title: Role of direct RhoA-phospholipase D1 interaction in mediating adenosine-induced protection from cardiac ischemia.
abstract: Activation of adenosine A1 or A3 receptors protects heart cells from ischemia-induced injury. The A3 receptor signals via RhoA and phospholipase D (PLD) to induce cardioprotection. The objective of the study was to investigate how RhoA activates PLD to achieve the anti-ischemic effect of adenosine A3 receptors. In an established cardiac myocyte model of preconditioning using the cultured chick embryo heart cells, overexpression of the RhoA-noninteracting PLD1 mutant I870R selectively blocked the A3 agonist (Cl-IBMECA, 10 nM)-induced cardioprotection. I870R caused a significantly higher percentage of cardiac cells killed in A3 agonist-treated than in A1 agonist (CCPA, 10 nM)-treated myocytes (ANOVA and posttest comparison, P<0.01). Consistent with its inhibitory effect on the PLD activity, I870R attenuated the Cl-IBMECA-mediated PLD activation. Cl-IBMECA caused a 41 +/- 15% increase in PLD activity in mock-transfected myocytes (P<0.01, paired t test) while having only a slight stimulatory effect on the PLD activity in I870R-transfected cells. To further test the anti-ischemic role of a direct RhoA-PLD1 interaction, atrial cardiac myocytes were rendered null for native adenosine receptors by treatment with irreversible A1 antagonist m-DITC-XAC and were selectively transfected with the human adenosine A1 or A3 receptor cDNA individually or they were cotransfected with cDNAs encoding either receptor plus I870R. I870R preferentially inhibited the human A3 receptor-mediated protection from ischemia. The RhoA-noninteracting PLD1 mutant caused a significantly higher percentage of cardiac cells killed in myocytes cotransfected with the human A3 receptor than in those cells expressing the human A1 receptor (ANOVA and posttest comparison, P<0.01). The present data provided the first demonstration of a novel physiological role for the direct RhoA-PLD1 interaction, that of potent protection from cardiac ischemia. The study further supported the concept that a divergent signaling mechanism mediates the anti-ischemic effect of adenosine A1 and A3 receptors.
authors.affiliation: Department of Cardiology, University of Connecticut Health Center, 263 Farmington Avenue, Farmington, Connecticut 06030, USA.
authors.forename: Susan
authors.initials: S
authors.lastname: Mozzicato
authors.forename: Bhalchandra V
authors.initials: BV
authors.lastname: Joshi
authors.forename: Kenneth A
authors.initials: KA
authors.lastname: Jacobson
authors.forename: Bruce T
authors.initials: BT
authors.lastname: Liang
mesh_terms: D000241:Adenosine / Q000627:therapeutic use*; D058907:Adenosine A1 Receptor Agonists; D058909:Adenosine A3 Receptor Agonists; D000818:Animals; D002478:Cells, Cultured; D002642:Chick Embryo; D004789:Enzyme Activation; D015870:Gene Expression; D006801:Humans; D019157:Ischemic Preconditioning, Myocardial; D008954:Models, Biological; D009154:Mutation; D017202:Myocardial Ischemia / Q000188:drug therapy* / Q000201:enzymology / Q000378:metabolism; D032383:Myocytes, Cardiac; D010739:Phospholipase D / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D043682:Receptor, Adenosine A1 / Q000235:genetics / Q000378:metabolism; D043684:Receptor, Adenosine A3 / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
chemical_list: D058907:Adenosine A1 Receptor Agonists; D058909:Adenosine A3 Receptor Agonists; D043682:Receptor, Adenosine A1; D043684:Receptor, Adenosine A3; D010739:Phospholipase D; C106801:phospholipase D1; D020742:rhoA GTP-Binding Protein; D000241:Adenosine