Coxsackievirus and adenovirus receptor (CAR) mediates atrioventricular-node function and connexin 45 localization in the murine heart
J. Clin. Invest. Byung-Kwan Lim, et al. 118:2758 doi:10.1172/JCI34777 [Go to this article.]

Figure 8
CAR associates with connexin 45, β-catenin, and ZO-1. (A) Whole-heart protein extracts were immunoprecipitated using IgG as a negative control and anti-CAR (top panels) or connexin 45 antibodies (bottom panels). (B) HeLa cells were transfected with connexin 45–Flag only (Cx45-Flag; lane 1) as control, with connexin 45–Flag and CAR (Cx45-Flag + CAR; lane 2), or with connexin 45–Flag and CAR lacking the PDZ domain–binding motif (Cx45-Flag + CARΔPDZ; lane 3). Connexin 45–Flag pulled down CAR and ZO-1 in the connexin 45–Flag– and CAR-expressing HeLa cells. In contrast, connexin 45–Flag was not able to precipitate CAR or ZO-1 after transfection of HeLa cells with CARΔPDZ. (C) To confirm the importance of the PDZ domain–binding motif of connexin 45, CAR was transfected with connexin 45–Flag (Cx45-Flag + CAR; lane 2) or connexin 45–Flag lacking the PDZ domain–binding motif (Cx45ΔPDZ-Flag + CAR; lane 3) or FHL2-Flag as control (FHL2-Flag + CAR; lane 1). Connexin 45 lacking the PDZ domain–binding motif did not coprecipitate with CAR. (D) Protein extracts from the ventricular myocardium were immunoprecipitated using IgG as a negative control or the anti-CAR antibody. β-catenin and CAR were detected in the precipitate. (E) CAR-Flag was precipitated with anti–β-catenin antibody in CAR-Flag–transfected HeLa cells. (F) β-catenin was precipitated with CAR-Flag from HeLa cells transfected with CAR-Flag but not with the negative control, FHL2-Flag.