Abstract P096: Eya4, a Transcription Cofactor Crucial in Acquired Heart Disease

Abstract

Introduction: We previously identified a mutation in the human transcriptional cofactor Eya4 (E193) as cause of familial dilated cardiomyopathy (DCM) and heart failure. Upon interaction with real transcription factors such as Six family members, Eya4 is recruited to and interacts with target genes. One of the few known Eya-Six targets expressed in the heart is the cyclin-dependent kinase inhibitor p27kip1, which has been shown to inhibit hypertrophic growth in adult cardiomyocytes, is. We therefore hypothesize that Eya4/Six1 regulates targets relevant in normal cardiac function. Methods and results: We examined p27 expression in response to Eya4 in permanent mammalian cell lines. Western blot analysis demonstrated that an overexpression of Eya4 led to a significant downregulation of p27, whereas the overexpression of E193 had no effect on p27 levels. Studies using a p27 promoter fragment including Six1 consensus sites cloned in front of a Luciferase reporter gene proved Eya4 acts as a suppressor of p27 already at the transcriptional level, whereas E193 could not sufficiently inhibit p27 expression. Further transfection and knockdown experiments revealed that under basal conditions an Eya4 overexpression decreased protein synthesis in primary cardiac myocytes. We constructed transgenic mouse models with a constitutive myocardial overexpression of E193 and Eya4 to study the precise role of Eya4 in the heart. First analysis of these animals using magnetic resonance imaging to visualize cardiac structures in detail show that an overexpression of E193 leads to an age related onset of DCM as seen in an increase in LVEDV, whereas Eya4 overexpressing mice show no signs of heart disease. Conclusion: In summary, we identified a mutation in Eya4 to cause DCM. We now provide evidence that the Eya4/Six1 signalling cascade is also relevant in more common forms of acquired heart disease. Eya4/Six1 seems to regulate the expression of p27kip1, an important inhibitor of the development of hypertrophy in postmitotic cardiomyocytes. Studies using E193 overexpressing mice support our hypothesis whereas tempering the Eya4/Six1 signalling cascade disturbs cardiac physiology.