Abstract P2074: Site-specific Phosphorylation Of Vinculin Regulates N-cadherin Junction Assembly

Abstract

Introduction: During physiological and pathological growth, cardiomyocytes adapt and remodel in response to changes in mechanical load. The cytoskeleton plays a central role in tissue remodeling as it both senses mechanical stress and mediates structural remodeling which affects the functional response within the cardiomyocyte. Abl kinase-dependent vinculin (VCL) phosphorylation at tyrosine (Y) residue 822 (pVCL-Y822) has been implicated in force-induced cytoskeletal remodeling and adhesion strengthening at cadherin junctions. However, the functional significance of VCL phosphorylation in the working myocardium remains unknown. Objective: To determine the role of vinculin-mediated mechanotransduction in cardiac development, homeostasis, and disease. Methods and Results: Using a phospho-specific antibody, we found that pVCL-Y822 correlates with dynamic junction remodeling in embryonic cardiomyocytes which declines in the postnatal heart when N-cadherin junctions assemble at the intercalated disc. Notably, pVCL-Y822 is upregulated in border zone cardiomyocytes following myocardial infarction. To investigate VCL-mediated mechanotransduction in the heart, CRISPR-mediated genome editing was used to mutate the tyrosine at position 822 to a non-phosphorylatable phenylalanine (F) in the mouse Vcl gene. Vcl Y822F mice were born in the expected Mendelian frequency and are viable. Blocking VCL phosphorylation causes a preferential reduction of mutant VCL protein at the myocyte termini together with the N-cadherin-catenin adhesion complex. Whereas, α5/β1 integrin and fibronectin are both increased along the lateral border of the Vcl Y822F postnatal cardiomyocytes. Vcl Y822F mice exhibit reduced cardiac function with age; however, cardiac remodeling is attenuated in the mutant hearts. Conclusion: This novel animal model demonstrates how blocking vinculin Y822 phosphorylation affects the balance between cadherin adhesions and integrin adhesions and thereby highlights the importance of understanding the crosstalk between the two adhesion systems and its implications for disease pathogenesis.