dictyNews Electronic Edition Volume 40, number 22 September 5, 2014 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 dictyNews, the Dicty Reference database and other useful information is available at dictyBase - http://dictybase.org. Follow dictyBase on twitter: http://twitter.com/dictybase ========= Abstracts ========= PTEN Redundancy: Overexpressing lpten, a Homolog of Dictyostelium discoideum ptenA, the Ortholog of Human PTEN, Rescues All Behavioral Defects of the Mutant ptenA- . Daniel F. Lusche, Deborah Wessels, Nicole A. Richardson, Kanoe B. Russell, Brett M. Hanson, Benjamin A. Soll, Benjamin H. Lin and David R. Soll. Monoclonal Antibody Research Institute and Developmental Studies Hybridoma Bank Department of Biology, The University of Iowa, Iowa City, IA 52242 PloS ONE, in press Mutations in the tumor suppressor gene PTEN are associated with a significant proportion of human cancers. Because the human genome also contains several homologs of PTEN, we considered the hypothesis that if a homolog, functionally redundant with PTEN, can be overexpressed, it may rescue the defects of a PTEN mutant. We have performed an initial test of this hypothesis in the model system Dictyostelium discoideum, which contains an ortholog of human PTEN, ptenA. Deletion of ptenA results in defects in motility, chemotaxis, aggregation and multicellular morphogenesis. D. discoideum also contains lpten, a newly discovered homolog of ptenA. Overexpressing lpten completely rescues all developmental and behavioral defects of the D. discoideum mutant ptenA-. This hypothesis must now be tested in human cells. Submitted by Daniel Lusche [daniel-lusche@uiowa.edu] --------------------------------------------------------------------------- Cell substratum adhesion during early development of Dictyostelium discoideum Marco Tarantola, Albert Bae, Danny Fuller, Eberhard Bodenschatz, Wouter-Jan Rappel and William F. Loomis PLoS ONE' in press Vegetative and developed amoebae of Dictyostelium discoideum gain traction and move rapidly on a wide range of substrata without forming focal adhesions. We used two independent assays to quantify cell-substrate adhesion in mutants and in wild-type cells as a function of development. Using a microfluidic device that generates a range of hydrodynamic shear stress, we found that substratum adhesion decreases at least 10 fold during the first 6 hr of development of wild type cells. This result was confirmed using a single-cell assay in which cells were attached to the cantilever of an atomic force probe and allowed to adhere to untreated glass surfaces before being retracted. Both of these assays showed that the decrease in substratum adhesion was dependent on the cAMP receptor CAR1 which triggers development. Vegetative cells missing talin as the result of a mutation in talA exhibited slightly reduced adhesive properties compared to vegetative wild-type cells. In sharp contrast to wild-type cells, however, these talA mutant cells did not show further reduction of adhesion during development such that after 5 hr of development they were significantly more adhesive than developed wild type cells. In addition, both assays showed that substrate adhesion was reduced in 0hr cells when the actin cytoskeleton was disrupted by latrunculin. Consistent with previous observations, substrate adhesion was also reduced in 0hr cells lacking the membrane proteins SadA or SibA as the result of mutations in sadA or sibA. However, there was no difference in the adhesion properties between wild type AX3 cells and these mutant cells after 6 hr of development, suggesting that neither SibA nor SadA play an essential role in substratum adhesion during aggregation. Our results provide a quantitative framework for further studies of cell substratum adhesion in Dictyostelium. Submitted by Bill Loomis [wloomis@ucsd.edu] --------------------------------------------------------------------------- Defective lysosome maturation and Legionella pneumophila replication in Dictyostelium ArfGAP ACAP-A mutant cells Nathalie Bailo, Pierre Cosson, Steve J. Charette, Valerie E. Paquet, Patricia Doublet, and Fran¨ois Letourneur J Cell Sci , doi:10.1242/jcs.154559 http://jcs.biologists.org/content/early/2014/09/01/jcs.154559.abstract Dictyostelium discoideum ACAP-A is an Arf-GTPase-activating protein involved in cytokinesis, cell migration and actin cytoskeleton dynamics. In mammalian cells, ACAP family members regulate endocytic protein trafficking. Here we explored the function of ACAP-A in the endocytic pathway of D. discoideum. In the absence of ACAP-A, reduced fusion efficacy of post-lysosomes with the plasma membrane resulted in the accumulation of post-lysosomes. Moreover, internalized fluid-phase showed extended intracellular transit time and transfer kinetics of phagocyted particles from lysosomes to post-lysosomes was reduced. Neutralization of lysosomal pH, one essential step in lysosome maturation, was also delayed. Whereas expression of ACAP-A-GFP in acapA- cells restored normal particle transport kinetics, a mutant ACAP-A protein with no GAP activity towards the small GTPase ArfA failed to complement this defect. Together these data support a role for ACAP-A in maturation of lysosomes into post-lysosomes through an ArfA-dependent mechanism. In addition, we reveal that ACAP-A is required for efficient intracellular growth of Legionella pneumophila, a pathogen known to subvert the endocytic host cell machinery for replication. This further emphasizes the role of ACAP-A in the endocytic pathway. Submitted by Francois Letourneur [francois.letourneur@univ-montp2.fr] ============================================================== [End dictyNews, volume 40, number 22]