dictyNews Electronic Edition Volume 39, number 19 July 5, 2013 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 ========= The AmpA protein functions by different mechanisms to influence early cell type specification and to modulate cell adhesion and actin polymerization in Dictyostelium discoideum Hoa N. Cost, Elizabeth F. Noratel and Daphne D. Blumberg Department of Biological Sciences, University of Maryland Baltimore County 1000 Hilltop Circle, Baltimore, Maryland 21250 Differentiation, in press. The Dictyostelium discoideum ampA gene encodes a multifunctional regulator protein that modulates cell-cell and cell-substrate adhesions and actin polymerization during growth and is necessary for correct cell type specification and patterning during development. Insertional inactivation of the ampA gene results in defects that define two distinct roles for the ampA gene during development. AmpA is necessary in a non-cell autonomous manner to prevent premature expression of a prespore gene marker. It is also necessary in a cell autonomous manner for the anterior like cells, which express the ampA gene, to migrate to the upper cup during culmination. It is also necessary to prevent excessive cell-cell agglutination when cells are developed in a submerged suspension culture. Here we demonstrate that a supernatant source of AmpA protein, added extracellularly, can prevent the premature mis-expression of the prespore marker. Synthetic oligopeptides are used to identify the domain of the AmpA protein that is important for preventing cells from mis-expressing the prespore gene. We further demonstrate that a factor capable of inducing additional cells to express the prespore gene marker accumulates extracellularly in the absence of AmpA protein. While secreted AmpA acts extracellularly to suppress prespore gene expression, the effects of AmpA on cell agglutination and on actin polymerization in growing cells are not due to an extracellular role of secreted AmpA protein. Rather, these effects appear to reflect a distinct cell autonomous role of the ampA gene. Finally, we show that secretion of AmpA protein is brought about by elevating the levels of expression of ampA so that the protein accumulates to an excessive level. Submitted by Daphne Blumberg [blumberg@umbc.edu] --------------------------------------------------------------------------- ForC lacks canonical formin activity but bundles actin filaments and is required and multicellular development of Dictyostelium cells Alexander Junemann (1), Moritz Winterhoff (1), Benjamin Nordholz (1), Klemens Rottner (2,3), Ludwig Eichinger (4), Ralph GrŠf (5), and Jan Faix (1) (1) Institute for Biophysical Chemistry, Hannover Medical School, Carl-Neuberg Stra§e 1, 30625 Hannover, Germany; (2) Institute of Genetics, University of Bonn, Karlrobert-Kreiten-Strasse 13, 53115 Bonn, Germany; (3) Helmholtz Centre for Infection Research, Inhoffenstrasse 7, 38124 Braunschweig, Germany; (4) Centre for Biochemistry, Medical Faculty, University of Cologne, Joseph-Stelzmann-Str. 52, 50931 Cologne, Germany; (5) Department of Cell Biology, University of Potsdam, Karl-Liebknecht-Str. 24-25, 14476 Potsdam-Golm, Germany. EJCB, in press Diaphanous-related formins (DRFs) drive the nucleation and elongation of linear actin filaments downstream of Rho GTPase signaling pathways. Dictyostelium formin C (ForC) resembles a DRF, except that it lacks a genuine formin homology domain 1 (FH1), raising the questions whether or not ForC can nucleate and elongate actin filaments. We found that a recombinant ForC-FH2 fragment does not nucleate actin polymerization, but moderately decreases the rate of spontaneous actin assembly and disassembly, although the barbed-end elongation rate in the presence of the formin was not markedly changed. However, the protein bound and crosslinked actin filaments into loose bundles of mixed polarity. Furthermore, ForC is an important regulator of morphogenesis since ForC-null cells are severely impaired in development resulting in the formation of aberrant fruiting bodies. Immunoblotting revealed that ForC is absent during growth, but becomes detectable at the onset of early aggregation when cells chemotactically stream together to form a multicellular organism, and peaks around the culmination stage. Fluorescence microscopy of cells ectopically expressing a GFP-tagged, N-terminal ForC fragment showed its prominent accumulation in the leading edge, suggesting that ForC may play a role in cell migration. In agreement with its expression profile, no defects were observed in random migration of vegetative mutant cells. Notably, chemotaxis of starved cells towards a source of cAMP was severely impaired as opposed to control. This was, however, largely due to a marked developmental delay of the mutant, as evidenced by the expression profile of the early developmental marker csA. In line with this, chemotaxis was almost restored to wild type levels after prolonged starvation. Finally, we observed a complete failure of phototaxis due to abolished slug formation and a massive reduction of spores consistent with forC promoter-driven expression of §-galactosidase in prespore cells. Together, these findings demonstrate ForC to be critically involved in regulation of the cytoskeleton during various stages of development. Submitted by Jan Faix [faix.jan@mh-hannover.de] ============================================================== [End dictyNews, volume 39, number 19]