Extracellular cAMP is sufficient to restore developmental gene expression and morphogenesis in Dictyostelium cells lacking the aggregation adenylyl cyclase (ACA).

Pitt, Geoffrey S.; Brandt, Raymond; Lin, Kenneth; Devreotes, Peter N.; Schaap, Pauline

doi:10.1101/gad.7.11.2172

Cited by 77 publications

(69 citation statements)

References 32 publications

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“…Since basal expression of these genes is completely absent in RM cells, while a low level of cAMP-pulse induced transcription is retained; it seems that also here PKA may be required for basal induction by CMF or PSF, rather than for pulse-induced expression. This is supported by findings that pulse-induced gene expression is normal in ACA null mutants [24]. PSF has not yet been characterized, but CMF encodes an 80 kDA glycoprotein.…”

Section: Resultssupporting

confidence: 67%

cAMP‐dependent protein kinase activity is essential for preaggregative gene expression in Dictyostelium

Schulkes

Schaap

1995

FEBS Letters

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Section: Resultssupporting

confidence: 67%

cAMP‐dependent protein kinase activity is essential for preaggregative gene expression in Dictyostelium

Schulkes

Schaap

1995

FEBS Letters

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“…Overexpression of PKA yields a more rapid, extended, and higher level of activation, suggesting that PKA may be essential for maintaining low basal levels and maximally stimulating ERK2 activation. The more extended activation in pka null cells may be associated with other roles of PKA, which is known to be required for multiple aspects of aggregation and later multicellular development (19,28).…”

Section: Discussionmentioning

confidence: 99%

The Dictyostelium Mitogen-activated Protein Kinase ERK2 Is Regulated by Ras and cAMP-dependent Protein Kinase (PKA) and Mediates PKA Function

Aubry

Maeda

Insall

et al. 1997

Journal of Biological Chemistry

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The chemoattractant cAMP, acting through serpentine cAMP receptors, results in a rapid and transient stimulation of the Dictyostelium mitogen-activated protein kinase ERK2 activity (1). In this study we show that other pathways required for aggregation, including Ras and cAMP-dependent protein kinase (PKA), are important regulators of ERK2 activation and adaptation. By examining both the level and kinetics of activation and adaptation of ERK2, we show that Ras is a negative regulator of ERK2. Activated Ras or disruption of a Ras GAP gene results in reduced ERK2 activation whereas disruption of putative Ras GEF or expression of dominant negative Ras proteins have a more rapid, higher, and extended activation. CRAC, a PH domain-containing protein required for adenylyl cyclase activation, is also required for proper ERK2 adaptation. PKA overexpression results in a more rapid, higher level of activation, whereas pka null cells show a lower level but more extended ERK2 activation. Furthermore, we show that constitutive expression of PKA catalytic subunit bypasses the requirement of ERK2 for aggregation and later development, indicating that PKA lies downstream from ERK2 and that ERK2 may regulate one or more components of the signaling pathway required for mediating PKA function, possibly by directly regulating PKA R or a protein controlling the intracellular level of cAMP.A cell's ability to respond to an extracellular signal involves both the activation of pathways and subsequent adaptation in which the cells are no longer fully responsive to the extracellular signal. This allows the cells to properly regulate the level and extent of the signaling pathway as well as adapt to changing environmental conditions. Well known examples are the pheromone pathway in yeast and the mammalian cell ␤-adrenergic receptor (2, 3). In Dictyostelium, aggregation is mediated by a periodic activation and adaptation of pathways regulated by G protein-coupled cARs 1 that bind extracellular cAMP as the ligand (4 -6). In response to extracellular cAMP, guanylyl and adenylyl cyclase activities are rapidly stimulated and then adapted. If cAMP levels are kept constant the cells remain nonresponsive, whereas removal of the cAMP, which in vivo occurs through its hydrolysis by an extracellular phosphodiesterase, allows the pathways to de-adapt within ϳ5 min.MAP kinase cascades regulate a variety of intracellular responses through the activation of cell surface receptors (1, 2, 7). During the preaggregation and aggregation stages of Dictyostelium development, the MAP kinase ERK2 is activated through cAMP and folate chemotactic receptors (1). 2 Stimulation with extracellular cAMP results in a Ͼ40-fold increase in ERK2 activity that peaks at ϳ50 s and thereupon adapts, reaching its basal level within 5-8 min. ERK2 activation requires the G protein-coupled cAMP receptors that mediate cAMP-stimulated adenylyl and guanylyl cyclase activation, chemotaxis, and gene expression. cAMP-mediated activation of ERK2 is partially independent of G␣2, the G␣ sub...

