Activated Alleles of the Schizosaccharomyces pombe gpa2(+) G alpha Gene Identify Residues Involved in GDP-GTP Exchange

作者：Ivey F Douglas; Taglia Francis X; Yang Fan; Lander Matthew M; Kelly David A; Hoffman Charles S^*

来源：Eukaryotic Cell, 2010, 9(4): 626-633.

DOI：10.1128/EC.00010-10

The Schizosaccharomyces pombe glucose/cyclic AMP (cAMP) signaling pathway includes the Gpa2-Git5-Git11 heterotrimeric G protein, whose Gpa2 G alpha subunit directly binds to and activates adenylate cyclase in response to signaling from the Git3 G protein-coupled receptor. To study intrinsic and extrinsic regulation of Gpa2, we developed a plasmid-based screen to identify mutationally activated gpa2 alleles that bypass the loss of the Git5-Git11 G beta gamma dimer to repress transcription of the glucose-regulated fbp1(+) gene. Fifteen independently isolated mutations alter 11 different Gpa2 residues, with all but one conferring a receptor-independent activated phenotype upon integration into the gpa2(+) chromosomal locus. Biochemical characterization of three activated Gpa2 proteins demonstrated an increased GDP-GTP exchange rate that would explain the mechanism of activation. Interestingly, the amino acid altered in the Gpa2(V90A) exchange rate mutant protein is in a region of Gpa2 with no obvious role in G alpha function, thus extending our understanding of G alpha protein structure-function relationships.

• 出版日期2010-4