Histone deacetylase (HDAC) 6 is known to deacetylate amino acid lysine in alpha-tubulin, a component of microtubules, at amino acid position 40. However, the functional role of HDAC6 in the progression of cardiac disease conditions such as pressure-overloaded hypertrophy remain uncertain. The involvement of HDAC6 in cardiac disease was examined using transgenic (TG) mice expressing human dominant-negative HDAC6 (HDAC6H216A, H611A, 4-fold increase) specifically in the heart. Overexpression of HDAC6H216A, H611A significantly increased acetylated alpha-tubulin in mouse hearts, suggesting that HDAC6 can regulate cardiac tubulin deacetylation. Neither histological alteration in the myocardium nor alteration of the cardiac function determined by echocardiography was observed in the HDAC6H216A, H611A TG mice 1 year of age or older. To analyze the role of HDAC6 and acetylated tubulin in disease conditions, we examined the role of HDAC6 in pressure-overloaded hypertrophy generated by transverse aortic constriction (TAC) surgery. A reduction in the shortening fraction was detected in the NTG mouse hearts 4 weeks after TAC surgery while a sustained shortening fraction was observed in the TAC HDAC6H216A, H611A TG mouse hearts, suggesting that inactivation of HADC6 with concomitant enhancement in acetylated tubulin can prevent cardiac disease in pressure-overloaded hypertrophy.