Interaction of gdt1 and protein kinase A (PKA) in the growth-differentiation-transition in Dictyostelium

The gdt1 gene is a negative regulator of the growth-differentiation-transit ion (GDT) in Dictyostelium. gdt1(-) cells express the GDT marker discoidin earlier and at higher levels and prematurely enter the differentiation path way. Protein kinase A is a positive regulator of the CDT and is required fo r multicellular development. Disruption of the PKA catalytic subunit or ove rexpression of a constitutively active mutant of the regulatory subunit res ults in cells which do not form multicellular aggregates and which show str ongly reduced levels of discoidin. We have created PKA(-)/gdt1(-) double mu tants and show that these display high levels of discoidin expression but n o aggregation, suggesting that gdt1 may be a downstream target of PKA in a branched signalling cascade initiating differentiation. Data obtained with the PKA inhibitor H89 support these result: in wild type cells H89 inhibits discoidin expression while in gdt1(-) mutants there is no obvious effect. However, since PKA(-)/gdt1(-) cells display less discoidin expression than the single gdt1 mutant, we propose that PKA and gdt1 are in two parallel in teracting pathways. To get insight into the mechanism how PKA may block gdt1, we have tested tw o putative PKA phosphorylation sites in the protein and found that one of t hem is efficiently phosphorylated by PKA in vitro. A model for the interpla y between PKA and gdt1 during the growth-differentiation-transition is disc ussed.