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Common tumour p53 mutations in immortalized cells from Hupki mice heterozygous at codon 72

Abstract

Codon 72 of human p53 gene is polymorphic, encoding arginine or proline. Here we report construction of a human p53 knock-in (Hupki) mouse encoding the codon 72pro variant. The new strain was crossed with the original Hupki mice (codon 72arg/arg) to obtain primary embryonic fibroblasts polymorphic at codon 72 or homozygous for codon 72pro. The fibroblasts, cultured under standard conditions, immortalized within 12 weeks and acquired p53 mutations similarly to Hupki codon 72arg/arg cells investigated previously. Sequencing of human p53 exons 4–9 in immortalized cultures revealed missense mutations found repeatedly in human tumours. In cell lines ensuing from benzo(a)pyrene-treated cultures the combined p53 mutation pattern from experiments with the 3 codon 72 genotypes showed a predominance of strand-biased G to T transversions (18 of 36 mutations), and mutations recurring at smokers' lung tumour hotspot codons 157 and 273, supporting involvement of tobacco carcinogens in shaping the mutation signature in lung cancers of smokers. Mutations in cell lines from unexposed cultures did not cluster at these codons and G to T transversions were uncommon (2 of 52 mutations) (Fisher's exact test P<0.0001). Most mutations (13/16) in cell lines derived from cells polymorphic at codon 72 were found on the proline allele, with loss of the arginine allele.

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Figure 1
Figure 2

Abbreviations

BaP:

benzo(a)pyrene

HUF:

Hupki embryonic fibroblasts

Hupki:

human p53 knock-in

References

  • Bergamaschi D, Samuels Y, Sullivan A, Zvelebil M, Breyssens H, Bisso A et al. (2006). iASPP preferentially binds p53 proline-rich region and modulates apoptotic function of codon 72-polymorphic p53. Nat Genet 38: 1133–1141.

    Article  CAS  PubMed  Google Scholar 

  • Chen JX, Zheng Y, West M, Tang M-s . (1998). Carcinogens preferentially bind at methylated CpG in the p53 mutational hot spots. Cancer Res 58: 2070–2075.

    CAS  PubMed  Google Scholar 

  • Dearth LR, Qian H, Wang T, Baroni TE, Zeng J, Chen SW et al. (2007). Inactive full-length p53 mutants lacking dominant wild-type p53 inhibition highlight loss of heterozygosity as an important aspect of p53 status in human cancers. Carcinogenesis 28: 289–298.

    Article  CAS  PubMed  Google Scholar 

  • Denissenko MF, Chen JX, Tang MS, Pfeifer GP . (1997). Cytosine methylation determines hot spots of DNA damage in the human p53 gene. Proc Natl Acad Sci USA 94: 3893–3898.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Denissenko MF, Pao A, Pfeifer GP, Tang M . (1998). Slow repair of bulky DNA adducts along the nontranscribed strand of the human p53 gene may explain the strand bias of transversion mutations in cancers. Oncogene 16: 1241–1247.

    Article  CAS  PubMed  Google Scholar 

  • Denissenko MF, Pao A, Tang M, Pfeifer GP . (1996). Preferential formation of benzo[a]pyrene adducts at lung cancer mutational hotspots in P53. Science 274: 430–432.

    Article  CAS  PubMed  Google Scholar 

  • Dumont P, Leu JI, Della PA, George DL, Murphy M . (2003). The codon 72 polymorphic variants of p53 have markedly different apoptotic potential. Nat Genet 33: 357–365.

    Article  CAS  PubMed  Google Scholar 

  • Feng Z, Hu W, Hu Y, Tang MS . (2006). Acrolein is a major cigarette-related lung cancer agent: preferential binding at p53 mutational hotspots and inhibition of DNA repair. Proc Natl Acad Sci USA 103: 15404–15409.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hahn WC, Weinberg RA . (2002). Modelling the molecular circuitry of cancer. Nat Rev Cancer 2: 331–341.

    Article  CAS  PubMed  Google Scholar 

  • Hainaut P, Hollstein M . (2000). p53 and human cancer: the first ten thousand mutations. Adv Cancer Res 77: 81–137.

    Article  CAS  PubMed  Google Scholar 

  • Hainaut P, Pfeifer GP . (2001). Patterns of p53 G → T transversions in lung cancers reflect the primary mutagenic signature of DNA-damage by tobacco smoke. Carcinogenesis 22: 367–374.

    Article  CAS  PubMed  Google Scholar 

  • Harvey DM, Levine AJ . (1991). P53 alteration is a common event in the spontaneous immortalization of primary BALB/c murine embryo fibroblasts. Genes Dev 5: 2375–2385.

    Article  CAS  PubMed  Google Scholar 

  • Hu Y, McDermott MP, Ahrendt SA . (2005). The p53 codon 72 proline allele is associated with p53 gene mutations in non-small cell lung cancer. Clin Cancer Res 11: 2502–2509.

