dictyNews Electronic Edition Volume 33, number 5 August 21, 2009 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. ========= Abstracts ========= CP250, a novel acidic coiled coil protein of the Dictyostelium centrosome, affects growth, chemotaxis and the nuclear envelope Rosemarie Blau-Wasser,*† Ursula Euteneuer,‡ Huajiang Xiong,* Berthold Gassen,*† Michael Schleicher,‡ Angelika A. Noegel*† *Center for Biochemistry, Medical Faculty, †Center for Molecular Medicine Cologne (CMMC) and Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, 50931 Köln, Germany, ‡Institute of Anatomy and Cell Biology and Center for Integrated Protein Science (CIPSM), Ludwig-Maximilians-University, 80336 München, Germany Mol. Biol. Cell, in press The Dictyostelium centrosome is a nucleus associated body consisting of a box-shaped core surrounded by the corona, an amorphous matrix functionally equivalent to the pericentriolar material of animal centrosomes which is responsible for the nucleation and anchoring of microtubules. Here we describe CP250 a component of the corona, an acidic coiled coil protein which is present at the centrosome throughout interphase while disappearing during prophase and reappearing at the end of late telophase. Amino acids 756-1148 of the 2,110 amino acids are sufficient for centrosomal targeting and cell cycle dependent centrosome association. Mutant cells lacking CP250 are smaller in size, growth on bacteria is delayed, chemotaxis is altered and development is affected which, in general, are defects observed in cytoskeletal mutants. Furthermore, loss of CP250 affected the nuclear envelope and led to reduced amounts and altered distribution of Sun-1, a conserved nuclear envelope protein that connects the centrosome to chromatin. Submitted by Angelika Nögel [noegel@uni-koeln.de] -------------------------------------------------------------------------------- Role of a Cytoplasmic Dual-Function Glycosyltransferase in O2-Regulation of Development in Dictyostelium Zhuo A. Wang(1), Hanke van der Wel(1), Yusof Vohra(2), Therese Buskas(2), Geert-Jan Boons(2), and Christopher M. West(1) 1Dept. of Biochemistry & Molecular Biology and Oklahoma Center for Medical Glycobiology, University of Oklahoma Health Sciences Center, Oklahoma City, OK  73104  USA; 2Dept. of Chemistry and Complex Carbohydrate Research Center, 315 Riverbend Road, University of Georgia, Athens, GA  30602  USA. J. Biol. Chem., in press In the social amoeba Dictyostelium, a terminal step in development is regulated by environmental O2. Prolyl 4-hydroxylase-1 (P4H1) was previously implicated in mediating the O2 signal, and P4H1-null cells require elevated O2 to culminate. The E3-ubiquitin ligase adaptor Skp1 is a P4H1 substrate, and here we investigate the function of PgtA, a dual function beta3-galactosyltransferase/alpha2-fucosyltransferase that contributes the 2nd and 3rd sugars of the pentasaccharide cap formed on Skp1 hydroxyproline. Although pgtA-null cells, whose Skp1 contains only a single sugar (N-acetylglucosamine or GlcNAc), show wild-type O2-dependence of culmination, cells lacking AgtA, an alpha3-galactosyltransferase required to extend the trisaccharide, require elevated O2 as for P4H1-null cells. Skp1 is the only detectable protein modified by purified PgtA added to pgtA-null extracts. The basis for specificity of PgtA was investigated using native Skp1 acceptor glycoforms and a novel synthetic peptide containing GlcNAcalpha1,4-hydroxy(trans)proline. Cysteine-alkylation of Skp1 strongly inhibited modification by the PgtA galactosyltransferase but not the fucosyltransferase. Furthermore, native and synthetic Skp1 glycopeptides were poorly galactosylated, not processively fucosylated, and negligibly inhibitory, whereas the fucosyltransferase was active toward small substrates. In addition, the galactosyltransferase exhibited an atypical concentration dependence on UDP-galactose. The results provide the first evidence that Skp1 is the functional target of P4H1 in O2-regulation, indicate a gatekeeper function for the beta3-galactosyltransferase in the PgtA-dual reaction, and identify an unexpected P4H1-dependent yet antagonistic function for PgtA that is reversed by AgtA. Submitted by Chris West [Cwest2@ouhsc.edu] -------------------------------------------------------------------------------- dictyExpress: A Dictyostelium discoideum gene expression database with