期刊
CIRCULATION RESEARCH
卷 88, 期 3, 页码 291-297出版社
LIPPINCOTT WILLIAMS & WILKINS
DOI: 10.1161/01.RES.88.3.291
关键词
A-kinase anchoring proteins; protein kinase A; cardiac myocyte; beta-adrenergic receptor; contractility
资金
- NHLBI NIH HHS [F32-HL10236, R01 HL 56256] Funding Source: Medline
- NIA NIH HHS [R01 AG 16613] Funding Source: Medline
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, Ca2+ 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 Ca2+ but no Significant difference between Ht31-expressing and control cells in the rate of Ca2+ cycling or amplitude of the Ca2+ 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 beta -adrenergic stimulation, cardiomyocyte function and substrate phosphorylation by PKA is regulated by targeting of PKA. by AKAPs.
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