Interacting proteins of protein kinase A regulatory subunit in Saccharomyces cerevisiae

Abstract

cAMP-dependent protein kinase mediates many extracellular signals in eukaryotes. The compartmentalization of PKA is an important level of control of the specificity of signal transduction mediated by cAMP. Unlike mammalian PKA for which proof insights in the mechanism that controls its localization through anchoring proteins (AKAPs) has been obtained, in the case of Saccharomyces cerevisiae PKA there was little information available. In this work, we present results that demonstrate the isolation and identification of yeast PKA regulatory subunit (Bcy1) associated proteins using a MS-based proteomic analysis and a bioinformatic approach. The verification of some of these interactions was assessed by immunoprecipitation, pull down and co-localization by subcellular fractionation. The key role of positively charged residues present in the interaction domain of the identified proteins was demonstrated. The defined interaction domain has therefore different molecular characteristics than conventional AKAP domains. Finally we assess initial experiments to visualize the physiological relevance of the interaction of both Ira2 and Hsp60 with Bcy1. Bcy1 interacts with Ira2 tethering PKA to the Ras complex and Hsp60 chaperone localizes PKA to mitochondria and has a role in the kinase stability.