Multiple myosin II heavy chain kinases: roles in filament assembly control and proper cytokinesis in Dictyostelium.
Myosin II filament assembly in Dictyostelium discoideum is regulated via phosphorylation of residues located in the carboxyl-terminal portion of the myosin II heavy chain (MHC) tail. A series of novel protein kinases in this system are capable of phosphorylating these residues in vitro, driving filament disassembly. Previous studies have demonstrated that at least three of these kinases (MHCK A, MHCK B, and MHCK C) display differential localization patterns in living cells. We have created a collection of single, double, and triple gene knockout cell lines for this family of kinases. Analysis of these lines reveals that three MHC kinases appear to represent the majority of cellular activity capable of driving myosin II filament disassembly, and reveals that cytokinesis defects increase with the number of kinases disrupted. Using biochemical fractionation of cytoskeletons and in vivo measurements via fluorescence recovery after photobleaching (FRAP), we find that myosin II overassembly increases incrementally in the mutants, with the MHCK A(-)/B(-)/C(-) triple mutant showing severe myosin II overassembly. These studies suggest that the full complement of MHC kinases that significantly contribute to growth phase and cytokinesis myosin II disassembly in this organism has now been identified.
Yumura
Shigehiko
S
Department of Biology, Faculty of Science, Yamaguchi University, Yamaguchi 753-8512, Japan.
Yoshida
Masashi
M
Betapudi
Venkaiah
V
Licate
Lucila S
LS
Iwadate
Yoshiaki
Y
Nagasaki
Akira
A
Uyeda
Taro Q P
TQ
Egelhoff
Thomas T
TT
D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D048749:Cytokinesis / Q000502:physiology*; D003599:Cytoskeleton / Q000201:enzymology*; D004023:Dictyostelium / Q000201:enzymology* / Q000254:growth & development; D017930:Genes, Reporter; D018995:Myosin Heavy Chains / Q000378:metabolism; D024462:Myosin Type II / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C033142:myosin-heavy-chain kinase; D024462:Myosin Type II; D018995:Myosin Heavy Chains