Roles of Cx43-associated protein kinases in suppression of gap junction-mediated chemical coupling by ischemic preconditioning

Abstract

Ischemic preconditioning (PC) suppresses chemical coupling of cardiomyocytes via gap junctions (GJs) during ischemia, which is an adjunct mechanism of protection. The aim of this study was to characterize roles of protein kinases in PC-induced GJ modulation. In isolated rat hearts, ventricular tissues were sampled before and after ischemia with or without PC, and intercalated disc-rich fractions were separated for immunoprecipitation and immunoblotting. Levels of protein kinase C (PKC)-ε, p38mitogen-activated protein kinase (MAPK)-α, and Src coimmunoprecipitated with connexin-43 (Cx43) were increased after ischemia, whereas p38MAPKβ was not detected in the Cx43 immunoprecipitates. PC did not modify the level of Cx43-Src complex after ischemia. However, PC enhanced Cx43-PKCε complex formation, which was abolished by PKCε translocation inhibitory peptide (TIP). In contrast, PC reduced Cx43-p38MAPKα complex level and p38MAPK activity in the Cx43 immunoprecipitates after ischemia. The effect of PC on Cx43-p38MAPKα interaction was mimicked by SB-203580, a p38MAPK inhibitor. PC reduced permeability of GJs to Lucifer yellow in the myocardium at 25 min after ischemia, and this effect was abolished by PKCε-TIP. SB-203580 increased the GJ permeability at 15 min after ischemia compared with that in untreated controls, but the difference became insignificant 25 min after ischemia. In conclusion, PC has distinct effects on interaction of GJ Cx43 with PKCε, p38MAPKα, and Src during ischemia. Suppression of GJ permeability during ischemia by PC is primarily achieved by enhanced interaction of Cx43 with PKCε, which overwhelms the counterbalancing effect of reduced Cx43-p38MAPKα interaction.

Suppression of PKCε-mediated mitochondrial connexin 43 phosphorylation at serine 368 is involved in myocardial mitochondrial dysfunction in a rat model of dilated cardiomyopathy

“…PKCε is an important cardioprotective mediator, which interacts with Cx43 and is required for the phosphorylation of Cx43 at the S368 PKC target site (8). In the present study, the expression and activity of PKCε were examined using western blot analysis to determine whether PKCε mediated the phosphorylation of Cx43 at S368 in the myocardium and mitochondria.…”

“…In addition, the phosphorylation status of Cx43 also affects its suppressive effect on cell proliferation (17). PKCε is essential for the phosphorylation of Cx43 at S368 and is important in ischemic preconditioning-induced cardioprotection by suppressing chemical coupling via Cx43 gap junction modulation (8). Similar to Cx43, PKCε is also located in the mitochondria, which suggests a possible interaction between Cx43 and PKCε in the mitochondria (18).…”

Exercise Preconditioning-Induced Early and Late Phase of Cardioprotection Is Associated With Protein Kinase C Epsilon Translocation

“…25, 26 Enhanced binding of PKCε to Cx43, a major gap junction (GJ) protein, might be the cause of PKCε translocating to intercalated disks. 27 Phosphorylation of Cx43 by PKCε plays a crucial role in δ-opioid-induced suppression of GJ permeability in ischemic myocardium. 28 During the late phase of EP, PKCε did not translocate to intercalated disks, but myocardial injury was still significantly attenuated, as it was during the early phase of EP.…”

Epac stimulation induces rapid increases in connexin43 phosphorylation and function without preconditioning effect

“…Upon activation, PKCε translocates from the cytosol to the particulate fraction (see [21] for review) and phosphorylates a number of targets, among them connexin43 (Cx43), one of the principal connexin isoforms that plays a major role in intercellular communication [37]. A PKC-induced closure of connexons has been proposed [26] as a possible mechanism by which PKCε activation may lead to preconditioning, the phenomenon which induces a significant cardioprotection. It can be achieved by intermittent ischemic episodes or some other interventions prior to prolonged ischemia (see [5] for review).…”