Regulation Of Myosins And Myosin Kinases (PAKs)

Heart, Lung, And Blood Institute

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Abstract

(1) Last year we found that chimeras consisting of the head, neck and light chains of Dictyostelium myosin II and the tails of either Acanthamoeba or chicken smooth muscle myosin II had about 25-fold more actin-activated MgATPase activity than unphosphorylated wild-type Dictyostelium myosin II and that their activities were unaffected by phosphorylation of either the regulatory light chain or the tail of the Acanthamoeba chimera. The chimeras made filaments essentially indistinguishable from the filaments of the parent myosin that was the source of the tail. This year we have quantified the ability of these chimeric myosins to support cytokinesis, ConA-capping, motility, chemotaxis and differentiation and development. Only differentiation and development fail to be supported when the chimeras are expressed in Dictyostelium heavy chain null cells. (2) MYOA, the single myosin I of Aspergillus nidulans, is essential for viability of this fungus. We find that MYOA mutants with no more than 1% of the MgATPase activity of wild type MYOA and in vitro motility activity too low to measure support essentially normal fungal morphology with only slight impairment of fungal growth rate and hyphal elongation implying that the essential function of MYOA is structural and not catalytic. This work was done in collaboration with Dr. Gregory May, University of Texas, Anderson Cancer Center. (3) The calmodulin-binding and autoregulatory regions of Acanthamoeba myosin I heavy chain kinase, a p21-activated kinase (PAK), have been determined. (4) We continue to look for structural changes in the tail of Acanthamoeba myosin II resulting from phosphorylation of the regulatory serines at the end of the tail. (5) We are investigating the function of the surface loop on the head of myosin that contains the TEDS-site by characterizing the properties in vivo and in vitro of myosins with point mutants, deletions and chimeras in this region.