A novel cytosolic regulator, Pianissimo, is required for chemoattractant receptor and G protein-mediated activation of the 12 transmembrane domain adenylyl cyclase in Dictyostelium
Abstract
Genetic analysis was applied to identify novel genes involved in G protein-linked pathways controlling development. Using restriction enzyme-mediated integration (REMI), we have identified a new gene,Pianissimo (PiaA), involved in cAMP signaling inDictyostelium discoideum. PiaA encodes a 130-kD cytosolic protein required for chemoattractant receptor and G protein-mediated activation of the 12 transmembrane domain adenylyl cyclase. InpiaA − null mutants, neither chemoattractant stimulation of intact cells nor GTPγS treatment of lysates activates the enzyme; constitutive expression of PiaA reverses these defects. Cytosols of wild-type cells that contain Pia protein reconstitute the GTPγS stimulation of adenylyl cyclase activity in piaA −lysates, indicating that Pia is directly involved in the activation. Pia and CRAC, a previously identified cytosolic regulator, are both essential for activation of the enzyme as lysates of crac−piaA − double mutants require both proteins for reconstitution. Homologs of PiaA are found in Saccharomyces cerevisiaeand Schizosaccaromyces pombe; disruption of the S. cerevisiae homolog results in lethality. We propose that homologs of Pia and similar modes of regulation of these ubiquitous G protein-linked pathways are likely to exist in higher eukaryotes.
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↵1 Corresponding author.
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E-MAIL pnd{at}welchlink.welch.jhu.edu; FAX (410) 955-5759.
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- Received June 13, 1997.
- Accepted September 18, 1997.
- Cold Spring Harbor Laboratory Press