Abstract 15042: Arrhythmogenic Cardiomyopathy Mutations Cause Disassembly of the Cx43 Forward Trafficking Machinery Which Can Be Rescued by GSK-3β Inhibition
Arrhythmogenic cardiomyopathy leads to a decrease in connexin43 (Cx43) immunosignal at the intercalated disc, despite unchanged total Cx43 protein. In an experimental model of arrhythmogenic cardiomyopathy, involving transfection of neonatal ventricular cardiomyocytes and HeLa cells with mutated plakoglobin (2057del2 plakoglobin, corresponding to Naxos disease in humans), we quantified the decrease in Cx43 at the intercalated disc by morphometric analysis of confocal slices, and analyzed the EB1-dependent microtubular trafficking machinery known to be responsible for forward trafficking of Cx43 cargo to the intercalated disc. Expression of mutated plakoglobin reduced Cx43 signal at the intercalated disc from 100±13% to 47±2%, n=9 (mean±S.E). In HeLa cells, a cell line known to express the full EB1-dependent trafficking machinery, mutated plakoglobin produced a marked disassembly of microtubules, characterized by a failure of alignment of microtubules towards the cell border, less developed EB1 protein comets at microtubule plus-ends, and a lack of a discernable microtubule organizing center (MTOC). Glycogen synthase kinase-3β (GSK-3β) is a known constitutive inhibitor of microtubule dynamics and EB1 activity. SB216763, a specific inhibitor of GSK-3β fully rescued the MTOC, microtubular alignment and EB1 activity at microtubule plus-ends.
These results suggest that decreased connexin43 at the cardiac cell-to-cell border in arrhythmogenic cardiomyopathy patients is attributable to disassembly of the microtubule based forward trafficking machinery. Furthermore, inhibition of GSK-3β, rescues both EB1 activity and microtubule dynamics to properly localize Cx43 at cell-to-cell borders.
Author Disclosures: S. Zhang: None. D.A. Kuhn: None. E. Kessler: None. A. Asimaki: None. V. Sottas: None. D. Vanhecke: None. B. Rothen-Rutishauser: None. J.E. Saffitz: None. R.M. Shaw: None. A.G. Kleber: Consultant/Advisory Board; Modest; Schiller Inc. Baar Switzerland.
This research has received full or partial funding support from the American Heart Association.
- © 2014 by American Heart Association, Inc.