Characterization of the molecular mechanisms involved in the regulation of AKAP‐Lbc activity

Abstract

A-kinase anchoring proteins (AKAPs) are a family of functionally related proteins which are classified on the basis of their ability to associate with the PKA holoenzyme inside cells. By tethering the holoenzyme at precise cell locations, in close proximity to physiological substrates, AKAPs favor specific PKA phosphorylation events. We recently identified a novel AKAP, called AKAP-Lbc, which also functions as a Rho-selective guanine nucleotide exchange factor (GEF). GEFs of the Dbl family are upstream activators of small GTPases of the Rho family and play fundamental roles in signaling pathways controlling many physiological processes such as cell proliferation, motility, growth and development. We demonstrated that AKAP-Lbc nucleates two opposite signaling pathways thereby controlling precisely the cellular level of GTP-bound RhoA. Accordingly, we previously showed that AKAP-Lbc Rho-GEF activity can be stimulated by the activated alpha-subunit of the heterotrimeric G protein G12, whereas 14-3-3 binding to AKAP-Lbc is required to maintain AKAP-Lbc inactive under basal conditions. Here we report that AKAP-Lbc forms homo-oligomers through a leucine zipper motif located in its C-terminal region and that homo-oligomerization maintains AKAP-Lbc in an inactive state under unstimulated conditions. Importantly, we could demonstrate that AKAP-Lbc oligomerization deficient mutants are completely resistant to 14-3-3-mediated inhibition. Taken together, our findings indicate that 14-3-3 can exert a negative regulation on AKAP-Lbc only when AKAP-Lbc adopts an oligomeric conformation.