T Kon1, H Adachi, K Sutoh. 1. Department of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, 3-8-1 Komaba, Meguro-ku, Tokyo 153-8902, Japan.
Abstract
BACKGROUND: The differentiation programme of Dictyostelium discoideum is initiated by starvation. Nutrient depletion triggers the differentiation of Dictyostelium cells through the transcriptional inactivation of some growth-phase genes, as well as through the transcriptional activation of essential genes required for the aggregation of the cells. The adenylyl cyclase (ACA) gene, acaA, is one of the earliest genes expressed following starvation. ACA produces intracellular and extracellular cAMP that drives further differentiation by inducing chemotaxis, developmental gene expression and morphogenesis of Dictyostelium cells. Although several genes have been identified as being essential for the initiation of differentiation process, such as the transcriptional activation of ACA expression, the molecular mechanisms of the growth/differentiation transition remain to be explored. RESULTS: Using insertional mutagenesis, we have isolated a mutant that does not aggregate upon starvation. The disrupted gene, amiB (aggregation minus B), is predicted to encode a novel protein of 298.9 kDa. When starved, amiB- cells produced an undetectable level of cAMP. Analyses of gene expression showed that amiB- cells fail to turn off the expression of one of the growth-phase genes, cprD, and to turn on the expression of ACA following starvation. The ectopic expression of ACA from a constitutive promoter rescued the differentiation and morphogenesis of amiB- mutants. Furthermore, the ectopic expression of a putative transcriptional factor DdMyb2 or a catalytic subunit of cAMP-dependent protein kinase (PKA-C), both of which are thought to be involved in ACA expression pathway(s), also rescued the starvation-induced ACA expression and further differentiation of the amiB- mutant. CONCLUSION: These results suggest that AmiB plays a role at the start of Dictyostelium differentiation through induction of the ACA expression which is essential for cAMP signalling.
BACKGROUND: The differentiation programme of Dictyostelium discoideum is initiated by starvation. Nutrient depletion triggers the differentiation of Dictyostelium cells through the transcriptional inactivation of some growth-phase genes, as well as through the transcriptional activation of essential genes required for the aggregation of the cells. The adenylyl cyclase (ACA) gene, acaA, is one of the earliest genes expressed following starvation. ACA produces intracellular and extracellular cAMP that drives further differentiation by inducing chemotaxis, developmental gene expression and morphogenesis of Dictyostelium cells. Although several genes have been identified as being essential for the initiation of differentiation process, such as the transcriptional activation of ACA expression, the molecular mechanisms of the growth/differentiation transition remain to be explored. RESULTS: Using insertional mutagenesis, we have isolated a mutant that does not aggregate upon starvation. The disrupted gene, amiB (aggregation minus B), is predicted to encode a novel protein of 298.9 kDa. When starved, amiB- cells produced an undetectable level of cAMP. Analyses of gene expression showed that amiB- cells fail to turn off the expression of one of the growth-phase genes, cprD, and to turn on the expression of ACA following starvation. The ectopic expression of ACA from a constitutive promoter rescued the differentiation and morphogenesis of amiB- mutants. Furthermore, the ectopic expression of a putative transcriptional factor DdMyb2 or a catalytic subunit of cAMP-dependent protein kinase (PKA-C), both of which are thought to be involved in ACA expression pathway(s), also rescued the starvation-induced ACA expression and further differentiation of the amiB- mutant. CONCLUSION: These results suggest that AmiB plays a role at the start of Dictyostelium differentiation through induction of the ACA expression which is essential for cAMP signalling.
Authors: Thomas Winckler; Negin Iranfar; Peter Beck; Ingo Jennes; Oliver Siol; Unha Baik; William F Loomis; Theodor Dingermann Journal: Eukaryot Cell Date: 2004-10