Co-occurrence in nature of different clones of the social amoeba,Dictyostelium discoideum

The social amoeba, Dictyostelium discoideum, produces a multicellular fruiting body and has become a model system for cell-cell interactions such as signalling, adhesion and development. However, unlike most multicellular organisms, it forms by aggregation of cells and, in the laboratory, forms genetic chimeras where there may be competition among clones. Here we show that chimera formation is also likely in nature, because different clones commonly co-occur on a very small scale. This suggests that D. discoideum will likely have evolved strategies for competing in chimeras, and that the function of some developmental genes will be competitive. Natural chimerism also makes D. discoideum a good model organism for the investigation of issues relating to coexistence and conflict between cells.     

Stalk size and altruism investment within and among populations of the social amoeba

Reproductive division of labour is common in many societies, including those of eusocial insects, cooperatively breeding vertebrates, and most forms of multicellularity. However, conflict over what is best for the individual vs. the group can prevent an optimal division of labour from being achieved. In the social amoeba Dictyostelium discoideum, cells aggregate to become multicellular and a fraction behaves altruistically, forming a dead stalk that supports the rest. Theory suggests that intra‐organismal conflict over spore–stalk cell fate can drive rapid evolutionary change in allocation traits, leading to polymorphisms within populations or rapid divergence between them. Here, we assess several proxies for stalk size and spore–stalk allocation as metrics of altruism investment among strains and across geographic regions. We observe geographic divergence in stalk height that can be partly explained by differences in multicellular size, as well as variation among strains in clonal spore–stalk allocation, suggesting within‐population variation in altruism investment. Analyses of chimeras comprised of strains from the same vs. different populations indicated genotype‐by‐genotype epistasis, where the morphology of the chimeras deviated significantly from the average morphology of the strains developed clonally. The significantly negative epistasis observed for allopatric pairings suggests that populations are diverging in their spore–stalk allocation behaviours, generating incompatibilities when they encounter one another. Our results demonstrate divergence in microbial social traits across geographically separated populations and demonstrate how quantification of genotype‐by‐genotype interactions can elucidate the trajectory of social trait evolution in nature.     

Cheating does not explain selective differences at high and low relatedness in a social amoeba

Background  

Altruism can be favored by high relatedness among interactants. We tested the effect of relatedness in experimental populations of the social amoeba Dictyostelium discoideum, where altruism occurs in a starvation-induced social stage when some amoebae die to form a stalk that lifts the fertile spores above the soil facilitating dispersal. The single cells that aggregate during the social stage can be genetically diverse, which can lead to conflict over spore and stalk allocation. We mixed eight genetically distinct wild isolates and maintained twelve replicated populations at a high and a low relatedness treatment. After one and ten social generations we assessed the strain composition of the populations. We expected that some strains would be out-competed in both treatments. In addition, we expected that low relatedness might allow the persistence of social cheaters as it provides opportunity to exploit other strains.     

A retinoblastoma ortholog controls stalk/spore preference in Dictyostelium

We describe rblA, the Dictyostelium ortholog of the retinoblastoma susceptibility gene Rb. In the growth phase, rblA expression is correlated with several factors that lead to 'preference' for the spore pathway. During multicellular development, expression increases 200-fold in differentiating spores. rblA-null strains differentiate stalk cells and spores normally, but in chimeras with wild type, the mutant shows a strong preference for the stalk pathway. rblA-null cells are hypersensitive to the stalk morphogen DIF, suggesting that rblA normally suppresses the DIF response in cells destined for the spore pathway. rblA overexpression during growth leads to G1 arrest, but as growing Dictyostelium are overwhelmingly in G2 phase, rblA does not seem to be important in the normal cell cycle. rblA-null cells show reduced cell size and a premature growth-development transition; the latter appears anomalous but may reflect selection pressures acting on social ameba.     

The costs and benefits of being a chimera

Most multicellular organisms are uniclonal. This is hypothesized to be because uniclonal organisms function better than chimeras (non-clonal organisms), owing to reduced levels of internal genetic conflict. We tested this idea using the social amoeba or slime mold Dictyostelium discoideum. When starving, the normally solitary amoebae aggregate to form a differentiated multicellular slug that migrates towards light and forms a fruiting body, facilitating the dispersal of spores. We added 10(7) amoebae to Petri plates containing 1, 2, 5 or 10 clones mixed together. We found an intrinsic cost to chimerism: chimeric slugs moved significantly less far than uniclonal slugs of the same size. However, in nature, joining with other clones to form a chimera should increase slug size, and larger slugs travel further. We incorporated this size effect into a second experiment by giving chimeras more cells than single clones (single clones had 10(6) cells, two-clone chimeras had 2 x 10(6) cells and so on). The uniclonal treatments then simulated a clone in a mixture that refuses to form chimeras. In this experiment, chimeras moved significantly further than the uniclonal slugs, in spite of the intrinsic cost. Thus, chimerism is costly, which may be why it evolves so seldom, but in D. discoideum the benefits of large size appear to compensate.     

