Mono-Heteromeric Configurations of Gap Junction Channels Formed by Connexin43 and Connexin45 Reduce Unitary Conductance and Determine both Voltage Gating and Metabolic Flux Asymmetry.

Guoqiang Zhong,Osman Ahmed,Nazem Akoum,Alonso P Moreno,Eric C Beyer,Daniel A Appadurai,Agustin D Martinez,Volodya Hayrapetyan

doi:10.3389/fphys.2017.00346

Abstract

In cardiac tissues, the expression of multiple connexins (Cx40, Cx43, Cx45, and Cx30.2) is a requirement for proper development and function. Gap junctions formed by these connexins have distinct permeability and gating mechanisms. Since a single cell can express more than one connexin isoform, the formation of hetero-multimeric gap junction channels provides a tissue with an enormous repertoire of combinations to modulate intercellular communication. To study further the perm-selectivity and gating properties of channels containing Cx43 and Cx45, we studied two monoheteromeric combinations in which a HeLa cell co-transfected with Cx43 and Cx45 was paired with a cell expressing only one of these connexins. Macroscopic measurements of total conductance between cell pairs indicated a drastic reduction in total conductance for mono-heteromeric channels. In terms of Vj dependent gating, Cx43 homomeric connexons facing heteromeric connexons only responded weakly to voltage negativity. Cx45 homomeric connexons exhibited no change in Vj gating when facing heteromeric connexons. The distributions of unitary conductances (γj) for both mono-heteromeric channels were smaller than predicted, and both showed low permeability to the fluorescent dyes Lucifer yellow and Rhodamine123. For both mono-heteromeric channels, we observed flux asymmetry regardless of dye charge: flux was higher in the direction of the heteromeric connexon for MhetCx45 and in the direction of the homomeric Cx43 connexon for MhetCx43. Thus, our data suggest that co-expression of Cx45 and Cx43 induces the formation of heteromeric connexons with greatly reduced permeability and unitary conductance. Furthermore, it increases the asymmetry for voltage gating for opposing connexons, and it favors asymmetric flux of molecules across the junction that depends primarily on the size (not the charge) of the crossing molecules.

Highlights

Connexins constitute a homologous family of proteins that oligomerize to form gap junction channels that allow communication between cells
We have shown that the co-expression of Cx45 and Cx43 induces the formation of mono-heteromeric channels with properties different than the ones recorded from their parental homomeric counterparts, including reduced unitary conductance, reduced permeability to large dyes and reduced transjunctional voltage dependence
Our data strongly suggest that this combination of connexins could be an effective physiological method to regulate the levels of communication between cells and in tissues with multiple cell types expressing more than one connexin type (e.g., Neurons, oligodendrocytes and astrocytes in brain, or fibroblasts, myocytes and SA node cells in heart), their membrane distribution and permeability can establish unique communication pathways among these cells, controlling the flow of information between them

Summary

Introduction

Connexins constitute a homologous family of proteins that oligomerize to form gap junction channels that allow communication between cells. Six connexins oligomerize to form gap junction connexons (hemi-channels). The heart, expresses multiple connexins with different functions including connexin (Cx43), connexin (Cx40), connexin (Cx45), and Cx 30.2 (Gros and Jongsma, 1996). These four proteins have been detected in the working and conductive regions of the heart (Bukauskas et al, 2006); Cx45 is more abundant in the sinoatrial node (site of impulse generation) and the atrioventricular node (Coppen et al, 1999b,a). Abnormal abundances and distributions of these connexins may contribute to cardiac pathologies such as arrhythmias (Saffitz et al, 1999; Thibodeau et al, 2010; Askar et al, 2012)

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