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1.  Overexpression of an activated rasG gene during growth blocks the initiation of Dictyostelium development. 
Molecular and Cellular Biology  1996;16(8):4156-4162.
Transformants that expressed either the wild-type rasG gene, an activated rasG-G12T gene, or a dominant negative rasG-S17N gene, all under the control of the folate-repressible discoidin (dis1gamma) promoter, were isolated. All three transformants expressed high levels of Ras protein which were reduced by growth in the presence of folate. All three transformants grew slowly, and the reduction in growth rate correlated with the amount of RasG protein produced, suggesting that RasG is important in regulating cell growth. The pVEII-rasG transformant containing the wild-type rasG gene developed normally despite the presence of high levels of RasG throughout development. This result indicates that the down regulation of rasG that normally occurs during aggregation of wild-type strains is not essential for the differentiation process. Dictyostelium transformants expressing the dominant negative rasG-S17N gene also differentiated normally. Dictyostelium transformants that overexpressed the activated rasG-G12T gene did not aggregate. The defect occurred very early in development, since the expression of car1 and pde, genes that are normally induced soon after the initiation of development, was repressed. However, when the transformant cells were pulsed with cyclic AMP, expression of both genes returned to wild-type levels. The transformants exhibited chemotaxis to cyclic AMP, and development was synergized by mixing with wild-type cells. Furthermore, cells that were pulsed with cyclic AMP for 4 h before being induced to differentiate by plating on filters produced small, but otherwise normal, fruiting bodies. These results suggest that the rasG-G12T transformants are defective in cyclic AMP production and that RasG - GTP blocks development by interfering with the initial generation of cyclic AMP pulses.
PMCID: PMC231412  PMID: 8754814
2.  Regulation of DdrasG gene expression during Dictyostelium development. 
Molecular and Cellular Biology  1990;10(3):918-922.
DdrasG gene expression during the early development of Dictyostelium discoideum has been examined in detail. The amount of DdrasG-specific mRNA increased approximately twofold during the first 2 to 3 h of development and then declined rapidly, reaching negligible levels by the aggregation stage. The increase in mRNA levels that occurred during the first 2 to 3 h of development also occurred during differentiation in cell suspensions and was enhanced when cells were shaken rapidly. This initial increase was unaffected by cell density. When cells were set up to differentiate on filters, the addition of a glucose-amino acid mixture slightly delayed differentiation and had a similar effect on the expression of the gene. The decline in DdrasG expression during development did not occur when cells were treated with cycloheximide, suggesting that the expression of a developmentally regulated gene product is essential for the reduction of DdrasG gene mRNA. There was no decrease in DdrasG mRNA level during differentiation in shake suspension, but the decrease did occur upon application of pulses of cyclic AMP to shaking cultures. The application of a continuously high level of cyclic AMP delayed the increase in expression of the gene and did not result in the subsequent decline. These results suggest that the induction of a functional cyclic AMP relay system is important in reducing DdrasG gene mRNA levels.
PMCID: PMC360932  PMID: 2154684
3.  Regulation of a ras-related protein during development of Dictyostelium discoideum. 
Recent work has shown that DNA sequences related to the mammalian ras proto-oncogenes are highly conserved in eucaryotic evolution. A monoclonal antibody (Y13-259) to mammalian p21ras specifically precipitated a 23,000-molecular-weight protein (p23) from lysates of Dictyostelium discoideum amoebae. Tryptic peptide analysis indicated that D. discoideum p23 was closely related in its primary structure to mammalian p21ras. p23 was apparently derived by post-translational modification of a 24,000-molecular-weight primary gene product. The amount of p23 was highest in growing amoebae, but declined markedly with the onset of differentiation such that by fruiting body formation there was less than 10% of the amoeboid level. The rate of p23 synthesis dropped rapidly during aggregation, rose transiently during pseudoplasmodial formation, and then declined during the terminal stages of differentiation. There was, therefore, a strong correlation between the expression of the ras-related protein p23 and cell proliferation of D. discoideum.
PMCID: PMC366674  PMID: 3920511
4.  Allergic reaction to chlormethiazole. 
British Medical Journal  1977;1(6056):290.
PMCID: PMC1604157  PMID: 837079
5.  Effect of cerulenin on the growth and differentiation of Dictyostelium discoideum. 
Journal of Bacteriology  1976;128(1):21-27.
The growth of Dictyostelium discoideum Ax-2 was inhibited completely by cerulenin at a concentration of 5 mug/ml. This inhibition of growth was found to be due to the inhibition of fatty acid synthesis. Acetate incorporation into a long-chain fatty acid was inhibited completely by cerulenin, and the growth inhibition could be reversed by inclusion of certain saturated fatty acids in the medium. Unsaturated fatty acids and sterols failed to reverse the inhibitory effect. The fatty acid and sterol compositions of cerulenin-treated cells were determined to establish whether the drug could be used to manipulate the organism's lipid composition. Only relatively small manipulations were obtained under the conditions employed in this study. Cerulenin inhibited differentiation but only at high concentrations (150 mug/ml). This inhibition could be reversed by palmitic acid, suggesting that the prime cause of the inhibition was an inhibition of fatty acid synthesis. Thus, it appears that continued fatty acid synthesis is required for the cellular process of differentiation in D. discoideum.
PMCID: PMC232821  PMID: 988014
6.  A ras-related gene from the lower eukaryote Dictyostelium that is highly conserved relative to the human rap genes. 
Nucleic Acids Research  1990;18(17):5265-5269.
The cellular slime mold Dictyostelium discoideum contains two ras genes, DdrasG and Ddras that are differentially expressed during development. We have characterized a gene that hybridized to both Ddras and DdrasG under low, but not under high stringency conditions. The deduced amino acid sequence is highly conserved with respect to the human rap (Krev-1, smg21) proteins and the corresponding gene has been designated Ddrap1. The Ddrap1 gene is expressed at all stages during development but is expressed maximally during the aggregation and culmination periods when the expression of Ddras and DdrasG is declining. During vegetative growth and early development Ddrap1 cDNA hybridizes to a single mRNA of 1.1 kb. As development progresses the level of this mRNA declines and messages of 1.0 and 1.3 kb appear.
PMCID: PMC332150  PMID: 2205839
7.  Expression of an activated rasD gene changes cell fate decisions during Dictyostelium development. 
Molecular Biology of the Cell  1997;8(2):303-312.
It has been previously demonstrated that the expression of an activated rasD gene in wild-type Dictyostelium cells results in formation of aggregates with multitips, instead of the normal single tips, and a block in further development. In an attempt to better understand the role of activated RasD development, we examined cell-type-specific gene expression in a strain stably expressing high levels of RasD[G12T]. We found that the expression of prestalk cell-specific genes ecmA and tagB was markedly enhanced, whereas the expression of the prespore cell-specific gene cotC was reduced to very low levels. When the fate of cells in the multitipped aggregate was monitored with an ecmA/lacZ fusion, it appeared that most of the cells eventually adopted prestalk gene expression characteristics. When mixtures of the [G12T]rasD cells and Ax3 cells were induced to differentiate, chimeric pseudoplasmodia were not formed. Thus, although the [G12T]rasD transformant had a marked propensity to form prestalk cells, it could not supply the prestalk cell population when mixed with wild-type cells. Both stalk and spore cell formation occurred in low cell density monolayers of the [G12T]rasD strain, suggesting that at least part of the inhibition of stalk and spore formation during multicellular development involved inhibitory cell interactions within the cell mass. Models for the possible role of rasD in development are discussed.
PMCID: PMC276081  PMID: 9190209

Results 1-7 (7)