Abstract 19069: Autoregulation of Connexin43 Gap Junction Formation by Internally Translated Isoforms

Abstract

During each heartbeat, cell-cell electrical coupling via connexin 43 (Cx43) gap junctions allows billions of individual cardiomyocytes to synchronously contract. The Cx43 protein turns over rapidly, rendering regulation of Cx43 trafficking a critical and continuous cellular need. Altered Cx43 trafficking during heart disease disrupts intercellular coupling and can contribute to the arrhythmias of sudden cardiac death. Better understanding of how Cx43 channels are transported to the cell surface will lead to therapies that can preserve normal electrical coupling of diseased heart muscle. We have identified a novel regulatory mechanism whereby internal translation of the coding sequence of Cx43 ( GJA1 ) mRNA generates N-terminally truncated isoforms that regulate trafficking of full length Cx43 channels. Biochemical analysis of human heart and cell lines reveals at least four additional Cx43 isoforms, with a 20 kDa isoform predominating. We find that in frame AUG codons within GJA1 mRNA represent isoform translation initiation sites and that their ablation arrests trafficking of full-length Cx43. The predominant 20 kDa isoform is sufficient to rescue this trafficking defect when expressed in trans , implicating an essential role as a trafficking chaperone for Cx43. Consistent with these data, expression of Cx43 truncated isoforms is enhanced through inhibition of the PI3K/AKT/mTOR pathway in cardiomyocytes, and concomitantly increases Cx43 gap junction plaque size. Such cap-independent internal translation initiation events may represent a common mechanism for auto-regulation of membrane protein trafficking which is likely altered during stress and, based on these studies, represents a potent target for therapeutic preservation of intercellular coupling in heart disease.