Evolutionary dynamics of altruism and cheating among social amoebas

Dictyostelium discoideum is a eukaryotic amoeba, which, when starvation is imminent, aggregates to form fruiting bodies consisting of a stalk of reproductively dead cells that supports spores. Because different clones may be involved in such aggregations, cheater strategies may emerge that allocate a smaller fraction of cells to stalk formation, thus gaining a reproductive advantage. In this paper, we model the evolutionary dynamics of allocation strategies in Dictyostelium under the realistic assumption that the number of clones involved in aggregations follows a random distribution. By determining the full course of evolutionary dynamics, we show that evolutionary branching in allocation strategies may occur, resulting in dimorphic populations that produce stalkless and stalked fruiting bodies. We also demonstrate that such dimorphisms are more likely to emerge when the variation in the number of clones involved in aggregations is large.     

The cold war of the social amoebae

When confronted with starvation, the amoebae of Dictyostelium discoideum initiate a developmental process that begins with cell aggregation and ends with a ball of spores supported on a stalk. Spores live and stalk cells die. Because the multicellular organism is produced by cell aggregation and not by growth and division of a single cell, genetically diverse amoebae may enter an aggregate and, if one lineage has a capacity to avoid the stalk cell fate, it may have a selective advantage. Such cheater mutants have been found among wild isolates and created in laboratory strains. The mutants raise a number of questions--how did such a cooperative system evolve in the face of cheating? What is the basis of self recognition? What genes are involved? How is cheating constrained? This review summarizes the results of studies on the social behavior of Dictyostelium and its relatives, including the familiar asexual developmental cycle and the lesser known, but puzzling, sexual cycle.     

A linear dominance hierarchy among clones in chimeras of the social amoeba Dictyostelium discoideum

Amoebae from different clones of Dictyostelium discoideum aggregate into a common slug, which migrates towards light for dispersal, then forms a fruiting body consisting of a somatic, dead stalk, holding up a head of living spores. Contributions of two clones in a chimera to spore and stalk are often unequal, with one clone taking advantage of the other's stalk contribution. To determine whether there was a hierarchy of exploitation among clones, we competed all possible pairs among seven clones and measured their relative representation in the prespore and prestalk stages and in the final spore stage. We found a clear linear hierarchy at the final spore stage, but not at earlier stages. These results suggest that there is either a single principal mechanism or additive effects for differential contribution to the spore, and that it involves more than spore/stalk competition.     

An invitation to die: initiators of sociality in a social amoeba become selfish spores

Greater size and strength are common attributes of contest winners. Even in social insects with high cooperation, the right to reproduce falls to the well-fed queens rather than to poorly fed workers. In Dictyostelium discoideum, formerly solitary amoebae aggregate when faced with starvation, and some cells die to form a stalk which others ride up to reach a better location to sporulate. The first cells to starve have lower energy reserves than those that starve later, and previous studies have shown that the better-fed cells in a mix tend to form disproportionately more reproductive spores. Therefore, one might expect that the first cells to starve and initiate the social stage should act altruistically and form disproportionately more of the sterile stalk, thereby enticing other better-fed cells into joining the aggregate. This would resemble caste determination in social insects, where altruistic workers are typically fed less than reproductive queens. However, we show that the opposite result holds: the first cells to starve become reproductive spores, presumably by gearing up for competition and outcompeting late starvers to become prespore first. These findings pose the interesting question of why others would join selfish organizers.     

Rapid antagonistic coevolution between strains of the social amoeba Dictyostelium discoideum

Social groups face a fundamental problem of overcoming selfish individuals capable of destroying cooperation. In the social amoeba Dictyostelium discoideum, there is evidence that some clones (‘cheaters’) contribute disproportionately to the viable spores in a fruiting body while avoiding the dead stalk cell fate. It remains unclear, however, whether this cheating is actually the product of selection. Here, I report the results of an experimental evolution study designed to test whether clones of D. discoideum will evolve resistance to cheating in the laboratory with genetic variation created only through spontaneous mutation. Two strains, one green fluorescent protein (GFP)-labelled and one wild-type, were allowed to grow and develop together before the wild-type strain was removed and replaced with a naïve strain evolving in parallel. Over the course of 10 social generations, the GFP-labelled strain reliably increased its representation in the spores relative to control populations that had never experienced the competitor. This competitive advantage extended to the non-social, vegetative growth portion of the life cycle, but not to pairwise competition with two other strains. These results indicate strong antagonism between strains, mediated by ample mutational variation for cheating and also suggest that arms races between strains in the wild may be common.     

