Increased Association of ZO-1 With Connexin43 During Remodeling of Cardiac Gap Junctions

Abstract

Abstract-The intercellular geometry of connexin43 (Cx43) gap junctional coupling is key to coordinated spread of electrical activation through the ventricle of the mammalian heart. A progressive redistribution of electrical and mechanical junctions into intercalated discs occurs during postnatal development. Breakdown of disc-localized pattern in the adult heart, to recapitulate immature distributions, is thought to be key to the genesis of conduction disturbance and arrhythmia. Recently, ZO-1 (a PDZ-MAGUK protein), has been suggested to have a role in generating coupling geometries between myocytes. We therefore investigated the codistribution of ZO-1 with Cx43 and N-cadherin in the adult rat ventricle using quantitative immunoconfocal and immunoelectron microscopy. These analyses indicated that, whereas ZO-1 and Cx43 codistribute within discs, only low to moderate point-by-point colocalization of Cx43 and ZO-1 is found within these domains compared with the relatively high level of colocalization between N-cadherin and ZO-1. By contrast, levels of association between Cx43 and ZO-1 increased rapidly and significantly (PϽ0.001) after partial or complete enzymatic dissociation of myocytes from intact ventricle-a treatment known to induce gap junction endocytosis. Coimmunoprecipitation using Cx43-and ZO-1-specific antibodies confirmed that significantly (PϽ0.03) increased ZO-1 is precipitated relative to Cx43 in freshly dissociated myocytes as compared with intact ventricle. On immunoblots, decreases in Cx43 relative mobility, consistent with increased phosphorylation, were observed following myocyte dissociation. The increased ZO-1-Cx43 association that occurs after remodeling of myocyte intercellular contacts indicates the possibility of unanticipated roles for ZO-1 in gap junction turnover during cardiac development and disease processes.

The Second PDZ Domain of Zonula Occludens-1 Is Dispensable for Targeting to Connexin43 Gap Junctions

“…The present findings regarding ZO-1 engagement of Cx43 at plaque edges accord with our previous finding that only a fraction of the total ZO-1 present in intercalated disks of ventricular cardiomyocytes colocalizes with Cx43 plaques (Barker et al 2002). More recently, others have focused on the spatial arrangement of ZO-1 relative to Cx43 GJs within cardiac tissues.…”

“…plaque size; we proposed that, as a consequence of disengagement of ZO-1 from Cx43 at the GJ edge, plaques expanded due to unregulated channel accretion . It has been variously proposed that ZO-1 enhances GJ assembly (Laing et al 2005), plays a role in GJ disassembly (Akoyev and Takemoto 2007), or mediates GJ internalization (Barker et al 2002;Segretain et al 2004). ZO-1 also has been implicated in the regulation of intercellular communication through Cx43 GJs (Akoyev and Takemoto 2007;Girao and Pereira 2007;Laing et al 2005;Toyofuku et al 2001;van Zeijl et al 2007).…”

N-cadherin haploinsufficiency affects cardiac gap junctions and arrhythmic susceptibility

“…The scaffold protein, ZO-1, interacts with the large carboxy terminus of Cx43 via a PDZ domain (28,33), and the cadherin/catenin complex interacts with ZO-1 via α-catenin and the actin cytoskeleton (34) thus forming a molecular linkage between the two protein complexes. ZO-1 is primarily localized with N-cadherin at the ICD, however during gap junction remodeling it is more closely associated with Cx43 (29). In osteoblastic cells, disruption of the Cx43/ZO-1 interaction using a dominant negative ZO-1 leads to a decrease in gap junction function as determined by dye transfer (35).…”

Defective Epidermal Barrier in Neonatal Mice Lacking the C-Terminal Region of Connexin43

“…This is most likely due to mutual influences of changes in cardiomyocyte architecture and electrophysiological properties. Perinatally, Cx43 and calcium-dependent cell contact proteins, including ZO-1, are dispersed around the sarcolemma of cardiomyocytes (Barker et al, 2002) before intercalated discs begin to form. To date, there are two possible explanations for this transition.…”