Dicty News Electronic Edition Volume 19, number 11 November 23, 2002 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to dicty@northwestern.edu. Back issues of Dicty-News, the Dicty Reference database and other useful information is available at DictyBase--http://dictybase.org. ============= Abstracts ============= A checkpoint of growth/differentiation transition in the cell cycle and its relevance to pattern formation in Dictyostelium development Yasuo Maeda*, Kazunori Sasaki and Aiko Amagai Department of Developmental Biology and Neurosciences, Graduate School of Life Sciences, Tohoku University, Aoba, Sendai 980-8578, Japan Recent Research Developments in Biophysics and Biochemistry, in press (a review article) ABSTRACT In general, growth and differentiation are mutually exclusive, but are cooperatively controlled during the course of development. The cellular slime mold Dictyostelium, a wonderful model organism, grows and multiplies as long as nutrients are supplied, and its differentiation is triggered by starvation. A strict checkpoint (PS-point) from which cells switch growth to differentiation has been specified in the cell cycle of D. discoideum Ax-2 cells. Thus, Ax-2 cells start differentiating from the PS-point in response to starvation, indicating that one needs to combine data on events occurring around the PS-point with those on starvation-induced events to understand the mechanism controlling growth/differentiation transition (GDT). As was expected from the presence of PS-point, the cell s positioning in cell masses and the subsequent cell-type choice occur greatly depending on the cell s phase in the cell cycle at the onset of starvation. Novel and somewhat unexpected functions of mitochondria in cell movement and differentiation are also referred. submitted by: Yasuo MAEDA [ymaeda@mail.cc.tohoku.ac.jp] ----------------------------------------------------------------------------- CONTRASTING ACTIVITIES OF THE AGGREGATIVE AND LATE PDSA PROMOTERS IN DICTYOSTELIUM DEVELOPMENT. Karin E. Weening1, Irene Verkerke-Van Wijk2, Christopher R. Thompson3, Richard H. Kessin4, Gregory J. Podgorski5 and Pauline Schaap1 1School of Life Sciences, University of Dundee, Dundee DD1 5EH, UK 2Department of Biology, University of Leiden, 2333 AL Leiden, The Netherlands 3MRC Laboratory of Molecular Biology, Cambridge, CB2 2QH, UK 4Department of Anatomy and Cell Biology, College of Physicians and Surgeons of Columbia University, New York, NY 10032, USA 5Department of Biology, Utah State University, Logan UT 84322-5305, USA Developmental Biology, in press ABSTRACT Expression of the Dictyostelium PdsA gene from the aggregative (PdA) and late (PdL) promoter is essential for aggregation and slug morphogenesis respectively. We studied the regulation of the PdA and PdL promoters in slugs using labile beta-galactosidase (gal) reporter enzymes. PdL was active in prestalk cells as was also found with stable gal. PdA activity decreased strongly in slugs from all cells except those at the rear. This is almost opposite to PdA activity traced with stable gal, where slugs showed sustained activity with highest levels at the front. PdA was down- regulated after aggregation irrespective of stimulation with any of the factors known to control gene expression. PdL activity was induced in cell suspension by cAMP and DIF acting in synergy. However, a DIF-less mutant showed normal PdL activity during development, suggesting that DIF does not control PdL in vivo. Dissection of the PdL promoter showed that all sequences essential for correct spatio-temporal control of promoter activity are downstream of the transcription start site in a region between -383 and -19 nucleotides relative to the start codon. Removal of nucleotides to position -364 eliminated responsiveness to DIF and cAMP, but normal PdL activity in prestalk cells in slugs was retained. Further 5' deletions abolished all promoter activity. This result also indicates that the induction by DIF and cAMP as seen in cell suspensions is not essential for PdL activity in normal development. submitted by: Pauline Schaap [PSchaap@lsstaff.dundee.ac.uk] ----------------------------------------------------------------------------- Formation of the outer layer of the Dictyostelium spore coat depends on the inner layer protein SP85/PsB Talibah Metcalf, Karen Kelley, Gregory W. Erdos, Lee Kaplan, and Christopher M. West Dept. of Anatomy and Cell Biology, College of Medicine, and Electron Microscopy Core