Summary
Interaction of p53 with mismatched DNA induces proteolytic cleavage with release of a 35-kDa protein fragment from the p53–DNA complexes. The 35-kDa cleavage product is activated for specific biochemical function(s) and may play a role in the cellular response to DNA damage (Molinari et al (1996) Oncogene: 2077–2086; Okorokov et al (1997) EMBO J 16: 6008–6017). In the present study we have asked if mutants of p53 retain the ability to undergo similar proteolytic cleavage, and compared sequence-specific ‘DNA contact’ with ‘structural’ mutants commonly found in human cancer. In addition, a series of phosphorylation site mutants were generated to investigate the possible effects of phosphorylation/dephosphorylation on the proteolytic cleavage of p53. All mutants tested bound to a mismatched DNA target in vitro. Moreover, studies in vitro and in vivo indicate that p53 mutants with intact conformational structure (as determined by immunoreactivity with PAb246 and PAb1620) retain the ability to undergo proteolytic cleavage similar, if not identical, to the wild-type p53 protein. Our results suggest that the capacity for p53 to bind mismatched DNA is independent of structural conformation of the central core domain. Proteolytic cleavage, however, is crucially dependent upon a wild-type conformation of the protein.
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References
Caelles, C, Helmberg, A & Karin, M (1994). p53-dependent apoptosis in the absence of transcriptional activation of p53-target genes. Nature 370: 220–223.
Cho, Y, Gorina, S, Jeffrey, PD & Pavletich, NP (1994). Crystal structure of a p53 tumor suppressor–DNA complex: understanding tumorigenic mutations. Science 265: 346–355.
El-Diery, WS, Tokino, T, Velculescu, VE, Levy, DB, Parsons, R, Trent, JM, Lin, D, Mercer, WE, Kinzler, KW & Vogelstein, B (1993). WAF1, a potential mediator of p53 tumour suppression. Cell 75: 817–825.
Funk, WD, Pak, DJ, Karas, RH, Wright, WE & Shay, JW (1992). A transcriptionally active DNA-binding site for human p53 protein complexes. Mol Cell Biol 12: 2866–2871.
Gannon, JV, Greaves, R, Iggo, R & Lane, DP (1990). Activating mutations in p53 produce a common conformational effect. A monoclonal antibody specific for the mutant form. EMBO J 9: 1595–1602.
Gorina, S & Pavletich, NP (1996). Structure of the p53 tumor suppressor bound to the ankyrin and SH3 domains of 53BP2. Science 274: 1001–1005.
Gottlieb, TM & Oren, M (1996). p53 in growth control and neoplasia. Biochim Biophys Acta 1287: 77–102.
Haupt, Y & Oren, M (1996). p53-mediated apoptosis: mechanisms and regulation. Behring Inst Mitt 97: 32–59.
Haupt, Y, Rowan, S, Shaulian, E, Vousden, KH & Oren, M (1995). Induction of apoptosis in HeLa cells by transactivation-deficient p53. Genes Dev 9: 2170–2183.
Jayaraman, J & Prives, C (1995). Activation of p53 sequence-specific DNA binding by short single strands of DNA requires the p53 C-terminus. Cell 81: 1021–1029.
Ko, LJ & Prives, C (1996). p53: puzzle and paradigm. Genes Dev 10: 1054–1072.
Lee, S, Elenbaas, B, Levine, A & Griffith, J (1995). p53 and its 14 kDa C-terminal domain recognize primary DNA damage in the form of insertion/deletion mismatches. Cell 81: 1013–1020.
Levine, AJ (1997). p53, the cellular gatekeeper for growth and division. Cell 88: 323–331.
Meek, DW (1994). Post-translational modification of p53. Semin Cancer Biol 5: 203–210.
Michalovitz, D, Halevy, O & Oren, M (1990). Conditional inhibition of transformation and of cell proliferation by a temperature-sensitive mutant of p53. Cell 62: 671–680.
Milne, DM, McKendrick, L, Jardine, LJ, Deacon, E, Lord, JM & Meek, DW (1996). Murine p53 is phosphorylated within the PAb421 epitope by protein kinase C in vitro, but not in vivo, even after stimulation with the phorbol ester o-tetradecanoylphorbol 13-acetate. Oncogene 13: 205–211.
Milner, J, Metcalf, EA & Cook, A (1991). Tumour suppressor p53: analysis of wild-type and mutant p53 complexes. Mol Cell Biol 11: 12–19.
Miyashita, T & Reed, JC (1995). Tumor suppressor p53 is a direct transcriptional activator of the human bax gene. Cell 80: 293–299.
Molinari, M, Okorokov, AL & Milner, J (1996). Interaction with damaged DNA induces selective proteolytic cleavage of p53 to yield 40-kDa and 35-kDa fragments competent for sequence-specific DNA binding. Oncogene 13: 2077–2086.
Mummenbrauer, T, Janus, F, Muller, B, Wiesmuller, L, Deppert, W & Grosse, F (1996). p53 Protein exhibits 3′-to-5′ exonuclease activity. Cell 85: 1089–1099.
Okorokov, AL, Ponchel, F & Milner, J (1997). Induced N- and C- terminal cleavage of p53: a core fragment of p53, generated by interaction with damaged DNA, promotes cleavage of the N-terminus of full-length p53, whereas ssDNA induces C-terminal cleavage of p53. EMBO J 16: 6008–6017.
Paetzel, M & Dalbey, RE (1997). Catalytic hydroxyl/amine dyads within serine proteases. Trends Biochem Sci 22: 28–31.
Steegenga, WT, van der Eb, AJ & Jochemsen, AG (1996). How phosphorylation regulates the activity of p53. J Mol Biol 263: 103–113.
Takenaka, I, Morin, F, Seizinger, BR & Kley, N (1995). Regulation of the sequence-specific DNA binding function of p53 by protein kinase C and protein phosphatases. J Biol Chem 270: 5405–5411.
Thukral, SK, Blain, GC, Chang, KK & Fields, S (1994). Distinct residues of human p53 implicated in binding to DNA, simian virus 40 large T antigen, 53BP1, and 53BP2. Mol Cell Biol 14: 8315–8321.
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Mee, T., Okorokov, A., Metcalfe, S. et al. Proteolytic cleavage of p53 mutants in response to mismatched DNA. Br J Cancer 81, 212–218 (1999). https://doi.org/10.1038/sj.bjc.6690679
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DOI: https://doi.org/10.1038/sj.bjc.6690679
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