Cardiac hypertrophy is the heart's response to hypertrophic stimuli and is associated with increased mortality. Vinexin-beta is a vinculin-binding protein that belongs to a family of adaptor proteins and mediates signal transduction and actin cytoskeleton organisation. A previous study has shown that Vinexin-beta is ubiquitously expressed and that it is highly expressed in the heart. However, a critical role for Vinexin-beta in cardiac hypertrophy has not been investigated. Therefore, to examine the role of Vinexin-beta in pathological cardiac hypertrophy, we used Vinexin-beta knockout mice and transgenic mice that over-express human Vinexin-beta in the heart. Cardiac hypertrophy was induced by aortic banding (AB). The extent of cardiac hypertrophy was quantitated by echocardiography and pathological and molecular analyses of heart samples. Our results demonstrated that Vinexin-beta overexpression in the heart markedly attenuated cardiac hypertrophy, fibrosis, and cardiac dysfunction, whereas loss of Vinexin-beta exaggerated the pathological cardiac remodelling and fibrosis response to pressure overload. Further analysis of the in vitro and in vivo signalling events indicated that beneficial Vinexin-beta effects were associated with AKT signalling abrogation. Our findings demonstrate for the first time that Vinexin-beta is a novel mediator that protects against cardiac hypertrophy by blocking the AKT signalling pathway.