Abstract 17525: Mutations in the Transcriptional Cofactor Eya4 Cause Age-Dependent Onset of Adverse Cardiac Physiology and Heart Failure
Introduction: The human mutation E193 in eyes absent homolog 4 (EYA4) was identified to cause hearing impairment followed by heart failure. We prove that Eya4, unlike the mutation E193, together with Six1 negatively regulates p27kip1, a cyclin-dependent kinase inhibitor shown to inhibit hypertrophic growth in adult cardiomyocytes. Here we define an important role for Eya4 in maintaining normal cardiac function.
Methods and results: We first examined the correlation of Eya4 and E193 upon p27 in vitro and demonstrate that an overexpression of Eya4 led to a significant decrease in p27 protein levels; E193-overexpression had a dominant negative effect, releasing Eya4 mediated transcriptional suppression of p27; knockdown of Eya4 with specific shRNA exerted opposing effects. We next generated transgenic (TG) mice with cardiac-specific overexpression of Eya4 and E193. EMSA technique was used to determine protein-DNA interactions in regards to Six1 consensus sites within the p27 promoter. It proved nuclear protein extracts of Eya4 TG mice had a stronger affinity for binding and thereby suppressing the p27 promoter than that of E193 TG animals, which is in line with our in vitro data. Magnetic resonance imaging along with hemodynamic measurements showed Eya4 TG mice developed hypertrophy under baseline conditions while E193 TG mice developed onset of DCM similar to patients carrying the E193 mutation. HE-stainings proved ventricular hypertrophy or dilation of the LV associated with a thinning of the myocardial wall, respectively. PSR-stainings showed interstitial fibrosis of myocardial tissue in both TG mouse models, characteristic for cardiac disease. We also observed that the Casein kinase 2-α-activity and levels of phosphorylated HDAC2 were significantly elevated in Eya4 TG mice, whereas they were significantly reduced in E193 animals compared to wildtype littermates.
In addition, we identified a new human mutation, E215, with phenotypes similar to those seen in E193 patients.
Conclusion: In summary, we identified a mutation in Eya4 to cause DCM. Our results indicate Eya4/Six1 regulates cardiac hypertrophic reactions via p27/CK2-α/HDAC2. Mutations within Eya4 disturb this signalling cascade, finally leading to age-related onset of cardiomyopathy.
- © 2013 by American Heart Association, Inc.