Dicty News Electronic Edition Volume 10, number 13 May 9, 1998 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to dicty@nwu.edu. Back issues of Dicty-News, the Dicty Reference database and other useful information is available at the Dictyostelium Web Page "http://dicty.cmb.nwu.edu/dicty/dicty.html" =========== Abstracts =========== Identification and Analysis of a Gene that is Essential for Morphogenesis and Prespore Cell Differentiation in Dictyostelium Hiroo Yasukawa, Sudhasri Mohanty, and Richard A. Firtel Dept. of Biology, Center for Molecular Genetics, UCSD, La Jolla DEVELOPMENT, in press SUMMARY We have identified a gene (PslA) that is expressed throughout Dictyostelium development and encodes a novel protein that is required for proper aggregation and subsequent cell-type differentiation and morphogenesis. pslA null (pslA-) cells produce large aggregation streams under conditions in which wild-type cells form discrete aggregates. Tips form along the stream, elongate to produce a finger, and eventually form a terminal structure that lacks a true sorus (spore head). More than half of the cells remain as a mass at the base of the developing fingers. The primary defect in the pslA- strain is the inability to induce prespore cell differentiation. Analyses of gene expression show a complete lack of prespore-specific gene expression and no mature spores are produced. In chimeras with wild-type cells, pslA- cells form the prestalk domain and normal, properly proportioned fruiting bodies can be produced. This indicates that pslA- cells are able to interact with wild-type cells and regulate patterning, even though pslA- cells are unable to express prespore cell-type-specific genes, do not participate in prespore cell differentiation, and do not produce pslA- spores in the chimeras. While pslA- cells produce mature, vacuolated stalk cells during multicellular development, pslA- cells are unable to do so in vitro in response to exogenous DIF (a morphogen required for prestalk and stalk cell differentiation). These results indicate that pslA- cells exhibit a defect in the prestalk/stalk cell pathways under these experimental conditions. Our results suggest that PslA’s primary function is to regulate prespore-cell determination very early in the prespore pathway via a cell-autonomous mechanism, possibly at the time of the initial prestalk/prespore cell-fate decision. Indirect immunofluorescence of myc-tagged PslA localizes the protein to the nucleus, suggesting that PslA may function to control the prespore pathway at the level of transcription. ------------------------------------------------------------------------- The helC gene encodes a putative DEAD-box RNA helicase required for development in Dictyostelium discoideum Laura M. Machesky, Robert H. Insall and Robert R. Kay Current Biology, vol. 8 iss. 10, in press Abstract DEAD-box RNA helicases, defined by the sequence Asp-Glu-Ala-Asp (DEAD, in single-letter amino-acid code), regulate RNA unwinding and secondary structure in an ATP-dependent manner in vitro and control mRNA stability and protein translation. Both yeast and mammals have large families of DEAD-box proteins, many of unknown function. We have disrupted a Dictyostelium discoideum gene, helC, which encodes helicase C, a member of the DEAD-box family of RNA helicases that shows strong homology to the product of the essential Saccharomyces cerevisiae gene dbp5 and to related helicases in mouse and Schizosaccharomyces pombe. The HelC protein also shows weaker homology to the translation initiation factor eIF-4a. Other DEAD-box-containing proteins, which are less closely related to HelC, have been implicated in developmental roles in Drosophila and Xenopus laevis; one example is the Xenopus Vasa-like protein (XVLP). In Drosophila and Xenopus, Vasa and XVLP, respectively, are specifically expressed in germ cells and are required for the establishment of tissue polarity during development. In yeast, DEAD-box helicases such as Prp8 are components of the spliceosome and connect pre-mRNA splicing with the cell cycle. Disruption of the helC gene in D. discoideum led to developmental asynchrony, failure to differentiate and aberrant morphogenesis. We postulate that one reason for the existence of large families of homologous DEAD-box proteins in yeast, mammals and Dictyostelium could be that some DEAD-box proteins have develop- mentally specific roles regulating protein translation or mRNA stability. ------------------------------------------------------------------------- LagC-Null and GBF-Null Define Key Steps in the Morphogenesis of Dictyostelium Mounds. Sujatha Sukumaran, Jason M. Brown, Richard A. Firtel and James G. McNally Devel. Biol., in press. ABSTRACT The transition to multicellularity is a key feature of the Dictyostelium life- cycle, and two genes, gbf and lagC, are known to play pivotal roles in regulating this developmental switch. lagC-null and gbf-null cells fail to induce cell-type-specific genes ordinarily expressed during multicellular development. The null mutants also share a similar morphological phenotype: mutant cells repeatedly aggregate to form a loose mound, disperse, and reform a mound, rather than proceeding to form a tip. To characterize defects in morphogenesis in these mutants, we examined cell motion in the mutant mounds. In analogy with the failed transition in gene expression, we found that lagC-null and gbf-null mounds failed to make a morphogenetic transition from random to rotational motion normally observed in the parent strain. One reason for this was the inability of the mutant mounds to establish a single, dominant signaling- wave center. This defect of lagC-null or gbf-null cells could be overcome by the addition of adenosine, which alters cAMP signaling, but then even in the presence of apparently normal signaling waves, cell motility was still aberrant. This motility defect, as well as the signaling-wave defect, could be overcome in lagC-null cells by overexpression of GBF, suggesting that lagC is dispensable if GBF protein levels are high enough. This set of morphogenetic defects that we have observed help define key steps in mound morphogenesis. These include the establishment of a dominant signaling-wave center and the capacity of cells to move directionally within the cell mass in response to guidance cues. ------------------------------------------------------------------------- [End Dicty News, volume 10, number 13]