Abstract 19570: Delineating a Novel Mechanism of Gap Junction Ubiquitination by Wwp1 in Human Cardiac Disease
Rationale: Gap junctions (GJ) mediate electrical coupling between cardiomyoctes and display mislocalization, decreased function, and/or downregulation in heart failure, in primary hypertrophic cardiomyopathy, and at infarction borders. This process, known as GJ remodeling, contributes to abnormal conduction and the formation of an arrhythmogenic substrate. In the ventricular myocardium, the main constituent of gap junctions is connexin (Cx) 43. Cx43 protein expression and function is post-translationally modified by phosphorylation and ubiquitination. We have recently shown that the ubiquitin ligase WWP1 is upregulated in patients with primary hypertrophic cardiomyopathy (n=2) as well as in hearts derived from patients that died from arrhythmogenic episodes associated with myocardial infarction or hypertension (n=2).
Hypothesis: We hypothesize that pathogenic GJ remodeling is mediated in part by the ubiquitination of Cx43 by Wwp1, resulting in its degradation and the production of an arrhythmogenic substrate.
Methods: Ubiquitination assays were performed in 293T cells. Cardiac structure and function was assessed in animals overexpressing Wwp1 (n=10) relative to wild type littermates (n=10) using routine histology, quantitative real time PCR, M-mode echocardiography, and electrocardiography. Cx43 protein levels and localization within myocardial tissue was determined via Western blotting and multi-immunofluorescence staining in conjunction with laser scanning confocal microscopy.
Results: Here we demonstrate that Wwp1 ubiquitinates Cx43 and this reaction is enhanced when cells are stimulated. The functional consequence of this interaction was confirmed in a novel mouse model. Mice overexpressing the ubiquitin ligase Wwp1 have concentric left ventricular hypertrophy, myofiber disarray, and a 90% decrease in Cx43 protein by 8 weeks of age when they die from ventricular arrhythmias.
Conclusions: Post-translation modification of Cx43 via both phosphorylation and ubiquitination is essential to GJ remodeling and the generation of an arrhythmogenic substrate. This mechanistic insight opens new avenues for potential therapeutic intervention in lethal ventricular arrhythmias associated with GJ remodeling.
- © 2010 by American Heart Association, Inc.