A 3.5-kDa polypeptide associated with the inner membrane of rat liver was found to be phosphorylated by [gamma-P-32]ATP, presumably via a cAMP-dependent kinase. The phosphorylation was modulated by [Ca2+] in the physiological range, with a minimum at 1 mu M and rising fourfold toward lower (10 nM) and higher (10 mu M) concentrations. Further characterization of the 3.5-kDa component showed that the polypeptide has the same electrophoretic mobility as subunit c of F0F1-ATPase and that it selectively binds to antibodies against subunit c.