an explorative data analysis web-based interface Gregor Rot*, Anup Parikh*, Tomaz Curk, Adam Kuspa, Gad Shaulsky and Blaz Zupan Faculty of Computer and Information Science, University of Ljubljana, SI-1000 Ljubljana, Slovenia Baylor College of Medicine, Houston, TX 77030, USA * equal contribution BMC Bioinformatics, in press Background Bioinformatics often leverages on recent advancements in computer science to support biologists in their scientific discovery process. Such efforts include the development of easy-to-use web interfaces to biomedical databases. Recent advancements in interactive web technologies require us to rethink the standard submit-and-wait paradigm, and craft bioinformatics web applications that share analytical and interactive power with their desktop relatives, while retaining simplicity and availability. Results We have developed dictyExpress, a web application that features a graphical, highly interactive explorative interface to our database that consists of more than 1000 Dictyostelium discoideum gene expression experiments. In dictyExpress, the user can select experiments and genes, perform gene clustering, view gene expression profiles across time, view gene co-expression networks, perform analyses of Gene Ontology term enrichment, and simultaneously display expression profiles for a selected gene in various experiments. Most importantly, these tasks are achieved through web applications whose components are seamlessly interlinked and immediately respond to events triggered by the user, thus providing a powerful explorative data analysis environment. Conclusions dictyExpress is a precursor for a new generation of web-based bioinformatics applications with simple but powerful interactive interfaces that resemble that of the modern desktop. While dictyExpress serves mainly the Dictyostelium research community, it is relatively easy to adapt it to other datasets. We propose that the design ideas behind dictyExpress will influence the development of similar applications for other model organisms. Submitted by Gad Shaulsky and Blaz Zupan [gadi@bcm.edu] -------------------------------------------------------------------------------- Self-organizing Actin Waves as Planar Phagocytic Cup Structures Günther Gerisch1, Mary Ecke1, Britta Schroth-Diez2, Silke Gerwig2, Ulrike Engel3, Lucinda Maddera4, and Margaret Clarke4 1 Max-Planck-Institut für Biochemie, Am Klopferspitz 18, D-82152 Martinsried, Germany. 2 Max-Planck-Institut für molekulare Zellbiologie und Genetik, Pfotenhauerstrasse 108, D-01307 Dresden, Germany. 3 Nikon Imaging Center der Universität Heidelberg, Bioquant BQ 0004, Im Neuenheimer Feld 267, D-69120 Heidelberg, Germany. 4 Program in Genetic Models of Disease, Oklahoma Medical Research Foundation, Oklahoma City, OK 73121, USA. Cell Adhesion & Migration, in press Actin waves that travel on the planar membrane of a substrate-attached cell underscore the capability of the actin system to assemble into dynamic structures by the recruitment of proteins from the cytoplasm. The waves have no fixed shape, can reverse their direction of propagation, and can fuse or divide. Actin waves separate two phases of the plasma membrane that are distinguished by their lipid composition. The area circumscribed by a wave resembles in its phosphoinositide content the interior of a phagocytic cup, leading us to explore the possibility that actin waves are in-plane phagocytic structures generated without the localized stimulus of an attached particle. Consistent with this view, wave-forming cells were found to exhibit a high propensity for taking up particles. Cells fed rod-shaped particles produced elongated phagocytic cups that displayed a zonal pattern that reflected in detail the actin and lipid pattern of free-running actin waves. Neutrophils and macrophages are known to spread on surfaces decorated with immune complexes, a process that has been interpreted as “frustrated” phagocytosis. We suggest that actin waves enable a phagocyte to scan a surface for particles that might be engulfed. Submitted by Günther Gerisch [gerisch@biochem.mpg.de] -------------------------------------------------------------------------------- Food searching strategy of amoeboid cells by starvation induced run length extension. Peter J.M. Van Haastert and Leonard Bosgraaf Department of Cell Biochemistry, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands