9600923 de Hostos The objective of this project is to define the function of the protein coronin. The Principal Investigator discovered coronin as a component of a complex of actin and myosin prepared in vitro from a cytoplasmic fraction of Dictyostelium discoideum. The protein is associated with the cortical actin cytoskeleton of Dictyostelium cells. It contains a sequence motif known as WD-40 repeats (or beta-transducin), which was first identified in the beta-subunits of trimeric G-proteins. Mutants lacking coronin are impaired in their ability to undergo cytokinesis and ameboid locomotion. A similar protein was subsequently identified by a Japanese group from human and bovine sources, suggesting that the significance of coronin extends beyond merely the biology of slime molds. Dictyostelium is an ideal model system for studying coronin, because the cells perform a variety of actin-based motile processes, and they are amenable to molecular genetic manipulations. Mutant cells will be examined by immunofluorescence, video, and electron microscopy in order to more precisely formulate a hypothesis that coronin is involved in the dynamics of extension or retraction of cortical processes during locomotion. In addition, the coronin mutants will be used to investigate the function and structure of the WD-40 repeats and to attempt to clarify the role of this sequence motif in general. These repeats are being found in a growing number of proteins and are thought to be involved in protein-protein interactions. The ability to genetically manipulate coronin expression in Dictyostelium cells makes this a particularly good system in which to investigate this problem. ***
