Editors' ChoiceProtein kinase A

A New Route to PKA Activation

Science's STKE  27 Sep 2005:
Vol. 2005, Issue 303, pp. tw343
DOI: 10.1126/stke.3032005tw343

Profirovic et al. uncovered a pathway from α-thrombin to cAMP-dependent protein kinase (PKA)-mediated phosphorylation of vasodilator-stimulated phosphoprotein (VASP), a membrane-associated protein implicated in regulation of actin structure and cell motility, that appears to be independent of adenosine 3′,5′-monophosphate (cAMP). α-Thrombin stimulation of cultured human vascular endothelial cells (HUVECs) elicited phosphorylation-dependent translocation of VASP to the cell periphery. α-Thrombin-dependent VASP phosphorylation was inhibited by expression of the regulators of G protein signaling (RGS) domain of p115RhoGEF, implicating Gα12 or Gα13 in the response. Moreover, constitutively active Gα13 stimulated VASP phosphorylation in human embryonic kidney (HEK) 293 cells and phosphorylation-dependent translocation in HUVECs. Depleting RhoA with small interfering RNA (siRNA) abolished VASP phosphorylation in response to α-thrombin, and expression of the botulinum C3 exotoxin (which inactivates RhoA) inhibited Gα13-mediated VASP phosphorylation, whereas expression of constitutively active RhoA promoted VASP phosphorylation. α-Thrombin-, Gα13-, and RhoA-mediated VASP phosphorylation were all sensitive to inhibition of PKA, and constitutively active Gα13 or RhoA stimulated PKA-dependent transcriptional activation of a reporter. However, neither α-thrombin nor constitutively active Gα13 or RhoA produced a detectable increase in intracellular cAMP. Previous research indicates that some PKA catalytic subunits exist in a complex with inhibitor of nuclear factor κB (IκB) and nuclear factor κB (NF-κB). Constitutively active Gα13 or RhoA promoted NF-κB activation, whereas a dominant-negative IκBα mutant (IκBαm, which inhibits IκB phosphorylation and degradation and the release of PKAc) abolished VASP phosphorylation in response to Gα13 or RhoA. Gα13-dependent VASP phosphorylation was blocked by a dominant-negative mutant of the mitogen-activated protein kinase kinase MEKK1, whereas activated MEKK1 stimulated IκBαm-sensitive VASP phosphorylation. Thus the authors conclude that α-thrombin triggers a novel pathway to PKA-mediated VASP phosphorylation that involves RhoA and MEKK1 activation and degradation of IκB.

J. Profirovic, M. Gorovoy, J. Niu, S. Pavlovic, T. Voyno-Yasenetskaya, A novel mechanism of G protein-dependent phosphorylation of vasodilator-stimulated phosphoprotein. J. Biol. Chem. 280, 32866-32876 (2005). [Abstract] [Full Text]