We studied subunit composition and Ca(++)-activated ATPase activity of myosin isolated from atria and ventricles of hearts explanted from patients suffering from idiopathic dilated cardiomyopathy. At variance with previously published data, we have been unable to detect in the ventricular subendocardial layers a significant amount of myosin atrial-like light chain 1 (ALC1), which has been reported to be related to some hemodynamic features of the hypertrophied and failing heart. Such a subunit was not visible in the septum and in the subepicardial layers either. On the contrary, in both atria a ventricular-like light chain 2 (VLC2) was found. The nature of this additional light chain was confirmed on the basis of two-dimensional electrophoresis and immunoblotting techniques with polyclonal antibodies reacting with VLC2. In these patients we also observed a depressed Ca(++)-activated ATPase activity, both in atrial and ventricular myosin. The explanation for this finding in ventricles still remains obscure since neither myosin light chains, nor myosin heavy chains showed any difference between patients with dilated cardiomyopathy and controls. On the contrary, in atria we clearly identified changes consistent with the expression of myosin heavy chains of ventricular type and VLC2, which can account for the depressed Ca(++)-activated ATPase activity.