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Kelch-repeat proteins interacting with the G protein Gpa2 bypass adenylate cyclase for direct regulation of protein kinase A in yeast
Tom Peeters,
Wendy Louwet,
Ruud Geladé,
David Nauwelaers,
Johan M. Thevelein, and
Matthias Versele*
Laboratory of Molecular Cell Biology, Katholieke Universiteit Leuven, and Department of Molecular Microbiology, Flanders Interuniversity Institute of Biotechnology (VIB), Kasteelpark Arenberg 31, B-3001 Leuven-Heverlee, Belgium
Edited by Gregory A. Petsko, Brandeis University, Waltham, MA, and approved July 5, 2006
Received for publication November 7, 2005.
Abstract:
The cAMP–PKA pathway consists of an extracellular ligand-sensitiveG protein-coupled receptor, a G protein signal transmitter,and the effector, adenylate cyclase, of which the product, cAMP,acts as an intracellular second messenger. cAMP activates PKAby dissociating the regulatory subunit from the catalytic subunit.Yeast cells (Saccharomyces cerevisiae) contain a glucose/sucrose-sensitiveseven-transmembrane domain receptor, Gpr1, that was proposedto activate adenylate cyclase through the G protein Gpa2. Consistently,we show here that adenylate cyclase binds only to active, GTP-boundGpa2. Two related kelch-repeat proteins, Krh1/Gpb2 and Krh2/Gpb1,are associated with Gpa2 and were suggested to act as G mimicsfor Gpa2, based on their predicted seven-bladed -propeller structure.However, we find that although Krh1 associates with both GDPand GTP-bound Gpa2, it displays a preference for GTP-Gpa2. Thestrong down-regulation of PKA targets by Krh1 and Krh2 doesnot require Gpa2 but is strictly dependent on both the catalyticand the regulatory subunits of PKA. Krh1 directly interactswith PKA by means of the catalytic subunits, and Krh1/2 stimulatethe association between the catalytic and regulatory subunitsin vivo. Indeed, both a constitutively active GPA2 allele anddeletion of KRH1/2 lower the cAMP requirement of PKA for growth.We propose that active Gpa2 relieves the inhibition imposedby the kelch-repeat proteins on PKA, thereby bypassing adenylatecyclase for direct regulation of PKA. Importantly, we show thatKrh1/2 also enhance the association between mouse R and C subunits,suggesting that Krh control of PKA has been evolutionarily conserved.
Key Words:Saccharomyces cerevisiae • signal transduction • glucose
Freely available online through the PNAS open access option.
Author contributions: T.P., W.L., R.G., D.N., J.M.T., and M.V.designed research; T.P., W.L., R.G., D.N., and M.V. performedresearch; T.P., W.L., R.G., D.N., J.M.T., and M.V. analyzeddata; and J.M.T. and M.V. wrote the paper.
Conflict of interest statement: No conflicts declared.
This paper was submitted directly (Track II) to the PNAS office.
*To whom correspondence should be addressed. E-mail: matthias.versele{at}bio.kuleuven.be
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protein Gpa2 bypass adenylate cyclase for direct regulation of protein kinase A in yeast
protein Gpa2. Consistently, we show here that adenylate cyclase binds only to active, GTP-bound Gpa2. Two related kelch-repeat proteins, Krh1/Gpb2 and Krh2/Gpb1, are associated with Gpa2 and were suggested to act as G
mimics for Gpa2, based on their predicted seven-bladed 
