Mutant p53 can delay growth arrest and loss of CDK2 activity in senescing human fibroblasts without reducing p21(WAF1) expression.
Functional wild-type p53 is required for human diploid fibroblasts (HDF) to enter an irreversible growth arrest known as replicative senescence. Experimentally, abrogation of p53 function by expression of human papillomavirus type 16 E6 or disruption of a key downstream effector p21 by homologous recombination both extended HDF life span. However, although sufficient to extend life span, p21 down-regulation is not necessary, because expression of a dominant-negative mutant p53 (143(ala)) extends life span without apparently decreasing p21 expression. Given the importance of p53 in cellular senescence and the general assumption that p21 may be the sole mediator of its action in this process, we have investigated how abrogation of p53 function can overcome senescence without lowering expression of p21. We have found up-regulated levels of the cyclin-dependent kinase 2 (cdk2) protein in HDF expressing 143(ala) mutant p53 as compared to senescent controls, together with an increase in p21-free cdk2 which, in conjunction with cyclin E, is able to form an active kinase which can phosphorylate the retinoblastoma protein. However, forced overexpression of cdk2 in near-senescent HDF failed to restore cdk2-associated kinase activity. Our data suggest that p53-mediated senescence depends on factor(s) other than p21 which modulate formation of cyclin E-cdk2 complexes.
Wyllie
Fiona
F
Department of Pathology, University of Wales College of Medicine, Heath Park, Cardiff, CF14 4XN, United Kingdom. wyllie@cf.ac.uk
Haughton
Michele
M
Bartek
Jiri
J
Rowson
Jan
J
Wynford-Thomas
David
D
D042846:CDC2-CDC28 Kinases; D018797:Cell Cycle Proteins / Q000032:analysis; D002455:Cell Division; D002460:Cell Line; D016922:Cellular Senescence; D019938:Cyclin D1 / Q000378:metabolism; D019927:Cyclin E / Q000378:metabolism; D051357:Cyclin-Dependent Kinase 2; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D016213:Cyclins / Q000032:analysis / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D009154:Mutation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000502:physiology*; D025521:Tumor Suppressor Proteins / Q000032:analysis
C493956:CDKN1A protein, human; D018797:Cell Cycle Proteins; D019927:Cyclin E; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D046911:Macromolecular Substances; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D019938:Cyclin D1; D050760:Cyclin-Dependent Kinase Inhibitor p27; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases