Thrombin regulates S-phase re-entry by cultured newt myotubes.

BACKGROUND

Adult urodele amphibians such as the newt have remarkable regenerative ability, and a critical aspect of this is the ability of differentiated cells to re-enter the cell cycle and lose their differentiated characteristics. Unlike mammalian myotubes, cultured newt myotubes are able to enter and traverse S phase, following serum stimulation, by a pathway leading to phosphorylation of the retinoblastoma protein. The extracellular regulation of this pathway is unknown.

RESULTS

Like their mammalian counterparts, newt myotubes were refractory to mitogenic growth factors such as the platelet-derived growth factor (PDGF), which act on their mononucleate precursor cells. Cultured newt myotubes were activated to enter S phase by purified thrombin in the presence of subthreshold amounts of serum. The activation proceeded by an indirect mechanism in which thrombin cleaved components in serum to generate a ligand that acted directly on the myotubes. The ligand was identified as a second activity present in preparations of crude thrombin and that was active after removal of all thrombin activity. It induced newt myotubes to enter S phase in serum-free medium, and it acted on myotubes but not on the mononucleate precursor cells. Cultured mouse myotubes were refractory to this indirect mechanism of S-phase re-entry.

CONCLUSIONS

These results provide a link between reversal of differentiation and the acute events of wound healing. The urodele myotube responds to a ligand generated downstream of thrombin activation and re-enters the cell cycle. Although this ligand can be generated in mammalian sera, the mammalian myotube is unresponsive. These results provide a model at the cellular level for the difference in regenerative ability between urodeles and mammals.