A developmentally regulated, putative serine/threonine protein kinase is essential for development in Dictyostelium.
Using PCR technology, we have cloned parts of three developmentally regulated putative serine/threonine kinases from Dictyostelium. All show significant homology to members of the cAMP-dependent protein kinase A/protein kinase C subfamilies. A genomic clone encoding one of these, DdPK3, has been isolated and sequenced. The open reading frame encodes a protein of 648 amino acids with the conserved kinase domain in the C-terminal half. The protein encoded by this gene is unusual in that it contains long homopolymer runs in the N-terminal half of the protein, including a long run of 88 amino acids in which 73 are glutamine residues. To examine the function of DdPK3, a gene disruption was created via homologous recombination. Ddpk3- cells do not aggregate by themselves but will co-aggregate with wild-type cells. However, after aggregation these cells are 'sloughed off' and do not proceed further through development, but are found as a discrete mass alongside the fruiting body formed by the wild-type cells. Analysis of signal transduction pathways indicates that cAMP pulse-induced expression of aggregation stage-specific genes is normal in Ddpk3- cells, as is induction of the prestalk gene Ddras in single cell assays. However, cAMP induction of the late promoters of cAMP receptor cAR1 and of two prespore-specific genes is absent under similar conditions. These cells show normal activation of adenylate cyclase and normal phosphorylation of the G alpha protein G alpha 2 in response to cAMP. The possible role of DdPK3 in Dictyostelium development is discussed.
Mann
S K
SK
Department of Biology, University of California, San Diego, La Jolla 92093.
Firtel
R A
RA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004023:Dictyostelium / Q000201:enzymology / Q000254:growth & development*; D005656:Fungal Proteins / Q000378:metabolism*; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011494:Protein Kinases / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases; D015398:Signal Transduction
D005656:Fungal Proteins; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases