Fast and slow isoforms of troponin I and troponin C. Distribution in normal rabbit muscles and effects of chronic stimulation

Polyclonal antibodies were raised against troponin I (TnI) and troponin C (TnC) purified from fast-twitch and slow-twitch rabbit muscles. These antibodies were used to elucidate the distribution of fast and slow isoforms of TnI and TnC in normal and chronically stimulated rabbit hind limb muscles by immunoblots of one-dimensional and two-dimensional electrophoreses. In contrast to the multiplicity of fast and slow troponin T (TnT) isoforms, TnI and TnC were present as unique fast and slow isoforms. Whereas no charge variants were detected for slow TnI, fast TnI was present in at least three charge variants. As judged from the results of alkaline phosphatase digestion, these charge variants represent differently phosphorylated forms. Fast and slow TnC both exist as two charge variants which, however, were unaffected by alkaline phosphatase treatment. Chronic low-frequency stimulation of fast-twitch muscles induced progressive increases in the slow isoforms of TnC and TnI at the expense of their fast isoforms. The extent of the fast-to-slow transition was more pronounced in the case of TnC than in that of TnI. Long-term stimulated muscles with a complete fast-to-slow transition, at the level of the TnT isoforms, still contained fast and slow isoforms of both TnI and TnC. The coexistence of fast and slow isoforms of the three troponin subunits in the transforming muscle was interpreted as indicating the presence of hybrid troponin molecules composed of fast and slow isoforms. Studies at the mRNA level showed changes similar to those at the protein level. However, in long-term stimulated muscles, the fast-to-slow transition of TnI was more pronounced at the mRNA level than at the protein level.