Myosin heavy-chain kinase A from Dictyostelium possesses a novel actin-binding domain that cross-links actin filaments.
Myosin heavy-chain kinase A (MHCK A) catalyses the disassembly of myosin II filaments in Dictyostelium cells via myosin II heavy-chain phosphorylation. MHCK A possesses a 'coiled-coil'-enriched domain that mediates the oligomerization, cellular localization and actin-binding activities of the kinase. F-actin (filamentous actin) binding by the coiled-coil domain leads to a 40-fold increase in MHCK A activity. In the present study we examined the actin-binding characteristics of the coiled-coil domain as a means of identifying mechanisms by which MHCK A-mediated disassembly of myosin II filaments can be regulated in the cell. Co-sedimentation assays revealed that the coiled-coil domain of MHCK A binds co-operatively to F-actin with an apparent K(D) of approx. 0.5 muM and a stoichiometry of approx. 5:1 [actin/C(1-498)]. Further analyses indicate that the coiled-coil domain binds along the length of the actin filament and possesses at least two actin-binding regions. Quite surprisingly, we found that the coiled-coil domain cross-links actin filaments into bundles, indicating that MHCK A can affect the cytoskeleton in two important ways: (1) by driving myosin II-filament disassembly via myosin II heavy-chain phosphorylation, and (2) by cross-linking/bundling actin filaments. This discovery, along with other supporting data, suggests a model in which MHCK A-mediated bundling of actin filaments plays a central role in the recruitment and activation of the kinase at specific sites in the cell. Ultimately this provides a means for achieving the robust and highly localized disruption of myosin II filaments that facilitates polarized changes in cell shape during processes such as chemotaxis, cytokinesis and multicellular development.
Russ
Misty
M
Department of Biology, University of North Carolina at Greensboro, Greensboro, NC 27402, USA.
Croft
Daniel
D
Ali
Omar
O
Martinez
Raquel
R
Steimle
Paul A
PA
D008841:Actin Cytoskeleton / Q000737:chemistry* / Q000378:metabolism*; D000199:Actins / Q000378:metabolism*; D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry* / Q000378:metabolism*; D004023:Dictyostelium / Q000201:enzymology*; D008840:Microfilament Proteins / Q000378:metabolism*; D010449:Peptide Mapping; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D015800:Protozoan Proteins; D014335:Tropomyosin / Q000378:metabolism
D000199:Actins; D008840:Microfilament Proteins; D015800:Protozoan Proteins; D014335:Tropomyosin; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C033142:myosin-heavy-chain kinase