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© The Rockefeller University Press, 0021-9525/1997//155 $5.00
The Journal of Cell Biology, Volume 136, Number 1, , 1997 155-165

Article

Newt Myotubes Reenter the Cell Cycle by Phosphorylation of the Retinoblastoma Protein



Elly M. Tanaka*, Alexander A.F. Gann*,§, Phillip B. Gates*, and Jeremy P. Brockes*,{ddagger}

* Ludwig Institute for Cancer Research and {ddagger} Department of Biochemistry and Molecular Biology, University College London, London W1P 8BT, United Kingdom; and § Division of Biological Sciences, Lancaster University, Lancaster LA1 4YQ, United Kingdom

Withdrawal from the cell cycle is an essential aspect of vertebrate muscle differentiation and requires the retinoblastoma (Rb) protein that inhibits expression of genes needed for cell cycle entry. It was shown recently that cultured myotubes derived from the Rb–/–mouse reenter the cell cycle after serum stimulation (Schneider, J.W., W. Gu, L. Zhu, V. Mahdavi, and B. Nadal-Ginard. 1994. Science (Wash. DC). 264:1467– 1471). In contrast with other vertebrates, adult urodele amphibians such as the newt can regenerate their limbs, a process involving cell cycle reentry and local reversal of differentiation. Here we show that myotubes formed in culture from newt limb cells are refractory to several growth factors, but they undergo S phase after serum stimulation and accumulate 4N nuclei. This response to serum is inhibited by contact with mononucleate cells. Despite the phenotypic parallel with Rb–/– mouse myotubes, Rb is expressed in the newt myotubes, and its phosphorylation via cyclin-dependent kinase 4/6 is required for cell cycle reentry. Thus, the postmitotic arrest of urodele myotubes, although intact in certain respects, can be undermined by a pathway that is inactive in other vertebrates. This may be important for the regenerative ability of these animals.

Abbreviations used in this paper: BrdU, bromodeoxyuridine; CDI, cyclin-dependent kinase inhibitor; CDK, cyclin-dependent kinase; CIP, calf intestinal phosphatase; GST, glutathione-S-transferase; Rb, retinoblastoma.

E.M. Tanaka was supported by fellowships from the Muscular Dystrophy Association, Inc. and The Helen Hay Whitney Foundation.

Address all correspondence to Jeremy P. Brockes, Ludwig Institute for Cancer Research, Department of Biochemistry and Molecular Biology, University College London, 91 Riding House Street, London W1P 8BT, United Kingdom. Tel.: (44) 171-878-4044. Fax: (44) 171-878-4040. e-mail: jerbro{at}ludwig.ucl.ac.uk


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