Exploiting new terrain: an advantage to sociality in the slime mold Dictyostelium discoideum

Understanding the ecological benefits of social actions is centralto explaining the evolution of social behavior. The social amoebaDictyostelium discoideum has been well studied and is a modelfor social evolution and development, but surprisingly littleis known about its ecology. When starving, thousands of thenormally solitary amoebae aggregate to form a differentiatedmulticellular organism known as a slug. The slug migrates towardthe soil surface where it metamorphoses into a fruiting bodyof hardy spores held up by a dead stalk comprising about one-fifthof the cells. Multicellularity in D. discoideum is thought tohave evolved to lift the spores above the hazards of the soilwhere spores can be picked up for long-distance dispersal. Here,we show that multicellularity has another advantage: local dispersalto new food sources. We find that cells shed by D. discoideumslugs during migration consume and remove bacteria in the pathof the slug, although slugs themselves do not breakup. We alsoshow that slugs are adept at local dispersal by comparing migrationof slugs with migration of individual cells of the mutant, CAP2,which cannot aggregate and so rely only on cellular movement.In particular, the solitary cells of the aggregation mutantare unable to cross a soil barrier, easily crossed by slugs.We propose that the exploitation of local food patches is animportant selective benefit favoring multicellular cooperationin D. discoideum.     

High relatedness in a social amoeba: the role of kin-discriminatory segregation

A major challenge for social theory is to explain the importance of kin discrimination for the evolution of altruism. One way to assess the importance of kin discrimination is to test its effects on increasing relatedness within groups. The social amoeba Dictyostelium discoideum aggregates to form a fruiting body composed of dead stalk and live spores. Previous studies of a natural population showed that where D. discoideum occurs in the soil, multiple clones are often found in the same small soil samples. However, actual fruiting bodies usually contain only one clone. We here performed experiments to gauge the effect of kin-discriminatory segregation on increasing relatedness. We mixed co-occurring clones from this population using a relatedness level found in small soil samples. We found a lower proportion of uniclonal fruiting bodies and a lower level of relatedness compared with natural fruiting bodies. We found that the amount of relatedness increase attributable to kin-discriminatory segregation was small. These findings suggest a relatively minor influence of kin-discriminatory segregation on relatedness in D. discoideum. We discuss our results comparing with the results of previous studies, including those of wild clones and laboratory mutants. We ask why wild clones of D. discoideum exhibit a low degree of kin-discriminatory segregation, and what alternative factors might account for high relatedness in D. discoideum.     

Species recognition in social amoebae

Aggregative multicellularity requires the ability of cells to recognise conspecifics. Social amoebae are among the best studied of such organisms, but the mechanism and evolutionary background of species recognition remained to be investigated. Here we show that heterologous expression of a single Dictyostelium purpureum gene is sufficient for D. discoideum cells to efficiently make chimaeric fruiting bodies with D. purpureum cells. This gene forms a bidirectional pair with another gene on the D. purpureum genome, and they are both highly polymorphic among independent wild isolates of the same mating group that do not form chimaeric fruiting bodies with each other. These paired genes are both structurally similar to D. discoideum tgrB1/C1 pair, which is responsible for clonal discrimination within that species, suggesting that these tgr genes constitute the species recognition system that has attained a level of precision capable of discriminating between clones within a species. Analysis of the available genome sequences of social amoebae revealed that such gene pairs exist only within the clade composed of species that produce precursors of sterile stalk cells (prestalk cells), suggesting concurrent evolution of a precise allorecognition system and a new ‘worker’ cell-type dedicated to transporting and supporting the reproductive cells.     

The trishanku gene and terminal morphogenesis in Dictyostelium discoideum

SUMMARY Multicellular development in the social amoeba Dictyostelium discoideum is triggered by starvation. It involves a series of morphogenetic movements, among them being the rising of the spore mass to the tip of the stalk. The process requires precise coordination between two distinct cell types—presumptive (pre-) spore cells and presumptive (pre-) stalk cells. Trishanku ( triA ) is a gene expressed in prespore cells that is required for normal morphogenesis. The triA⁻ mutant shows pleiotropic effects that include an inability of the spore mass to go all the way to the top. We have examined the cellular behavior required for the normal ascent of the spore mass. Grafting and mixing experiments carried out with tissue fragments and cells show that the upper cup, a tissue that derives from prestalk cells and anterior-like cells (ALCs), does not develop properly in a triA⁻ background. A mutant upper cup is unable to lift the spore mass to the top of the fruiting body, likely due to defective intercellular adhesion. If wild-type upper cup function is provided by prestalk and ALCs, trishanku spores ascend all the way. Conversely, Ax2 spores fail to do so in chimeras in which the upper cup is largely made up of mutant cells. Besides proving that under these conditions the wild-type phenotype of the upper cup is necessary and sufficient for terminal morphogenesis in D. discoideum , this study provides novel insights into developmental and evolutionary aspects of morphogenesis in general. Genes that are active exclusively in one cell type can elicit behavior in a second cell type that enhances the reproductive fitness of the first cell type, thereby showing that morphogenesis is a cooperative process.     

