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AKAP-Mediated Targeting of Protein Kinase A Regulates Contractility in Cardiac Myocytes

2001; Lippincott Williams & Wilkins; Volume: 88; Issue: 3 Linguagem: Inglês

10.1161/01.res.88.3.291

1524-4571

Mary Alexander Fink, Daniel R. Zakhary, Julie A. Mackey, Mary A. Russell, Carolyn Apperson-Hansen, Derek S. Damron, Meredith Bond,

Ion channel regulation and function

Abstract —Compartmentalization of cAMP-dependent protein kinase A (PKA) by A-kinase anchoring proteins (AKAPs) targets PKA to distinct subcellular locations in many cell types. However, the question of whether AKAP-mediated PKA anchoring in the heart regulates cardiac contractile function has not been addressed. We disrupted AKAP-mediated PKA anchoring in cardiac myocytes by introducing, via adenovirus-mediated gene transfer, Ht31, a peptide that binds the PKA regulatory subunit type II (RII) with high affinity. This peptide competes with endogenous AKAPs for RII binding. Ht31P (a proline-substituted derivative), which does not bind RII, was used as a negative control. We then investigated the effects of Ht31 expression on RII distribution, Ca 2+ cycling, cell shortening, and PKA-dependent substrate phosphorylation. By confocal microscopy, we showed redistribution of RII from the perinuclear region and from periodic transverse striations in Ht31P-expressing cells to a diffuse cytosolic localization in Ht31-expressing cells. In the presence of 10 nmol/L isoproterenol, Ht31-expressing myocytes displayed an increased rate and amplitude of cell shortening and relaxation compared with control cells (uninfected and Ht31P-expressing myocytes); with isoproterenol stimulation we observed decreased time to 90% decline in Ca 2+ but no significant difference between Ht31-expressing and control cells in the rate of Ca 2+ cycling or amplitude of the Ca 2+ transient. The increase in PKA-dependent phosphorylation of troponin I and myosin binding protein C on isoproterenol stimulation was significantly reduced in Ht31-expressing cells compared with controls. Our results demonstrate that, in response to β-adrenergic stimulation, cardiomyocyte function and substrate phosphorylation by PKA is regulated by targeting of PKA by AKAPs.