InsP3R-associated cGMP kinase substrate determines inositol 1,4,5-trisphosphate receptor susceptibility to phosphoregulation by cyclic nucleotide-dependent kinases

Abstract

Ca2+ release through inositol 1,4,5-trisphosphate receptors (InsP3R) can be modulated by numerous factors, including input from other signal transduction cascades. These events shape the spatio-temporal characteristics of the Ca2+ signal and provide fidelity essential for the appropriate activation of effectors. In this study, we investigate the regulation of Ca2+ release via InsP3R following activation of cyclic nucleotide-dependent kinases in the presence and absence of expression of a binding partner InsP3R-associated cGMP kinase substrate (IRAG). cGMP-dependent kinase (PKG) phosphorylation of only the S2+ InsP3R-1 subtype resulted in enhanced Ca2+ release in the absence of IRAG expression. In contrast, IRAG bound to each InsP3R subtype, and phosphorylation of IRAG by PKG attenuated Ca2+ release through all InsP3R subtypes. Surprisingly, simply the expression of IRAG attenuated phosphorylation and inhibited the enhanced Ca2+ release through InsP3R-1 following cAMP-dependent protein kinase (PKA) activation. In contrast, IRAG expression did not influence the PKA-enhanced activity of the InsP3R-2. Phosphorylation of IRAG resulted in reduced Ca2+ release through all InsP3R subtypes during concurrent activation of PKA and PKG, indicating that IRAG modulation is dominant under these conditions. These studies yield mechanistic insight into how cells with various complements of proteins integrate and prioritize signals from ubiquitous signaling pathways. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.