    Article  CAS  PubMed  Google Scholar 

  • Liu Z, Belharazem D, Muehlbauer KR, Nedelko T, Knyazev Y, Hollstein M . (2007). Mutagenesis of human p53 tumor suppressor gene sequences in embryonic fibroblasts of genetically-engineered mice. Genet Eng (NY) 28: 45–54.

    Article  Google Scholar 

  • Liu Z, Hergenhahn M, Schmeiser HH, Wogan GN, Hong A, Hollstein M . (2004). Human tumor p53 mutations are selected for in mouse embryonic fibroblasts harboring a humanized p53 gene. Proc Natl Acad Sci USA 101: 2963–2968.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Liu Z, Muehlbauer KR, Schmeiser HH, Hergenhahn M, Belharazem D, Hollstein MC . (2005). p53 mutations in benzo(a)pyrene-exposed human p53 knock-in murine fibroblasts correlate with p53 mutations in human lung tumors. Cancer Res 65: 2583–2587.

    Article  CAS  PubMed  Google Scholar 

  • Luo JL, Yang Q, Tong WM, Hergenhahn M, Wang ZQ, Hollstein M . (2001). Knock-in mice with a chimeric human/murine p53 gene develop normally and show wild-type p53 responses to DNA damaging agents: a new biomedical research tool. Oncogene 20: 320–328.

    Article  CAS  PubMed  Google Scholar 

  • Marin MC, Jost CA, Brooks LA, Irwin MS, O'Nions J, Tidy JA et al. (2000). A common polymorphism acts as an intragenic modifier of mutant p53 behaviour. Nat Genet 25: 47–54.

    Article  CAS  PubMed  Google Scholar 

  • Nelson HH, Wilkojmen M, Marsit CJ, Kelsey KT . (2005). TP53 mutation, allelism and survival in non-small cell lung cancer. Carcinogenesis 26: 1770–1773.

    Article  CAS  PubMed  Google Scholar 

  • Olivier M, Eeles R, Hollstein M, Khan MA, Harris CC, Hainaut P . (2002). The IARC TP53 database: new online mutation analysis and recommendations to users. Hum Mutat 19: 607–614.

    Article  CAS  PubMed  Google Scholar 

  • Parrinello S, Samper E, Krtolica A, Goldstein J, Melov S, Campisi J . (2003). Oxygen sensitivity severely limits the replicative lifespan of murine fibroblasts. Nat Cell Biol 5: 741–747.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pfeifer GP, Hainaut P . (2003). On the origin of G to t transversions in lung cancer. Mutat Res 526: 39–43.

    Article  CAS  PubMed  Google Scholar 

  • Pietsch EC, Humbey O, Murphy ME . (2006). Polymorphisms in the p53 pathway. Oncogene 25: 1602–1611.

    Article  CAS  PubMed  Google Scholar 

  • Song H, Hollstein M, Xu Y . (2007). P53 gain-of-function cancer mutants induce genetic instability by inactivating ataxia-telangiectasia mutated. Nat Cell Biol 9: 573–580.

    Article  CAS  PubMed  Google Scholar 

  • Storey A, Thomas M, Kalita A, Harwood C, Gardiol D, Mantovani F et al. (1998). Role of a p53 polymorphism in the development of human papillomavirus-associated cancer. Nature 393: 229–234.

    Article  CAS  PubMed  Google Scholar 

  • Szymanowska A, Jassem E, Dziadziuszko R, Borg A, Limon J, Kobierska-Gulida G et al. (2006). Increased risk of non-small cell lung cancer and frequency of somatic TP53 gene mutations in Pro72 carriers of TP53 Arg72Pro polymorphism. Lung Cancer 52: 9–14.

    Article  PubMed  Google Scholar 

  • Thomas M, Kalita A, Labrecque S, Pim D, Banks L, Matlashewski G . (1999). Two polymorphic variants of wild-type p53 differ biochemically and biologically. Mol Cell Biol 19: 1092–1100.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • vomBrocke J, Schmeiser HH, Reinbold M, Hollstein M . (2006). MEF immortalization to investigate the ins and outs of mutagenesis. Carcinogenesis 27: 2141–2147.

    Article  CAS  Google Scholar 

  • Zienolddiny S, Ryberg D, Arab MO, Skaug V, Haugen A . (2001). Loss of heterozygosity is related to p53 mutations and smoking in lung cancer. Br J Cancer 84: 226–231.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

This work was funded by the Deutsche Krebshilfe, the Yorkshire Cancer Research UK charity, and the National Cancer Institute USA.

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Correspondence to M Hollstein.

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Reinbold, M., Luo, JL., Nedelko, T. et al. Common tumour p53 mutations in immortalized cells from Hupki mice heterozygous at codon 72. Oncogene 27, 2788–2794 (2008). https://doi.org/10.1038/sj.onc.1210932

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