The gap junctional protein connexin 36 (Cx36) has been co-purified with the lipid raft protein caveolin-1 (Cav-1). The relevance of an interaction between the two proteins is unknown. In this study, we explored the significance of Cav-1 interaction in the context of intracellular and membrane transport of Cx36. Coimmunoprecipitation assays and Förster resonance energy transfer analysis (FRET) were used to confirm the interaction between the two proteins in the Neuro 2a cell line. We found that the Cx36 and Cav-1 interaction was dependent on the intracellular calcium levels. By employing different microscopy techniques, we demonstrated that Cav-1 enhances the vesicular transport of Cx36. Pharmacological interventions coupled with cell surface biotinylation assays and FRET analysis revealed that Cav-1 regulates membrane localization of Cx36. Our data indicate that the interaction between Cx36 and Cav-1 plays a role in the internalization of Cx36 by a caveolin-dependent pathway.
Keywords: FRET; TIRF; caveolin-1; connexin 36; lipid raft; protein interaction; transport.