CSM News Electronic Edition Volume 7, number 4 August 17, 1996 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to CSM-News@worms.cmb.nwu.edu. Back issues of CSM-News, the CSM Reference database and other useful information is available by anonymous ftp from worms.cmb.nwu.edu [165.124.233.50], via Gopher at the same address, or by World Wide Web at the URL "http://worms.cmb.nwu.edu/dicty.html" =========== Abstracts =========== The role of the cortical cytoskeleton: F-actin crosslinking proteins protect against osmotic stress, ensure cell size, cell shape and motility, and contribute to phagocytosis and development Francisco Rivero, Bernd K=F6ppel*, Barbara Peracino1, Salvatore Bozzaro1, Florian Siegert2, Cornelis J. Weijer2#, Michael Schleicher3, Richard Albrecht and Angelika A. Noegel4 Max-Planck-Institut fur Biochemie, Am Klopferspitz 18a, D-82152 Martinsried, Germany, 1Dipartimento di Scienze Cliniche e Biologiche, Ospedale S. Luigi Gonzaga, Regione Gonzole 10, 10043 Orbassano-Torino, Italy, 2Zoologisches Institut, Ludwig-Maximilians-Universitat, Luisenstrasse 14, 80333 Munchen, 3Institut fur Zellbiologie, Ludwig-Maximilians-Universitat, Schillerstrasse 42, 80336 Munchen, Germany J. Cell Sci., in press SUMMARY We generated Dictyostelium double mutants lacking the two F-actin crosslinking proteins a-actinin and gelation factor by inactivating the corresponding genes via homologous recombination. Here we investigated the consequences of these deficiencies both at the single cell level and at the multicellular stage. We found that loss of both proteins severely affected growth of the mutant cells in shaking suspension, and led to a reduction of cell size from 12 um in wild type cells to 9 um in mutant cells. Morover the cells did not exhibit the typical polarized morphology of aggregating Dictyostelium cells but had a more rounded cell shape, and also exhibited an increased sensitivity towards osmotic shock and a reduced rate of phagocytosis. Development was heavily impaired and never resulted in the formation of fruiting bodies. Expression of developmentally regulated genes and the final developmental stages that were reached varied however with the substrates on which the cells were deposited. On phosphate buffered agar plates the cells were able to form tight aggregates and mounds and to express prespore and prestalk cell specific genes. Under these conditions the cells could perform chemotactic signalling and cell behavior was normal at the onset of multicellular development as revealed by time-lapse video microscopy. Double mutant cells were motile but speed was reduced by approximately 30% as compared to wild type. These changes were reversed by expressing the gelation factor in the mutant cells. We conclude that the actin assemblies that are formed and/or stabilized by both F-actin crosslinking proteins have a protective function during osmotic stress and are essential for proper cell shape and motility. ------------------------------------------------------------------- RtoA links initial cell type choice to the cell cycle in Dictyostelium Salli A. Wood, Robin R. Ammann, Debra A. Brock, Lily Li, Timothy Spann, and Richard H. Gomer Howard Hughes Medical Institute, Department of Biochemistry and Cell Biology, Rice University, Houston, TX 77251-1892 Development, in press Abstract In Dictyostelium initial cell type choice is correlated with the cell-cycle phase of the cell at the time of starvation. We have isolated a mutant, ratioA (rtoA), with a defect in this mechanism that results in an abnormally high percentage of prestalk cells in both aggregates and cells starved at low density. The rtoA gene has been cloned and sequenced and encodes a novel 39.8 kD protein with 10 repeats of a 11 aa serine-rich sequence, possible N-terminal transmembrane and ATP/GTP binding domains, and a potential tyrosine phosphorylation site. The cell-cycle length and percentage of cells in M, S, and G1 phases are normal in rtoA cells, indicating that the increase in prestalk cells is not caused by a change in the cell cycle. In the wild-type, prestalk cells differentiate from one sister of each sister pair of cells in S or early G2-phase at starvation and prespore cells from one sister of pairs of cells in late G2 or M-phase at starvation. In rtoA mutants, both prestalk and prespore cells originate randomly from cells in any phase of the cell cycle at starvation. As in wild-type, the sister cells of the differentiated rtoA cells were null. These data suggest that RtoA does not participate in asymmetric cell division but is involved in the pathway that monitors cell-cycle phase at the time of starvation and uses this information to select initial cell-type. --------------------------------------------------------------------- [End CSM News, volume 7, number 4]