We investigated the effect of pH on isometric tension in actin filament-reconstituted and thin filament-reconstituted bovine cardiac muscle fibers in the pH range of 6.0-7.4. Thin filament was reconstituted from purified G-actin with either bovine cardiac tropomyosin (Tm) or rabbit skeletal Tm in conjunction with cardiac or skeletal troponin (Tn). Results showed that isometric tension decreased linearly with a decrease in pH. The slope of the pH-tension relation, DeltaF/DeltapH (Deltarelative tension/Deltaunit pH), was 0.28 and 0.44 in control cardiac fibers and skeletal fibers, respectively. In actin filament-reconstituted fibers without regulatory proteins, DeltaF/DeltapH was 0.62, namely larger than that in cardiac or skeletal fibers. When reconstituted with cardiac Tm-Tn complex (nTm), DeltaF/DeltapH recovered to 0.32, close to the value obtained in control cardiac fibers. When reconstituted with skeletal nTm, DeltaF/DeltapH recovered to 0.48, close to the value for control skeletal fibers. To determine whether Tm or Tn is responsible for the inhibitory effects of nTm on the tension decrease caused by reduced pH, thin filament was reconstituted with cardiac Tm and skeletal Tn, or with skeletal Tm and cardiac Tn. When cardiac Tm was used, pH dependence of isometric tension coincided with that of control cardiac fibers. When skeletal Tm was used, the pH dependence coincided with that of control skeletal fibers. Furthermore, closely similar results were obtained in fibers reconstituted with actin and either cardiac or skeletal Tm without Tn. These results demonstrate that Tm but not Tn modulates the pH dependence of active tension.