Troponin in embryonic chick skeletal muscle cell in vitro: an immunoelectron microscopic study
In developing skeletal muscle, myosin and actin are synthesized and polymerized into filamentous forms, thick and thin filaments, respectively. These myofilaments of two varieties have been shown, with the use of "decoration with heavy meromyosin" technique, to be arranged at the right polarity and spatial position as those seen in mature myofibrils from the initial phases of myofibrillogenesis. The question arises as to whether regulatory proteins are distributed along embryonic thin filaments from such early stages of development. In order to clarify this problem, the fine structural localization of troponin was investigated by the use of immunoelectron microscopy.Troponin and its components (troponin C [TN-C], I [TN-I] and T [TN-T]) were prepared from adult chicken breast muscles. Rabbit antisera against each of these troponin components were prepared. The serum was fractioned by ethanol or ammonium sulphate fractionation. Myogenic cells from 12-day chick embryonic thigh muscles were grown in monolayer and, after 2-8 days in vitro, the cultures were treated as follows: (i) They were immersed in 50% glycerol. The suspension of separated thin filaments was obtained by gentle homogenizing the cells.