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Signaling from beta-adrenoceptor to L-type calcium channel: identification of a novel cardiac protein kinase A target possessing similarities to AHNAK

Item Type:Article
Title:Signaling from beta-adrenoceptor to L-type calcium channel: identification of a novel cardiac protein kinase A target possessing similarities to AHNAK
Creators Name:Haase, H. and Podzuweit, T. and Lutsch, G. and Hohaus, A. and Kostka, S. and Lindschau, C. and Kott, M. and Kraft, R. and Morano, I.
Abstract:A novel calcium channel-associated protein of approximately 700 kDa has been identified in mammalian cardiomyocytes that undergoes substantial cAMP-dependent protein kinase (PKA) phosphorylation. It was therefore designated as phosphoprotein 700 (pp700). The pp700 interacts specifically with the beta(2) subunit of cardiac L-type calcium channels as revealed by coprecipitation experiments using affinity-purified antibodies against different calcium channel subunits. It is surprising that amino acid sequence analysis of pig pp700 revealed homology to AHNAK-encoded protein, which was originally identified in human cell lines of neural crest origin as 700-kDa phosphoprotein. Cardiac AHNAK expression was assessed on mRNA level by reverse transcriptase-polymerase chain reaction. Sequence-directed antibodies raised against human AHNAK recognized pp700 in immunoblotting and immunoprecipitation experiments, confirming the homology between both proteins. Anti-AHNAK antibodies labeled preferentially the plasma membrane of cardiomyocytes in cryosections of rat cardiac tissue and isolated cardiomyocytes. Sarcolemmal pp700/AHNAK localization was not influenced by stimulation of either the PKA or the protein kinase C pathway. In back-phosphorylation studies with cardiac biopsies, we identified distinct pp700 pools. The membrane-associated fraction of pp700 underwent substantial in vivo phosphorylation on beta-adrenergic receptor stimulation by isoproterenol, whereas the cytoplasmic fraction of pp700 was not accessible to endogenous PKA. It is important that in vivo phosphorylation occurred in that pp700 fraction which coprecipitated with the calcium channel beta subunit. We hypothesize that both phosphorylation of pp700 and its coupling to the beta subunit play a physiological role in cardiac beta-adrenergic signal transduction.
Keywords:cAMP Kinase, Protein Phosphorylation, Cardiomyocytes, Animals, Swine
Source:FASEB Journal
ISSN:0892-6638
Publisher:Federation of American Societies for Experimental Biology (U.S.A.)
Volume:13
Number:15
Page Range:2161-2172
Date:December 1999
PubMed:View item in PubMed

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