IMMUNOCHEMICAL ANALYSIS OF MYOSIN HEAVY-CHAIN DURING AVIAN MYOGENESIS INVIVO AND INVITRO

Monoclonal antibodies (McAb) against the myosin H chain (MHC) of adult chicken pectoralis muscle were tested for reactivity with pectoralis myosin at selected stages of chick development in vivo and in vitro. Three such McAb, MF 20 and MF 14, which bind to light meromyosin, and MF 30, which binds to myosin subfragment 2 (S2), were used to assay the appearance and accumulation of specific MHC epitopes with: indirect, solid phase radioimmune assay (RIA), immunoautoradiography and immunofluorescence microscopy. McAb MF 20 bound strongly and equivalently to MHC at all stages of embryonic development in vivo. In contrast, the MF 30 epitope was barely detectable at 12 d of incubation but its concentration rose rapidly just before hatching. No detectable binding of MF 14 to pectoralis myosin could be measured during myogenesis in vivo until 1 wk after hatching. Immunofluorescence studies revealed that all 3 epitopes accumulate in the same myocytes of the developing pectoralis muscle. Since all 3 McAb bound with high activity to native and denatured forms of myosin, it is unlikely that differential antibody reactivity can be explained by conformational changes in myosin during development in vivo. When myogenesis in vitro was monitored using the same McAb, MF 20 bound to the MHC at all stages tested while reactivity of MF 30 and MF 14 with myosin from cultured muscle was never observed. Three different immunochemical states of the MHC during development in vivo of chick pectoralis muscle and the absence of later occurring immunochemical transitions in the MHC of cultured embryonic muscle were indicated.