Dicty News Electronic Edition Volume 25, number 7 September 30, 2005 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to dicty@northwestern.edu or by using the form at http://dictybase.org/db/cgi-bin/dictyBase/abstract_submit. Back issues of Dicty-News, the Dicty Reference database and other useful information is available at dictyBase - http://dictybase.org. ============= Abstracts ============= Ammonium transporter C of Dictyostelium discoideum is required for correct prestalk gene expression and for regulating the choice between slug migration and culmination Janet H. Kirsten, Yanhua Xiong, Andrew J. Dunbar, Meena Rai, Charles K. Singleton * Department of Biological Sciences, Vanderbilt University, VU Station B 351634, Nashville TN 37235-1634, USA Developmental Biology, in press. Ammonium transporter C (AmtC) is one of three transporters in Dictyostelium that have been proposed to regulate entry and exit of ammonia in a cell type dependent manner and to mediate ammonia signaling. Previous work demonstrated that disruption of the amtC gene results in a slugger phenotype in which the cells remain as migrating slugs when they should form fruiting bodies. More detailed studies on the null strain revealed that differentiation of prestalk cell types was delayed and maintenance of prestalk cell gene expression was defective. There was little orno expression of ecmB, a marker for the initiation of culmination. Normal expression of CudA, a nuclear protein required for culmination, was absent in the anterior prestalk zone. The absence of CudA within the tip region was attributable to the lack of adenylyl cyclase A mRNA in the slug tips. Disruption of the histidine kinase gene dhkC in the amtC null strain restored STATa and CudA expression and the ability to culminate. The results suggest that the lack of nuclear t ranslocation of STATa results from low cAMP due to a misregulated and overactive DhkC phosphorelay in the amtC null strain. Submitted by: Charles Singleton [charles.k.singleton@vanderbilt.edu] ----------------------------------------------------------------------------- Role of calcium-dependent actin-bundling proteins: characterization of Dictyostelium mutants lacking fimbrin and the 34 kDa protein Claudia Pikzack(1), Josef Prassler(2), Ruth Furukawa(3), Marcus Fechheimer(3) and Francisco Rivero(1) (1) Zentrum fŸr Biochemie and Zentrum fŸr Molekulare Medizin, Medizinische FakultŠt, UniversitŠt zu Kšln, Joseph-Stelzmann-Str. 52, 50931 Kšln, Germany (2) Max-Planck-Institut fŸr Biochemie, Am Klopferspitz 18a, 82152 Martinsried, Germany (3) Department of Cellular Biology, University of Georgia, Athens, GA 30602, USA Cell Motility and the Cytoskeleton, in press Dictyostelium discoideum two abundant proteins display calcium-regulated bundling activity, fimbrin and the 34 kDa protein (ABP34). Using a GFP fusion we observed transient localization of fimbrin at the phagocytic cup and macropinosomes. The distribution of truncated constructs encompassing t he EF hands and the first actin-binding domain (EA1) or both actin-binding domains devoid of EF hands (A1A2) was indistinguishable from that of the full length protein. The role of fimbrin and a possible functional overlap with ABP34 was investigated in fim- and double 34-/fim- mutants. Except for a moderate cell size defect, fim- mutants did not show defects in growth, endocytosis, exocytosis and chemotaxis. Double mutants were characterized by a small cell size and a defect in morphogenesis resulting in small fruiting bodies and a low spore yield. The cell size defect could not be overcome by expression of fimbrin fragments EA1 or A1A2, suggesting that both bundling activity and regulation by calcium are important. Induction of filopod formation in 34-/fim- cells was not impaired, indicating that both proteins are dispensable for this process. We searched in the Dictyostelium genome database for fimbrin-like proteins that could compensate for the fimbrin defect and identified three unconventional fimbrins and two more proteins with actin-binding domains of the type present in fimbrins. Submitted by: Francisco Rivero [francisco.rivero@uni-koeln.de] ----------------------------------------------------------------------------- Specific host genes required for the killing of Klebsiella bacteria by phagocytes. Mohammed Benghezal, Marie-Odile Fauvarque, RŽgis Tournebize, Romain Froquet, Anna Marchetti, Evelyne Bergeret, Bernard Lardy, GŽrard Klein, Philippe Sansonetti, Steve J. Charette, Pierre Cosson Cellular Microbiology, in press The amoeba Dictyostelium discoideum shares many traits with mammalian macrophages, in particular the ability to phagocytose and kill bacteria. In response, pathogenic bacteria use conserved mechanisms to fight amoebae and mammalian phagocytes. Here we developed an assay using Dictyostelium to monitor phagocyte-bacteria interactions. Genetic analysis revealed that the virulence of Klebsiella pneumoniae measured by this test is very similar to that observed in a mouse pneumonia model. Using this assay, two new host resistance genes (PHG1 and KIL1) were identified and shown to be involved in intracellular killing of K. pneumoniae by phagocytes. Phg1 is a member of the 9TM family of proteins, and Kil1 is a sulfotransferase. The loss of PHG1 resulted in Dictyostelium susceptibility to a small subset of bacterial species including K. pneumoniae. Remarkably, Drosophila mutants deficient for PHG1 also exhibited a specific susceptibility to K. pneumoniae infections. Systematic analysis of several additional Dictyostelium mutants created a two-dimensional virulence array, where the complex interactions between host and bacteria are visualized. Submitted by: Pierre Cosson [Pierre.Cosson@medecine.unige.ch] ============================================================================== [End Dicty News, volume 25, number 7]