Abstract 546: GJA1-20k Protects the Heart From Ischemic Injury by Inducing Mitochondrial Biogenesis and Metabolic Quiescence

Abstract

Connexin 43 (Cx43) is a cell-cell communication gap junction protein that is encoded by the GJA1 gene and is expressed in all organ systems. It was recently found that the GJA1 mRNA undergoes alternative translation to generate N-terminally truncated isoforms, of which GJA1-20k is the most abundant. We have recently identified that GJA1-20k has a strong tropism for mitochondria and that GJA1-20k overexpression is sufficient to rescue mitochondrial localization to the cell periphery, limiting organelle network collapse upon exposure to oxidative stress. Given the protective role of GJA1-20k on mitochondrial network and the importance of preserving mitochondrial integrity in preventing cardiac ischemic injury, we therefore asked the question whether exogenous GJA1-20k can increase myocardial survival when subjected to stress. Using high resolution confocal imaging, we find that H 2 O 2 -induced oxidative stress in cardiomyocytes causes an upregulation of GJA1-20k independent of full length Cx43 expression. Furthermore, we find that GJA1-20k is upregulated and preferentially enriched in mitochondria after acute ischemia/reperfusion injury in isolated mouse hearts. We also find that exogenous GJA1-20k, when delivered by adenovirus to isolated adult mouse cardiomyocytes, was sufficient to improve cell survival and viability during H 2 O 2 -induced oxidative stress. When delivered in vivo using an Adeno-associated virus serotype 9 (AAV9), GJA1-20k exhibited potent cardioprotective effects by reducing ischemic injury size in a mouse model of myocardial infarction. AAV9-mediated gene transfer of GJA1-20k also resulted in increased cardiac mitochondrial content as observed by electron microscopy. Yet 20k-increased mitochondrial content in cardiomyocytes is associated with reduced mitochondrial membrane potential as well as respiration as measured by Seahorse assay, indicating a quiescent metabolic state resistant to stress. In conclusion, we have identified GJA1-20k as a stress response protein that increases mitochondrial biogenesis and reduces metabolic reactivity, providing powerful cardioprotection against ischemic insults.