Reversal of myogenic terminal differentiation by SV40 large T antigen results in mitosis and apoptosis

Abstract

Terminally differentiated skeletal muscle myotubes are arrested in the G0 phase of the cell cycle, and this arrest is not reversed by stimulation with serum or growth factors. The myotubes have been shown to be refractory to apoptosis even under low serum conditions. When the SV40 large T antigen is induced in the C2SVTts11 myotubes, which stably harbor the T antigen gene linked to an inducible promoter, the terminally differentiated cells reenter the cell cycle to resume nuclear DNA replication representing S phase. We show here that the large T-expressing myotubes further proceeded to M phase represented by the appearance of mitotic figures with centrosomes, condensed chromosomes, and mitotic spindles. The myotubes eventually cleaved and midbodies were formed at the cleavage sites of the cytoplasm. In some cases actin filaments, reminiscent of the contractile rings, accumulated at the cleavage furrows. Thus, terminally differentiated myotubes remain able to resume at least one round of the cell cycle and consequently are considered to be capable of dedifferentiation. A subset of myotubes expressing large T did not undergo mitosis. Some of them were degenerative and contained deformed giant nuclei and pulverized nuclei. The others suffered apoptotic cell death, which was identified by morphological changes of the nuclei and the labeling with dUTP at the ends of chromatin DNA fragments. The induction of apoptosis was unlikely to be confined to a particular phase of the cell cycle. These results imply that terminally differentiated myotubes also retain a complete set of machinery for apoptosis.