Laboratory, ICBR, University of Florida, Gainesville, FL 32610-0235 USA Microbiology, in press ABSTRACT The Dictyostelium spore is surrounded by a 220-micron thick trilaminar coat that consists of inner and outer electron dense layers surrounding a central region of cellulose microfibrils. In previous studies, a mutant strain (TL56) lacking three proteins associated with the outer layer exhibited increased permeability to macromolecular tracers, suggesting that this layer contributes to the coat permeability barrier. Electron microscopy now shows that the outer layer is incomplete in the coats of this mutant, and consists of a residual regular array of punctate electron densities. The outer layer is also incomplete in a mutant lacking a cellulose-binding protein associated with the inner layer, and these coats are deficient in an outer layer protein and another coat protein. To examine the mechanism by which this inner layer protein, SP85, contributes to outer layer formation, various domain fragments were overexpressed in forming spores. Most of these exert dominant negative effects similar to the deletion of outer layer proteins but one construct, consisting of a fusion of the N-terminal and Cys-rich C1-domain, induces a dense mat of novel filaments at the surface of the outer layer. Biochemical studies show that the C1-domain binds cellulose, and a combination of site-directed mutations that inhibits its cellulose-binding activity suppresses outer layer filament induction. The results suggest that, in addition to a previously described early role in regulating cellulose synthesis, SP85 subsequently contributes a cross- bridging function between cellulose and other coat proteins to organize previously unrecognized structural elements in the outer layer of the coat. submitted by: Chris West [westcm@anatomy.med.ufl.edu] ----------------------------------------------------------------------------- A model for cGMP signal transduction in Dictyostelium in perspective of 25 years of cGMP research Leonard Bosgraaf and Peter J.M. van Haastert Department of Biochemistry, University of Groningen,Nijenborgh 4, 9747 AG Groningen, the Netherlands Journal of Muscle Research and Cell Motility, Special Issue: Dictyostelium Ed. Dietmar J. Manstein, in press Abstract About twenty-five years ago, it was discovered that cGMP is produced intracellularly when cells are exposed to chemoattractants. Shortly hereafter, cGMP-degrading and cGMP-producing activities were found in Dictyostelium lysates, as well as cGMP-binding activity. In the eighties several mutants were isolated displaying aberrant cGMP metabolism as well as altered chemotaxis, suggesting that cGMP plays a role in chemotaxis. Recently we identified the genes encoding guanylyl cyclases, cGMP- phosphodiesterases and cGMP-binding proteins, that probably constitute the main components of the cGMP signal transduction pathway. The encoding proteins appear to be very different from proteins with the same function in metazoa, and therefore their biochemistry is not always easily interpreted. The aim of this review is to discuss the previously obtained biochemical and functional data on cGMP in Dictyostelium in the perspective of the recently identified genes. submitted by: P.J.M.van.Haastert [P.J.M.van.Haastert@chem.rug.nl] ----------------------------------------------------------------------------- cGMP signalling: different ways to create a pathway Jeroen Roelofs1 , Janet L. Smith2 and Peter J.M. Van Haastert1 1, Department of Biochemistry, University of Groningen, Nijenborgh 4, 9747 AG Groningen, the Netherlands 2, Boston Biomedical Research Institute, 64 Grove Street, Watertown, Massachusetts 02472-2829, U.S.A. Trends in Genetics, in press. Abstract A novel cGMP signalling cascade was uncovered in Dictyostelium, a eukaryote that diverged from the lineage to metazoa after plants but before yeast. In both Dictyostelium and metazoa, the ancient cAMP binding (cNB) motif of bacterial CAP has been modified and assembled with other domains into cGMP- target proteins. The domain structures of these cGMP targets, as well as the enzymes responsible for cGMP synthesis and degradation, are entirely different between Dictyostelium and metazoa, suggesting that different cGMP-signalling pathways evolved in these two lineages. submitted by: P.J.M.van.Haastert [P.J.M.van.Haastert@chem.rug.nl] ----------------------------------------------------------------------------- [End Dicty News, volume 19, number 11]