The importance of connexin 43 and its phosphorylation for mitochondrial function and myocardial ischemia/reperfusion injury

• Applicant: Privatdozentin Dr. Kerstin Böngler
• Subject Area: Anatomy and Physiology
• Term: from 2018 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 411490269

Connexin 43 (Cx43) is localized at gap junctions of the cardiomyocyte sarcolemma, but is also present at the inner membrane of subsarcolemmal mitochondria. Here, the protein is involved in dye uptake and potassium influx, suggesting that Cx43 also forms a channel at the mitochondria. Furthermore, mitochondrial Cx43 influences oxygen consumption and calcium handling. A reduction in mitochondrial Cx43 is associated with a loss of cardioprotection following ischemic preconditioning (IPC). Gap junctional Cx43 is phosphorylated at multiple sites by different kinases, and also mitochondrial Cx43 is phosphorylated. The exact residues, which are phosphorylated in mitochondrial Cx43 under physiological conditions, after ischemia/reperfusion without and with IPC are unknown and will be identified using phospho-specific antibodies. Cx43 phosphorylation will be studied in wildtype mice as well as in mice, in which specific phosphorylation sites within Cx43 (targeted by protein kinase C (Cx43PKCmut mice), casein kinase 1 (Cx43CK1mut mice), and mitogen-activated protein kinase (Cx43MK4mut mice)), are rendered phosphorylation-insensitive. We aim to identify proteins which interact with mitochondrial Cx43. Mitochondrial function will be analyzed in mitochondria from Cx43PKCmut, Cx43CK1mut und Cx43MK4mut mice, and will be compared to that of wildtype mice. The importance of Cx43 phosphorylation will be studied by subjecting wildtype and Cx43 phosphorylation-insensitive mice to myocardial ischemia/reperfusion without or with ischemic preconditioning and myocardial infarct size will be quantified.Prior data on the importance of Cx43 for mitochondrial function and ischemia/reperfusion injury were obtained in models with genetic reduction or pharmacological inhibition of Cx43. To evaluate the effect of enhanced amounts of Cx43, we generated mice, in which the expression of Cx43 can be induced in cardiomyocytes by the administration of tamoxifen. The amount of gap junctional and mitochondrial Cx43 will be analyzed in these mice and compared to that of wildtype mice. The importance of Cx43 for heart function will be characterized by echocardiography. In addition, mitochondrial function will be studied in Cx43-overexpressing mice. In mitochondria from aged myocardium, the amount of mitochondrial Cx43 is reduced. This Cx43 reduction is associated with a loss of infarct size reduction by ischemic preconditioning. In a rescue experiment, Cx43 overexpression will be induced in aged mice and infarct size will be determined after ischemia/reperfusion without and with ischemic preconditioning.

• DFG Programme: Research Grants