Despite intense investigation over the past century, the molecular mechanisms that regulate maintenance and adaptation of the heart during postnatal development are poorly understood. Myocardin is a remarkably potent transcriptional coactivator expressed exclusively in cardiac myocytes and smooth muscle cells during postnatal development. Here we show that myocardin is required for maintenance of cardiomyocyte structure and sarcomeric organization and that cell-autonomous loss of myocardin in cardiac myocytes triggers programmed cell death. Mice harboring a cardiomyocyte-restricted null mutation in the myocardin gene (Myocd) develop dilated cardiomyopathy and succumb from heart failure within a year. Remarkably, ablation of the Myocd gene in the adult heart leads to the rapid-onset of heart failure, dilated cardiomyopathy, and death within a week. Myocd gene ablation is accompanied by dissolution of sarcomeric organization, disruption of the intercalated disc, and cell-autonomous loss of cardiomyocytes via apoptosis. Expression of myocardin/serum response factor-regulated myofibrillar genes is extinguished, or profoundly attenuated, in myocardin-deficient hearts. Conversely, proapoptotic factors are induced and activated in myocardin-deficient hearts. We conclude that the transcriptional coactivator myocardin is required for maintenance of heart function and ultimately cardiomyocyte survival.