CSM News Electronic Edition Volume 9, number 5 19 August, 1997 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to CSM-News@nwu.edu. Back issues of CSM-News, the CSM Reference database and other useful information is available at the Dictyostelium Web Page "http://dicty.cmb.nwu.edu/dicty/dicty.html" =========== Abstracts =========== "Proteolysis activated protein kinase in Dictyostelium discoideum" Departamento de Bioquimica and Instituto de Investigaciones Biomedicas CSIC, Facultad de Medicina de la Universidad Autonoma de Madrid, Madrid, Spain Molecular and Cellular Biochemistry., in press Alicia Nunez and Margarita Fernandez In the search for MBP phosphorylating activities in Dictyostelium discoideum, we have found a proteolysis-activated protein kinase. This activity which is distributed between the soluble and the particulate fractions of the cell, uses MBP and histone as substrate and has a molecular mass of 140 kDa as detected in an "in situ" assay. This protein kinase has several features shared by the protein kinase C family, such as substrate specificity and sensitivity to proteolysis, but its molecular mass is much larger than that described for the known protein kinase C isoforms. To better characterize this activity we have studied its sensitivity to several protein kinase C inhibitors and activators. This protein kinase is activated neither by phorbol ester nor by phosphatidylserine or Ca2+. The activity is inhibited by staurosporine and PKC z pseudosubstrate, but is not affected by the specific protein kinase C inhibitor bisindolylmaleimide. These data lead us to propose that proteolytically activated Dictyostelium protein kinase belongs to the recently described protein kinase C-related family. ------------------------------------------------------------------------- Molecular Characterization of a Dictyostelium G-protein a-Subunit Required for Development Maureen A. Brandon1, Susan Voglmaier2 and A. Afshan Siddiqi3 1Wayne State University, Department of Surgery and Department of Biochemistry and Molecular Biology, Detroit, MI 48201, USA 2The Johns Hopkins University, Department of Biological Chemistry, Baltimore, MD 21205, USA 3Wayne State University, Department of Surgery, Detroit, MI 48201, USA Gene, in press Abstract Dictyostelium discoideum utilizes G-protein regulated transmembrane signaling systems to implement its developmental program. This report describes the Dictyostelium G-protein a-subunit, Ga3, and demonstrates that it is required for normal development. Ga3 is the largest of the four completely sequenced Dictyostelium G-protein a-subunits. The difference in size is due to variability in the N-terminal regions. The regions which are affected by the increased size of Ga3 are the Gbg binding region and the helical domain that protects the guanine nucleotide cleft. ga3- mutants created by gene disruption fail to aggregate. However, when treated with exogenous pulses of cAMP that mimic the endogenous cAMP oscillations, they are able to aggregate, but development arrests at the tipped mound stage. This conditional developmental phenotype suggests that Ga3 is required for production of the cAMP signal. --------------------------------------------------------------------- Ga3 REGULATES THE cAMP SIGNALING SYSTEM IN DICTYOSTELIUM Maureen A. Brandon1 and Gregory J. Podgorski2 1Department of Surgery and Department of Biochemistry and Molecular Biology, Wayne State University, Detroit, MI 48201 2Department of Biology and Graduate Program in Molecular Biology, Utah State University, Logan, UT 84322 Molecular Biology of the Cell, in press ABSTRACT The Dictyostelium discoideum developmental program is initiated by starvation and its progress depends on G-protein regulated transmembrane signaling. Disruption of the Dictyostelium G-protein a-subunit Ga3 (ga3- ) blocks development unless the mutant is starved in the presence of artificial cAMP pulses. The function of Ga3 was investigated by examining the expression of several components of the cAMP transmembrane signaling system in the ga3- mutant. Cyclic AMP receptor 1 (cAR11) protein and cyclic nucleotide phosphodiesterase (PDE), phosphodiesterase inhibitor (PDI) and aggregation-stage adenylyl cyclase (ACA) mRNA expression were absent or greatly reduced when cells were starved without exogenously applied pulses of cAMP. However, cAR1 protein and ACA mRNA expression were restored by starving the ga3- cells in the presence of exogenous cAMP pulses. Adenylyl cyclase activity was also reduced in ga3- cells starved without exogenous cAMP pulses compared to similarly treated wild-type cells, but was elevated to a level two-fold greater than wild-type cells in ga3- cells starved in the presence of exogenous cAMP pulses. These results suggest that Ga3 is essential in early development because it controls the expression of components of the transmembrane signaling system. --------------------------------------------------------------------- Lumazine-like fluorescence in a mass of spores of the cellular slime mold, Dictyostelium discoideum Saburo Uchiyama 1, Shin-ichi Nagai 1 and Keizo Maruyama 2 1 Department of Biology, Dokkyo University School of Medicine, Mibu, Tochigi 321-02, Japan 2 Laboratory of Biology, Faculty of Integrated Human Studies, Kyoto University, Kyoto 606, Japan J. Plant Res., in press Abstract Using a fluorospectrophotometer, we examined the fluorescence of a crude preparation from the spore masses of Dictyostelium discoideum. Fluorescence emission spectra and excitation spectra suggested that the fluorescence of the crude preparation was a lumazine-like fluorescence rather than a pterin-like fluorescence. By using a microspectrophotometer, we observed in situ the fluorescence emission of a lumazine-like substance localized only in the spore mass of the fruiting body. --------------------------------------------------------------------- [End CSM News, volume 9, number 5]