PLoS ONE, in press Food searching strategies of animals are key to their success in heterogeneous environments. The optimal search strategy may include specialized random walks such as Levy walks with heavy power-law tail distributions, or persistent walks with preferred movement in a similar direction. We have investigated the movement of the soil amoebae Dictyostelium searching for food. Dictyostelium cells move by extending pseudopodia, either in the direction of the previous pseudopod (persistent step) or in a different direction (turn). The analysis of ~4000 pseudopodia reveals that step and turn pseudopodia are drawn from a probability distribution that is determined by cGMP/PLA2 signaling pathways. Starvation activates these pathways thereby suppressing turns and inducing steps. As a consequence, starved cells make very long nearly straight runs and disperse over ~30-fold larger areas, without extending more or larger pseudopodia than vegetative cells. This ‘win-stay/lose-shift’ strategy for food searching is called Starvation Induced Run-length Extension. The SIRE walk explains very well the observed differences in search behavior between fed and starving organisms such as bumble-bees, flower bug, hoverfly and zooplankton. Submitted by Peter Van Haastert [p.j.m.van.haastert@rug.nl] -------------------------------------------------------------------------------- Navigation of chemotactic cells by parallel signaling to pseudopod persistence and orientation. Leonard Bosgraaf and Peter J.M. Van Haastert Department of Cell Biochemistry, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands PLoS ONE, in press The mechanism of chemotaxis is one of the most interesting issues in modern cell biology. Recent work shows that shallow chemoattractant gradients do not induce the generation of pseudopods, as has been predicted in many models. This poses the question of how else cells can steer towards chemoattractants. Here we use a new computational algorithm to analyze the extension of pseudopods by Dictyostelium cells. We show that a shallow gradient of cAMP induces a small bias in the direction of pseudopod extension, without significantly affecting parameters such as pseudopod frequency or size. Persistent movement, caused by alternating left/right splitting of existing pseudopodia, amplifies the effects of this bias by up to 5-fold. Known players in chemotactic pathways play contrasting parts in this mechanism; PLA2 and cGMP signal to the cytoskeleton to regulate the splitting process, while PI 3-kinase and soluble guanylyl cyclase mediate the directional bias. The coordinated regulation of pseudopod generation, orientation and persistence by multiple signaling pathways allows eukaryotic cells to detect extremely shallow gradients. Submitted by Peter Van Haastert [p.j.m.van.haastert@rug.nl] -------------------------------------------------------------------------------- The local cell curvature guides pseudopodia towards chemoattractants. Peter J.M. Van Haastert and Leonard Bosgraaf Department of Cell Biochemistry, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands HFSP Journal, in press Many eukaryotic cells use pseudopodia for movement towards chemoattractants. We developed a computer algorithm to identify pseudopodia, and analyzed how pseudopodia of Dictyostelium cells are guided towards cAMP. Surprisingly, the direction of a pseudopod is not actively oriented towards the gradient, but is always perpendicular to the local cell curvature. The gradient induces a bias in the position where the pseudopod emerges: pseudopodia more likely emerge at the side of the cell closer to the gradient where perpendicular pseudopodia are pointed automatically towards the chemoattractant. A mutant lacking the formin dDia2 is not spherical but has many invaginations. Although pseudopodia still emerge at the side closer to the gradient, the surface curvature is so irregular that many pseudopodia are not extended towards cAMP. The results imply that the direction of pseudopod extension, and therefore also the direction of cell movement, is dominated by two aspects: the position at the cell surface where a pseudopod emerges, and the local curvature of the membrane at that position. Submitted by Peter Van Haastert [p.j.m.van.haastert@rug.nl] ============================================================== [End dictyNews, volume 33, number 5]