Functional Interactions Between Mechanical Junction Proteins, Connexin43 and the Voltage-Gated Sodium Channel Complex in the Heart.

Abstract

Desmosomes and adherens junctions provide mechanical continuity between cardiac cells, whereas gap junctions allow for cell-cell electrical/metabolic coupling. These structures reside at the cardiac intercalated disc (ID). Also at the ID is the voltage-gated sodium channel (VGSC) complex. Functional interactions between desmosomes, gap junctions, and VGSCs have been demonstrated. Separate studies show, under various conditions, decreased abundance of gap junctions at the ID, and redistribution of connexin43 (Cx43) to plaques oriented parallel to fiber direction (gap junction “lateralization”). The mechanisms of Cx43 lateralization, and the fate of desmosomal and VGSC molecules in the setting of Cx43 remodeling, remain understudied. To study remodeling we employed the sheep pulmonary hypertension model. We found that Cx43 lateralization is a part of a complex remodeling that includes mechanical and gap junctions, but may exclude components of VGSC. Cx43/desmosomal remodeling was accompanied by lateralization of two microtubule-associated proteins relevant for Cx43 trafficking: EB1 and the kinesin protein Kif5b. Thus, we speculate that lateralization results from redirectionality of microtubule-mediated forward trafficking. Remodeling of junctional complexes may preserve electrical synchrony under conditions that disrupt ID integrity. We then further focused on the importance of the expression of Cx43 and desmosomal protein, PKP2, on the function of VGSCs in the heart. Studies of PKP2 and Cx43 deficiency (PKP2+/- and Cx43-/-) mice models demonstrated that these proteins affect the distribution and function of VGSC. In an attempt to reveal the mechanism for Cx43-mediated regulation of VGSCs we analyzed the ability of Cx43 to stabilize tubulin and for the plus-end of the microtubules to reach its anchoring point at sites rich in the adherens junction protein N-cadherin. Using Cx43 null mice and the immortal cell line of cardiac origin (HL-1) we were able to show that, in the heart, connexin43 regulates the function of VGSCs via its tubulin binding domain. Overall, the data presented in this thesis further illustrate the intimate functional interactions of the proteins residing at the ID and reinforces the idea of these proteins working together as parts of macromolecular complexes.