Arachidonic acid is a chemoattractant for Dictyostelium discoideum cells.

Cyclic AMP (cAMP)is a natural chemoattractant of the social amoeba Dictyostelium discoideum. It is detected by cell surface cAMP receptors. Besides a signalling cascade involving phosphatidylinositol 3,4,5-trisphosphate (PIP3), Ca2+ signalling has been shown to have a major role in chemotaxis. Previously, we have shown that arachidonic acid (AA) induces an increase in the cytosolic Ca2+ concentration by causing the release of Ca2+ from intracellular stores and activating influx of extracellular Ca2+. Here we report that AA is a chemoattractant for D. discoideum cells differentiated for 8-9 h. Motility towards a glass capillary filled with an AA solution was dose-dependent and qualitatively comparable to cAMP-induced chemotaxis. Ca2+ played an important role in AA chemotaxis of wild-type Ax2 as ethyleneglycol-bis(b-aminoethyl)-N,N,N',N'-tetraacetic acid (EGTA) added to the extracellular buffer strongly inhibited motility. In the HM1049 mutant whose iplA gene encoding a putative Ins(1,4,5)P3 -receptor had been knocked out, chemotaxis was only slightly affected by EGTA. Chemotaxis in the presence of extracellular Ca2+ was similar in both strains. Unlike cAMP, addition of AA to a cell suspension did not change cAMP or cGMP levels. A model for AA chemotaxis based on the findings in this and previous work is presented.

A model for cell type localization in the migrating slug of Dictyostelium discoideum based on differential chemotactic sensitivity to cAMP and differential sensitivity to suppression of chemotaxis by ammonia.

The three basic cell types in the migrating slug of Dictyostelium discoideum show differential chemotactic response to cyclic AMP (cAMP) and differential sensitivity to suppression of the chemotaxis by ammonia.The values of these parameters indicate a progressive maturation of chemotactic properties during the transdifferentiation of slug cell types.We present a model that explains the localization of the three cell types within the slug based on these chemotactic differences and on the maturation of their chemotactic properties.

Signaling molecules involved in the transition of growth to development of Dictyostelium discoideum.

The social amoeba Dictyostelium discoideum, a powerful paradigm provides clear insights into the regulation of growth and development. In addition to possessing complex individual cellular functions like a unicellular eukaryote, D. discoideum cells face the challenge of multicellular development. D. discoideum undergoes a relatively simple differentiation process mainly by cAMP mediated pathway. Despite this relative simplicity, the regulatory signaling pathways are as complex as those seen in metazoan development. However, the introduction of restriction-enzyme-mediated integration (REMI) technique to produce developmental gene knockouts has provided novel insights into the discovery of signaling molecules and their role in D. discoideum development. Cell cycle phase is an important aspect for differentiation of D. discoideum, as cells must reach a specific stage to enter into developmental phase and specific cell cycle regulators are involved in arresting growth phase genes and inducing the developmental genes. In this review, we present an overview of the signaling molecules involved in the regulation of growth to differentiation transition (GDT), molecular mechanism of early developmental events leading to generation of cAMP signal and components of cAMP relay system that operate in this paradigm.

Effect of arsenic on cell growth of the cellular slime mould, Dictyostelium discoideum.

The growing D. discoideum cells were killed in a dose-dependent manner when exposed to 100 and 140 ppm of arsenic (As2O3) at mid-log phase for 20 min. Reduced plaque sizes and changed cell and colony morphologies were observed in the treated cells. Endocytotic functions (both phagocytosis and pinocytosis) were also inhibited in the treated cells. Arsenic treated cell showed a lower DNA and protein synthetic activities. These findings are discussed in relation to known mechanism of action of the heavy metal on growth-related cellular functions.

Ammonia differentially suppresses the cAMP chemotaxis of anterior-like cells and prestalk cells in Dictyostelium discoideum.

A drop assay for chemotaxis to cAMP confirms that both anterior-like cells (ALC) and prestalk cells (pst cells) respond to cAMP gradients. We present evidence that the chemotactic response of both ALC and pst cells is suppressed by ammonia, but a higher concentration of ammonia is required to suppress the response in pst cells. ALC show a chemotactic response to cAMP when moving on a substratum of prespore cells in isolated slug posteriors incubated under oxygen. ALC chemotaxis on a prespore cell substratum is suppressed by the same concentration of ammonia that suppresses ALC chemotaxis on the agar substratum in drop assays. Chemotaxis suppression is mediated by the unprotonated (NH3) species of ammonia. The observed suppression, by ammonia, of ALC chemotaxis to cAMP supports our earlier hypothesis that ammonia is the tip-produced suppressor of such chemotaxis. We discuss implications of ammonia sensitivity of pst cells and ALC with regard to the movement and localization of ALC and pst cells in the slug and to the roles played by ALC in fruiting body formation. In addition, we suggest that a progressive decrease in sensitivity to ammonia is an important part of the maturation of ALC into pst cells.

Mechanism of cAMP-induced H(+)-efflux of Dictyostelium cells: a role for fatty acids.

Aggregating Dictyostelium cells release protons when stimulated with cAMP. To find out whether the protons are generated by acidic vesicles or in the cytosol, we permeabilized the cells and found that this did not alter the cAMP-response. Proton efflux in intact cells was inhibited by preincubation with the V-type H(+) ATPase inhibitor concanamycin A and with the plasma membrane H(+) ATPase blocker miconazole. Surprisingly, miconazole also inhibited efflux in permeabilized cells, indicating that this type of H(+) ATPase is present on intracellular vesicles as well. Vesicular acidification was inhibited by miconazole and by concanamycin A, suggesting that the acidic vesicles contain both V-type and P-type H(+) ATPases. Moreover, concanamycin A and miconazole acted in concert, both in intact cells and in vesicles. The mechanism of cAMP-induced Ca2(+)-fluxes involves phospholipase A2 activity. Fatty acids circumvent the plasma membrane and stimulate vesicular Ca2(+)-efflux. Here we show that arachidonic acid elicited H(+)-efflux not only from intact cells but also from acidic vesicles. The target of regulation by arachidonic acid seemed to be the vesicular Ca2(+)-release channel.

The precision of regulation in Dictyostelium discoideum: implications for cell-type proportioning in the absence of spatial pattern.

We have made careful counts of the exact number of spore, stalk and basal disc cells in small fruiting bodies of Dictyostelium discoideum (undifferentiated amoebae are found only rarely and on average their fraction is 4.96 x 10(-4)). (i) Within aggregates of a given size, the relative apportioning of amoebae to the main cell types occurs with a remarkable degree of precision. In most cases the coefficient of variation (c.v.) in the mean fraction of cells that form spores is within 4.86%. The contribution of stalk and basal disc cells is highly variable when considered separately (c.v.'s upto 25% and 100%, respectively), but markedly less so when considered together. Calculations based on theoretical models indicate that purely cell-autonomous specification of cell fate cannot account for the observed accuracy of proportioning. Cell-autonomous determination to a prestalk or prespore condition followed by cell type interconversion, and stabilised by feedbacks, suffices to explain the measured accuracy. (ii) The fraction of amoebae that differentiates into spores increases monotonically with the total number of cells. This fraction rises from an average of 73.6% for total cell numbers below 30 and reaches 86.0% for cell numbers between 170 and 200 (it remains steady thereafter at around 86%). Correspondingly, the fraction of amoebae differentiating into stalk or basal disc decreases with total size. These trends are in accordance with evolutionary expectations and imply that a mechanism for sensing the overall size of the aggregate also plays an essential role in the determination of cell-type proportions.