Abstract
The human p53 protein isoforms are expressed in several cell lines and modulate p53 tumor suppressor activity, mainly through modulation of gene expression (1–4). Thus, identifying the pattern of p53 isoforms expression in cell lines is a key step for future studies of the p53 network (5). At the moment, the detection of p53 protein isoforms is based on the use of a panel of antibodies allowing their identification by comparing their molecular weights and their detection pattern by different antibodies (6). Here, classical protocols supplemented with technical know-how are described to detect p53 protein isoforms at protein level by Western blotting and immunoprecipitation. Furthermore, a simple method to study the impact of p53 protein isoforms on p53 transcriptional activity through luciferase reporter gene assays is provided.
Virginie Marcel and Marie P. Khoury have contributed equally to this work.
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References
Bourdon J-C, Fernandes K, Murray-Zmijewski F, Liu G, Diot A, Xirodimas DP, Saville MK, Lane DP (2005) p53 isoforms can regulate p53 transcriptional activity. Genes Dev 19:2122–2137
Aoubala M, Murray-Zmijewski F, Khoury MP, Fernandes K, Perrier S, Bernard H, Prats AC, Lane DP, Bourdon J-C (2011) p53 directly transactivates Δ133p53α, regulating cell fate outcome in response to DNA damage. Cell Death Differ 18:248–258
Fujita K, Mondal AM, Horikawa I, Nguyen GH, Kumamoto K, Sohn JJ, Bowman ED, Mathe EA, Schetter AJ, Pine SR, Ji H, Vojtesek B, Bourdon J-C, Lane DP, Harris CC (2009) p53 isoforms Delta133p53 and p53beta are endogenous regulators of replicative cellular senescence. Nat Cell Biol 11:1135–1142
Marcel V, Vijayakumar V, Fernandez-Cuesta L, Hafsi H, Sagne C, Hautefeuille A, Olivier M, Hainaut P (2010) p53 regulates the transcription of its Delta133p53 isoform through specific response elements contained within the TP53 P2 internal promoter. Oncogene 29:2691–2700
Marcel V, Dichtel-Danjoy ML, Sagne C, Hafsi H, Ma D, Ortiz-Cuaran S, Olivier M, Hall J, Mollereau B, Hainaut P, Bourdon J-C (2011) Biological functions of p53 isoforms through evolution: lessons from animal and cellular models. Cell Death Differ 18(12):1815–24
Khoury MP, Bourdon J-C (2010) The isoforms of the p53 protein. Cold Spring Harb Perspect Biol 2:a000927
Courtois S, Verhaegh G, North S, Luciani MG, Lassus P, Hibner U, Oren M, Hainaut P (2002) DeltaN-p53, a natural isoform of p53 lacking the first transactivation domain, counteracts growth suppression by wild-type p53. Oncogene 21:6722–6728
Marcel V, Perrier S, Aoubala M, Ageorges S, Groves MJ, Diot A, Fernandes K, Tauro S, Bourdon J-C (2010) Δ160p53 is a novel N-terminal p53 isoform encoded by Δ133p53 transcript. FEBS Lett 584:4463–4468
Yin Y, Stephen CW, Luciani MG, Fåhraeus R (2002) p53 stability and activity is regulated by Mdm2-mediated induction of alternative p53 translation products. Nat Cell Biol 4:462–467
Murray-Zmijewski F, Lane DP, Bourdon J-C (2006) p53/p63/p73 isoforms: an orchestra of isoforms to harmonise cell differentiation and response to stress. Cell Death Differ 13:962–972
Ollman M, Young JM, Di Como CJ, Karim F, Belvin M, Robertson S, Whittaker K, Demsky M, Fisher WW, Buchman A, Duyk G, Friedman L, Prives C, Kopczynski C (2000) Drosophila p53 is a structural and functional homolog of the tumor suppressor p53. Cell 101:91–101
Brodsky MH, Nordstrom W, Tsang G, Kwan E, Rubin GM, Abrams JM (2000) Drosophila p53 binds a damage response element at the reaper locus. Cell 101:103–113
Lee K-C, Goh WLP, Xu M, Kua N, Lunny D, Wong JS, Coomber D, Vojtesek B, Lane EB, Lane DP (2008) Detection of the p53 response in zebrafish embryos using new monoclonal antibodies. Oncogene 27:629–640
Acknowledgments
This work was supported by Cancer Research UK (C8/A6613). V.M. is supported by Breast Cancer Campaign, M.P.K., K.F., A.D., and J.C.B. are supported by Cancer Research UK.
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Marcel, V., Khoury, M.P., Fernandes, K., Diot, A., Lane, D.P., Bourdon, JC. (2013). Detecting p53 Isoforms at Protein Level. In: Deb, S., Deb, S. (eds) p53 Protocols. Methods in Molecular Biology, vol 962. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-236-0_2
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DOI: https://doi.org/10.1007/978-1-62703-236-0_2
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