Genes involved in cell type specification and morphogenesis in Dictyostelium discoideum

Abstract

Calcium-Dependent Regulation of Gene Expression and Morphogenesis in Dictyostelium discoideum

Abstract and Synopsis Introduction Differentiation and morphogenesis occur in concert to ensure the development of multicellular organisms. Dictyostelium discoideum, a simple eukaryote, illustrates features of multicellular development common to metazoa. Its life cycle begins with a unicellular growth phase, followed by a multicellular phase triggered by starvation. Amoebae aggregate via chemotaxis to cyclic AMP (cAMP), forming a slug with anterior presumptive stalk (pst) cells and posterior presumptive spore (psp) cells. Calcium levels influence cell fate: high calcium predisposes cells to pst fate, while low calcium favors psp fate.

Calcium and Gene Expression

RAP-PCR Analysis: Identified cDNAs differentially expressed under altered calcium conditions.

Alpha Mannosidase: Repressed by high calcium; regulation at transcript level matched enzyme activity.

Beta Glucosidase: Similarly repressed by high calcium and induced by low calcium.

Pharmacological Agents: BHQ altered enzyme activity, implicating contractile vacuole calcium stores.

PSF-Responsive Genes: Alpha mannosidase, cAMP phosphodiesterase (PDE), and discoidin I were all repressed by high calcium and induced by low calcium, though quantitative responses varied.

Novel Calcium-Regulated Gene

A partial cDNA repressed by low calcium corresponded to a full-length gene encoding two ORFs: one similar to asparaginyl tRNA synthetase and the other to glutaminyl tRNA synthetase.

These represent the first aminoacyl tRNA synthetases with calcium-dependent expression.

Random Mutagenesis and the trishanku (tri) Mutant

REMI mutagenesis identified mutants with stalky fruiting bodies.

The tri mutant showed defects in aggregation and terminal differentiation: aggregates broke apart, and spore masses failed to ascend the stalk.

The tri gene encodes a novel protein with a BTB protein-protein interaction domain.

Knockout and ectopic expression confirmed its role.

Expression analysis:

Weak during growth phase.

Developmentally regulated, peaking at slug stage.

Primarily expressed in prespore cells.

Phenotype analysis suggested increased trans-differentiation between pst and psp cells rather than improper cell fate choice.

The mutation was cell-autonomous, as wild-type cells could not rescue the phenotype in chimeras.

Calcium alteration in tri null cells revealed an additional morphogenetic role of calcium in regulating spore mass ascent.

Conclusions Calcium regulates lysosomal enzymes and PSF-responsive genes, repressing activity at high levels and inducing activity at low levels.

Novel calcium-dependent regulation was identified in aminoacyl tRNA synthetases.

The tri gene plays a critical role in morphogenesis, with its mutation leading to stalky phenotypes and defective spore mass ascent.

Calcium influences both differentiation and morphogenetic processes, linking cell fate decisions with structural development.