Internal translation of the connexin 43 transcript

Abstract

BackgroundConnexin 43 (Cx43), the most widely expressed gap junction protein, is associated with a number of physiological and pathological conditions. Many functions of Cx43 have been shown to be independent of gap junction formation and only require the expression of Cx43 C-terminal fragments. Recent evidence demonstrated that naturally occurring C-terminal isoforms can be generated via internal translation.FindingsHere, we confirm that C-terminal domains of Cx43, particularly the major 20-kDa isoform, can be independently generated and regulated by internal translation of the same single GJA1 gene transcript that encodes full-length Cx43. Through direct RNA transfection experiments, we provide evidence that internal translation is not due to a bona fide cap-independent IRES-mediated mechanism, as upstream ribosomal scanning or translation is required. In addition to the mTOR pathway, we show for the first time, using both inhibitors and cells from knockout mice, that the Mnk1/2 pathway regulates the translation of the main 20-kDa isoform.ConclusionsInternal translation of the Cx43 transcript occurs but is not cap-independent and requires translation upstream of the internal start codon. In addition to the PI3K/AKT/mTOR pathway, the major 20-kDa isoform is regulated by the Mnk1/2 pathway. Our results have major implications for past and future studies describing gap junction-independent functions of Cx43 in cancer and other pathological conditions. This study provides further clues to the signalling pathways that regulate internal mRNA translation, an emerging mechanism that allows for increased protein diversity and functional complexity from a single mRNA transcript.

Connexin 43 and CaV1.2 Ion Channel Trafficking in Healthy and Diseased Myocardium

“…This is the first evidence that alternative translation is possible for human ion channels and in human heart. These results have since been supported by a separate report showing that the GJA1–20k isoform is expressed in many cell lines that express high levels of full length Cx43 57 . In addition, it has also been reported that this 20 kDa isoform is induced by hypoxic stimuli in the mouse brain and is the result of internal translation from an IRES element 58 .…”

ASC Induces Apoptosis via Activation of Caspase-9 by Enhancing Gap Junction-Mediated Intercellular Communication

“…Gap junction assembly by connexin 43 was documented to propagate cell-killing signals initially generated by a single cell spontaneously initiating apoptosis to surrounding cells in a bladder carcinoma cell line [40]. Since the expression of connexin 43 is regulated by the PI3K/AKT/mTOR and Mnk1/2 pathways [41], the role of ASC in gap junction signaling that occurs at a high density needs to be clarified.…”

Cardiomyocyte protein trafficking: Relevance to heart disease and opportunities for therapeutic intervention

“…Cx43 is the first mammalian ion channel shown to be subjected to alternative translation, which has already been confirmed by two separate reports [38, 39]. It has also been found that the 20 kDa isoform (GJA1-20K) assists with Cx43 forward trafficking.…”

“…Given the recent findings of alternative translation of smaller Cx43 isoforms, which facilitate the forward trafficking of full length Cx43, and the increases in such alternative translation by inhibiting mTOR and Mnk1/2 [37, 39], use of existing mTOR and Mnk1/2 inhibitors could increase plaque density. As mTOR inhibitors are already widely used in the fields of organ transplant and immunosuppression, therapeutic rescue could potentially be available as a novel use of an existing agent.…”