CSM News Electronic Edition Volume 4, number 17 May 13, 1995 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to CSM-News@worms.cmsbio.nwu.edu. Back issues of CSM-News, the CSM Reference database and other useful information is available by anonymous ftp from worms.cmsbio.nwu.edu [165.124.233.50], via Gopher at the same address, or by World Wide Web at the URL "http://worms.cmsbio.nwu.edu/dicty.html" ========== Absracts ========== Single-heade myosin II acts as a dominant-negative mutation in Dictyostelium. C. Geoffrey Burns, Denis A. Larochelle, Harold Erickson, Mary Reedy and Arturo De Lozanne Department of Cell Biology, Duke University Medical Center Durham, NC 27710 Proc. Natl. Acad. Sci. U.S.A., in press. Summary: Conventional myosin II is an essential protein for cytokinesis, capping of cell surface receptors and development of Dictyostelium cells. Myosin II also plays an important role in the polarization and movement of cells. All conventional myosins are double-headed molecules but the significance of this structure is not understood since single-headed myosin II can produce movement and force in vitro. We found that expression of the tail portion of myosin II in Dictyostelium led to the formation of single-headed myosin II in vivo. The resultant cells contain an approximately equal ratio of double- and single-headed myosin II molecules. Surprisingly, these cells were completely blocked in cytokinesis and capping of concanavalin A receptors although development into fruiting bodies was not impaired. We found that this phenotype is not due to defects in myosin light chain phosphorylation. These results show that single-headed myosin II cannot function properly in vivo and that it acts as a dominant-negative mutation for myosin II function. These results suggest the possibility that cooperativity of myosin II heads is critical for force production in vivo. ---------------------------------------------------------------------- ABERRANT cGMP-BINDING ACTIVITY IN NON-CHEMOTACTIC DICTYOSTELIUM DISCOIDEUM MUTANTS Hidekazu Kuwayama1, Gerhard T. Viel1, Shuji Ishida2 and Peter J.M. Van Haastert1* 1. Department of Biochemistry, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands 2. Department of Botany, Faculty of Science, Kyoto University, Kyoto, 606-01, Japan Biochem. Biophys. Acta, in press. Abstract The kinetics of cGMP-binding to the major cGMP-binding activity in Dictyostelium were investigated in ten non-chemotactic mutants (KI mutants; KI-1 10). A wild-type cell contains about 3000 binding sites with a Kd of 1.5 nM. cGMP may dissociate from these binding sites with fast (F-type) or slow (S-type) kinetics, and DNA has been shown to promote the conversion of F- to S-type of cGMP-binding. The ten mutants were placed in four classes, based on equilibrium and non-equilibrium binding properties and the effect of DNA. Class I mutants (KI-1, 3 and 8) have normal cGMP- binding properties. Class II mutants (KI-2, 6 and 7) show increased Kd values but nearly normal Bmax, normal F/S ratio and normal effects of DNA. Class III mutants (KI-4, 5 and 10) have a strongly decreased Kd and increased Bmax, nearly all binding sites are of the S- type and DNA does not affect the binding; apparently these mutants have a cGMP- binding protein locked in the S-form. cGMP-binding in class IV mutant (KI-9) is normal except that the number of binding sites is increased about three-fold. The finding of seven mutants with altered cGMP-binding in ten non-chemotactic mutants suggests that the cGMP-binding activity plays an important role in the chemotactic signal transduction pathway. ---------------------------------------------------------------------- [End CSM News, volume 4, number 17]