Functional analysis of a cardiac myosin rod in Dictyostelium discoideum.
Manipulation of the single conventional myosin heavy chain (mhc) gene in Dictyostelium discoideum (Dd) has delineated an essential role for the filament-forming, or light meromyosin (LMM) domain of the myosin molecule in cytokinesis, development, and in the capping of cell surface receptors (see Spudich: Cell Regulation 1:1-11, 1989; Egelhoff et al.: Journal of Cell Biology, 112:677-688, 1991a). In order to assess the functional relationship between sarcomeric and cytoplasmic myosins, a chimeric gene encoding the Dd myosin head and subfragment 2 fused to rat beta cardiac LMM was transfected into both wild-type and Dd mhc null cells. Chimeric myosin was organized into dense cortical patches in the cytoplasm of both wild-type and Dd mhc null cells. Although null cells expressing chimeric mhc at approximately 10% of Dd mhc levels were unable to grow in shaking suspension or to complete development, chimeric myosin was able to rescue capping of cell surface receptors, to associate with filamentous actin, and to localize to the correct subcellular position during aggregation. Deletion of 29 amino acids in the rod corresponding to a previously defined filament assembly competent region eliminated the cortical patches and the posterior localization during chemotaxis. Taken together, these observations suggest that sarcomeric and cytoplasmic myosin rods are functionally interchangeable in several aspects of nonmuscle motility.
LeBlanc-Straceski
J M
JM
Department of Cell Biology, Albert Einstein College of Medicine, Bronx, New York 10461.
Fukui
Y
Y
Sohn
R L
RL
Spudich
J A
JA
Leinwand
L A
LA
D000199:Actins / Q000032:analysis; D000818:Animals; D001483:Base Sequence; D002633:Chemotaxis / Q000502:physiology; D002872:Chromosome Deletion; D004023:Dictyostelium / Q000737:chemistry*; D016368:Frameshift Mutation; D008969:Molecular Sequence Data; D009206:Myocardium / Q000737:chemistry; D015879:Myosin Subfragments / Q000032:analysis / Q000235:genetics / Q000648:ultrastructure*; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics / Q000648:ultrastructure; D014162:Transfection
D000199:Actins; D015879:Myosin Subfragments; D011993:Recombinant Fusion Proteins