOD subclass. In a similar manner to other formins, FHODI has primarily been found to regulate the polymerisation of actin-based structures in cells. In muscle, actin is a major constituent of both skeletal and cardiac myocytes, with the thin filament system of sarcomeres and the cytoskeleton being partly composed of actin. FHOD proteins have previously been highlighted as important regulators of muscle cell biology since FHOD3, a close relative of FHODI was shown to be essential for myofibrillar maintenance. There is little known about the regulation of actin-based structures in muscle cells. We therefore aimed to characterise FHODI and probe into its involvement in myofibrillar and cytoskeletal regulation. In this study of FHOD 1 we addressed various aspects of the protein including its expression pattern and localisation, function, regulation, and novel interacting partners. Insight into the expression pattern of FHODI was gained by examining relative protein levels in different tissues, including both healthy and diseased heart. Subcellular localisation was addressed in a number of fluorescence microscopy experiments through antibody localisation studies in muscle tissue and cultured cells and through transient transfection of GFP tagged constructs. Expression of GFP tagged fragments and mutants helped to delineate the functional distribution of the FHODI molecule in cells. The function of the protein was further probed via molecular knockdown by RNAi and by looking at the capacity of FHOD 1 to polymerise actin in cells. The role of form ins in the heart was more broadly addressed by drug inhibition of their actin polymerising capacity. Previous studies have suggested that the Rho family of small GTPases as well as the Src kinases regulate FHOD 1
Thesis, Dissertation, English, 2015