Bestatin inhibits cell growth, cell division, and spore cell differentiation in Dictyostelium discoideum

Bestatin methyl ester (BME) is an inhibitor of Zn(2+)-binding aminopeptidases that inhibits cell proliferation and induces apoptosis in normal and cancer cells. We have used Dictyostelium as a model organism to study the effects of BME. Only two Zn(2+)-binding aminopeptidases have been identified in Dictyostelium to date, puromycin-sensitive aminopeptidase A and B (PsaA and PsaB). PSA from other organisms is known to regulate cell division and differentiation. Here we show that PsaA is differentially expressed throughout growth and development of Dictyostelium, and its expression is regulated by developmental morphogens. We present evidence that BME specifically interacts with PsaA and inhibits its aminopeptidase activity. Treatment of cells with BME inhibited the rate of cell growth and the frequency of cell division in growing cells and inhibited spore cell differentiation during late development. Overexpression of PsaA-GFP (where GFP is green fluorescent protein) also inhibited spore cell differentiation but did not affect growth. Using chimeras, we have identified that nuclear versus cytoplasmic localization of PsaA affects the choice between stalk or spore cell differentiation pathway. Cells that overexpressed PsaA-GFP (primarily nuclear) differentiated into stalk cells, while cells that overexpressed PsaAΔNLS2-GFP (cytoplasmic) differentiated into spores. In conclusion, we have identified that BME inhibits cell growth, division, and differentiation in Dictyostelium likely through inhibition of PsaA.     

Complex genotype interactions influence social fitness during the developmental phase of the social amoeba Dictyostelium discoideum

When individuals interact, phenotypic variation can be partitioned into direct genetic effects (DGEs) of the individuals’ own genotypes, indirect genetic effects (IGEs) of their social partners’ genotypes and epistatic interactions between the genotypes of interacting individuals (‘genotype‐by‐genotype (G×G) epistasis’). These components can all play important roles in evolutionary processes, but few empirical studies have examined their importance. The social amoeba Dictyostelium discoideum provides an ideal system to measure these effects during social interactions and development. When starved, free‐living amoebae aggregate and differentiate into a multicellular fruiting body with a dead stalk that holds aloft viable spores. By measuring interactions among a set of natural strains, we quantify DGEs, IGEs and G×G epistasis affecting spore formation. We find that DGEs explain most of the phenotypic variance (57.6%) whereas IGEs explain a smaller (13.3%) but highly significant component. Interestingly, G×G epistasis explains nearly a quarter of the variance (23.0%), highlighting the complex nature of genotype interactions. These results demonstrate the large impact that social interactions can have on development and suggest that social effects should play an important role in developmental evolution in this system.     

Taking the plunge. Terminal differentiation in Dictyostelium

During the last stage of Dictyostelium development a motile, cylindrical slug transforms into an immotile, stalked fruiting body and the constituent cells change from amoebae to either refractile spores or vacuolated stalk cells. Analysis of this process using genetics and simple culture techniques is becoming a powerful way of investigating a number of conserved signal transduction processes. A common pathway activating cAMP-dependent protein kinase (PKA) triggers the maturation of spore cells and those stalk cells forming the stalk. It uses a eukaryotic version of the 'bacterial' two-component phospho-relay system to control cAMP breakdown. A second pathway, inhibiting the GSK3 protein kinase, might control the maturation of a distinct set of stalk cells at the base of the fruiting body.     

Interspecies and intraspecies interactions in social amoebae

The stable co‐existence of individuals of different genotypes and reproductive division of labour within heterogeneous groups are issues of fundamental interest from the viewpoint of evolution. Cellular slime moulds are convenient organisms in which to address both issues. Strains of a species co‐occur, as do different species; social groups are often genetically heterogeneous. Intra‐ and interspecies 1 : 1 mixes of wild isolates of Dictyostelium giganteum and D. purpureum form chimaeric aggregates, following which they segregate to varying extents. Intraspecies aggregates develop in concert and give rise to chimaeric fruiting bodies that usually contain more spores (reproductives) of one component than the other. Reproductive skew and variance in the proportion of reproductives are positively correlated. Interspecies aggregates exhibit almost complete sorting; most spores in a fruiting body come from a single species. Between strains, somatic compatibility correlates weakly with sexual compatibility. It is highest within clones, lower between strains of a species and lowest between strains of different species. Trade‐offs among fitness‐related traits (between compatible strains), sorting out (between incompatible strains) and avoidance (between species) appear to lie behind coexistence.