Role of cAMP-Dependent Protein Kinase in Controlling Aggregation and Postaggregative Development inDictyostelium

Abstract

We have examined the role of cAMP-dependent protein kinase (PKA) in controlling aggregation and postaggregative development inDictyostelium.We previously showed that cells in which the gene encoding the PKA catalytic subunit has been disrupted (pkacat−cells) are unable to aggregate [S. K.O. Mann and R.A. Firtel (1991). A developmentally regulated, putative serine/threonine protein kinase is essential for development inDictyostelium. Mech. Dev.35, 89–102]. We show thatpkacat−cells are unable to activate adenylyl cyclase in response to cAMP stimulation due to the inability to express the aggregation-stage, G-protein-stimulated adenylyl cyclase (ACA). Constitutive expression of ACA from an actin promoter results in a high level of Mn2+-stimulated adenylyl cyclase activity and restores chemoattractant- and GTPγS-stimulated adenylyl cyclase activity but not the ability to aggregate. Similarly, expression of the constitutively active, non-G protein-coupled adenylyl cyclase ACG inpkacat−cells also does not restore the ability to aggregate, although ACG can complement cells in which theACAgene has been disrupted. These results indicate thatpkacat−cells lack multiple, essential aggregation-stage functions. As the mound forms, high, continuous levels of extracellular cAMP functioning through the cAMP serpentine receptors activate a transcriptional cascade that leads to cell-type differentiation and morphogenesis. The first step is the induction and activation of the transcription factor GBF and downstream postaggregative genes, followed by the induction of prestalk- and prespore-specific genes. We show thatpkacat−cells induce postaggregative gene expression in response to exogenous cAMP, but the level of induction of some of these genes, includingGBF,is reduced.SP60(a prespore-specific gene) is not induced andecmA(a prestalk-specific gene) is induced to very low levels. Expressing GBF constitutively inpkacat−cells restoresecmAexpression to a moderate level, butSP60is not detectably induced. Overexpression of PKAcat from the Actin 15 (Act15), ecmAprestalk, and thePKAcatpromoters inpkacat−cells results in significant aberrant spatial patterning of prestalk and prespore cells, as determined bylacZ reporter studies. Our studies identify new, essential regulatory roles for PKA in mediating multicellular development.