Cx43 has distinct mobility within plasma-membrane domains, indicative of progressive formation of gap-junction plaques

Connexin 43 (Cx43) is a dynamic molecule, having a short halflife of only a few hours. In this study, we use fluorescent-proteintagged Cx43 variants to examine Cx43 delivery to the cell surface, its residency status in various cell-surface membrane domains and its mobility characteristics. Rapid time-lapse imaging led to the identification of Cx43 being delivered to cell-surface domains that lacked a contacting cell, and also to its localization within membrane protrusions. Fluorescence recovery after photobleaching (FRAP) was used to investigate the mobility state of cell-surface-localized Cx43. Cx43 mobility within clustered cell-surface profiles of Cx43 could be categorized into those with generally a high degree of lateral mobility and those with generally a low degree of lateral mobility. Cx43 mobility was independent of cluster size, yet the C-terminal domain of Cx43 regulated the proportion of gap-junction-like clusters that acquired a low Cx43 mobility state. Collectively, these studies show that Cx43 establishes residency at all cell-surface membrane domains, and progressively acquires assembly states that probably reflect differences in either channel packing and/or its interactions with Cx43-binding proteins.