The biochemical, structural and cellular characterization of myosin-V

Abstract

The myosin superfamily includes about 16 classes of structurally distinct molecules. While the discovery of new classes of myosin is becoming more remote, new members of existing classes are still being identified in many organisms and cell types. The present challenge is to define the molecular and biochemical properties and exact cellular roles of each of these myosins. We are investigating the biochemical, structural and cellular properties of myosin-V, a class of unconventional myosins featuring an extended neck domain consisting of 6 tandem repeats of the IQ-motif, potential binding sites for calmodulin and other light chains, and a structurally distinct tail domain which we suppose to be involved in cellular localization, anchoring and/or other specific functions of this myosin. We are using several approaches including protein purification and biochemical assays, DNA recombinant techniques on the chicken myosin-V sequence, transfection and immunological probes on cultured melanocytes and in the yeast S.cerevisiae, video and confocal microscopy for determining phenotype parameters and immuno-ultrastructural studies on these cells and on subcellular fractions, such as synaptosome fractions. This project is a continuation of our research over the past 15 years. We believe it is fair to say that our laboratories have made a considerable contribution to the fact that myosin-V is one of the best characterized "unconventional" myosins at the present time. The overall questions being addressed in this project are "What are the structural/functional relations between the domain organization and structure of myosin-V and its biochemical properties?" ''What exact role(s) does myosin-V play in the cell and how does this reflect on the dynamics of the cytoskeleton and cellular motility?" We expect to make further contributions to the basic knowledge on the function and regulation of myosin motors in eukaryotic cells, but also there are clear implications for involvement of myosin-V in medical questions. For example, mutations in myosin-V are re1ated to Griscelli's disease, whose sufferers manifest grave disfunctions in their immunological and neurological systems. Also, myosin-V is essential for normal functioning of melanocytes, having effects on pigmentation and quite possibly tumor growth. Thus, our investigation of the molecular and cellular properties of myosin-V has potential relevance and application to the biomedical area. (AU)