Distinct Signaling Pathways Mediate Cardiomyocyte Phospholipase D Stimulation by Endothelin-1 and Thrombin

Abstract

Several G protein-coupled receptors which stimulate phospholipase C (PLC) also activate phospholipase D (PLD) in cardiomyocytes. Here, we characterized PLD activation in neonatal rat cardiomyocytes by the PLC-stimulatory thrombin receptor PAR1, in comparison to the endothelin-1 receptor ETAR, which induces PLD stimulation by activation of protein kinase C (PKC) δ and ϵ. Similar to ETAR, activation of PAR1 induced PLD stimulation, which, however, was insensitive to PKC inhibition. Furthermore, in contrast to ETAR, PLD stimulation by PAR1 was suppressed by overexpression of regulators of G protein signaling specific for G12-type G proteins and treatment with brefeldin A, an inhibitor of guanine nucleotide exchange factors for ADP-ribosylation factor (ARF) GTPases. On the other hand, inactivation of Rho GTPases by Clostridium difficile toxin B and treatment with general tyrosine kinase inhibitors suppressed PAR1- and ETAR- as well as phorbol ester-induced PLD stimulation and was associated with a fall in the cellular level of phosphatidylinositol 4,5-bisphosphate (PIP2). We conclude that, in contrast to ETAR-PLD coupling, PAR1-induced cardiomyocyte PLD stimulation is PKC-independent and mediated by G12-type G proteins and ARF GTPases, while Rho and tyrosine kinases regulate PLD stimulation by either receptor, apparently by controlling the cellular level of PIP2, a common regulator of PLD activity.