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“…However, these cells can respond to cAMP signals: after repeated treatment with extracellular cAMP, the cells will form aggregates and multicellular structures (Pitt et al, 1993). To determine whether such treatment with extracellular stimuli can rescue the SN mutants, the cells were starved in suspension, provided with exogenous cAMP at 6-min intervals for 5 h, and then plated on nonnutrient agar.…”

Section: Developmental Phenotypes Of Sn Mutant Cells After Treatment mentioning

confidence: 99%

“…When a mixture of G␤ null cells and wild-type cells is allowed to develop together, the spores from the resulting fruiting bodies are all derived from wildtype cells . In contrast, synag mutant cells, such as aca Ϫ and crac Ϫ cells, cannot aggregate alone but are able to form chimeric structures when mixed with wild-type cells (Pitt et al, 1993;Insall et al, 1994). To isolate G␤ alleles that resemble the phenotype seen in synag mutants, we transformed cells of a G␤ null parent strain with a library of randomly mutagenized G␤ cDNA and screened for transformants The G␤ null cells were transformed with a library of randomly mutagenized g␤ cDNA.…”

Section: Isolation Of Sn G␤ Mutationsmentioning

confidence: 99%

“…Cells impaired in ACA activation because of deletion of the genes encoding the enzyme ACA or cytosolic proteins that are required for ACA activation, such as Aimless (a RasGEF), CRAC, and Pianissimo, are defective in producing cAMP signals and do not aggregate. However, unlike G␤ null cells, they can respond to cAMP signals generated by wildtype cells in a mixture and can differentiate into viable spores in chimeric fruiting bodies (Pitt et al, 1993;Insall et al, 1994Insall et al, , 1996Chen et al, 1997). These mutants are designated as "synag.…”

Section: Introductionmentioning

confidence: 99%

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Selection of Gβ Subunits with Point Mutations That Fail to Activate Specific Signaling Pathways In Vivo: Dissecting Cellular Responses Mediated by a Heterotrimeric G Protein inDictyostelium discoideum

Jin¹,

Amzel

Devreotes³

et al. 1998

MBoC

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In Dictyostelium discoideum, a unique G␤ subunit is required for a G protein-coupled receptor system that mediates a variety of cellular responses. Binding of cAMP to cAR1, the receptor linked to the G protein G2, triggers a cascade of responses, including activation of adenylyl cyclase, gene induction, actin polymerization, and chemotaxis. Null mutations of the cAR1, G␣2, and G␤ genes completely impair all these responses. To dissect specificity in G␤␥ signaling to downstream effectors in living cells, we screened a randomly mutagenized library of G␤ genes and isolated G␤ alleles that lacked the capacity to activate some effectors but retained the ability to regulate others. These mutant G␤ subunits were able to link cAR1 to G2, to support gene expression, and to mediate cAMP-induced actin polymerization, and some were able to mediate to chemotaxis toward cAMP. None was able to activate adenylyl cyclase, and some did not support chemotaxis. Thus, we separated in vivo functions of G␤␥ by making point mutations on G␤. Using the structure of the heterotrimeric G protein displayed in the computer program CHAIN, we examined the positions and the molecular interactions of the amino acids substituted in each of the mutant G␤s and analyzed the possible effects of each replacement. We identified several residues that are crucial for activation of the adenylyl cyclase. These residues formed an area that overlaps but is not identical to regions where bovine Gt␤␥ interacts with its regulators, G␣ and phosducin.

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Extracellular cAMP is sufficient to restore developmental gene expression and morphogenesis in Dictyostelium cells lacking the aggregation adenylyl cyclase (ACA).

Cited by 77 publications

References 32 publications

cAMP‐dependent protein kinase activity is essential for preaggregative gene expression in Dictyostelium

cAMP‐dependent protein kinase activity is essential for preaggregative gene expression in Dictyostelium

The Dictyostelium Mitogen-activated Protein Kinase ERK2 Is Regulated by Ras and cAMP-dependent Protein Kinase (PKA) and Mediates PKA Function

Selection of Gβ Subunits with Point Mutations That Fail to Activate Specific Signaling Pathways In Vivo: Dissecting Cellular Responses Mediated by a Heterotrimeric G Protein inDictyostelium discoideum

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