Regulation of protein kinase D activity in adult myocardium: Novel counter-regulatory roles for protein kinase CF, and protein kinase A

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Abstract

Protein kinase D (PKD) is activated downstream of protein kinase C (PKC) in many cell types, although little is known about the mechanisms that regulate PKD in adult myocardium. Exposure of cultured adult rat ventricular myocyles (ARVM) to phorbol 12-myristate 13-acetate (PMA; 100 nM for 5 min) activated PKD, as evidenced by significantly increased phosphorylation at Ser744/8 (PKC phosphorylation sites) and Ser916 (autophosphorylation site). PKD activation occurred concomitantly with translocation of the enzyme from the cytosolic to the particulate fraction. The role of PKC was confirmed by pretreatment (15 min) of ARVM with the PKC X inhibitors GF I 109203X (1 mu M) and Ro31-8220 (I PM), both of which prevented PKD phosphorylation on subsequent exposure to PMA. Exposure of ARVM to endothelin-1 (ET1; 100 nM for 10 min) also activated PKD by a PKC-dependent mechanism. To determine the PKC isoform(s) involved in the ET1-induced PKD activation, ARVM were infected with adenoviral vectors encoding dominant-negative (DN) mutants of 13KC alpha, PKC delta and PKC epsilon. Expression of DN-PKC alpha and DN-PKC delta had little effect on ET1-induced PKD activation, whilst this was significantly attenuated by expression of DN-PKC epsilon, indicating that PKC epsilon plays a predominant role in the pertinent ETI signaling pathway. Intriguingly, prior exposure to the adenylyl cyclase activator forskolin (I mu M for 5 min) or the beta-adrenergic agonist isoprenaline (100 nM for 5 min) markedly attenuated ET1-induced PKD activation, but not PMA-induced PKD activation. The ET1-induced response was rescued when protein kinase A (PKA) was inhibited (H89, 10 mu M) before exposure to isoprenaline. These results show that ET1-induced PKD activation in ARVM is mediated by PKC, primarily the PKC epsilon isoform, and is suppressed by PKA activation. (c) 2007 Elsevier Inc. All rights reserved
Original languageEnglish
Pages (from-to)686 - 695
Number of pages10
JournalJournal of Molecular and Cellular Cardiology
Volume43
Issue number6
DOIs
Publication statusPublished - Dec 2007

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