Skip to main content
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1989 May;9(5):2163–2172. doi: 10.1128/mcb.9.5.2163

Characterization of the human p53 gene promoter.

S P Tuck 1, L Crawford 1
PMCID: PMC363010  PMID: 2664471

Abstract

Transcriptional deregulation of the p53 gene may play an important part in the genesis of some tumors. We report here an accurate determination of the transcriptional start sites of the human p53 gene and show that the majority of p53 mRNA molecules do not contain a postulated stem-loop structure at their 5' ends. Recombinant plasmids of the human p53 promoter-leader region fused to the bacterial chloramphenicol acetyltransferase gene (cat) were constructed. After transfection into rodent or human cells, a 350-base-pair fragment spanning the promoter region conferred 4% of the CAT activity mediated by the simian virus 40 early promoter/enhancer. We monitored the efficiency with which 15 3' and 5' promoter deletion constructs initiated transcription. Our results show that an 85-base-pair fragment, previously thought to have resided in exon 1, is all that is required for full promoter activity.

Full text

PDF
2170

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aviv H., Leder P. Purification of biologically active globin messenger RNA by chromatography on oligothymidylic acid-cellulose. Proc Natl Acad Sci U S A. 1972 Jun;69(6):1408–1412. doi: 10.1073/pnas.69.6.1408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Banks L., Matlashewski G., Crawford L. Isolation of human-p53-specific monoclonal antibodies and their use in the studies of human p53 expression. Eur J Biochem. 1986 Sep 15;159(3):529–534. doi: 10.1111/j.1432-1033.1986.tb09919.x. [DOI] [PubMed] [Google Scholar]
  3. Bendori R., Resnitzky D., Kimchi A. Changes in p53 mRNA expression during terminal differentiation of murine erythroleukemia cells. Virology. 1987 Dec;161(2):607–611. doi: 10.1016/0042-6822(87)90159-0. [DOI] [PubMed] [Google Scholar]
  4. Berk A. J., Sharp P. A. Sizing and mapping of early adenovirus mRNAs by gel electrophoresis of S1 endonuclease-digested hybrids. Cell. 1977 Nov;12(3):721–732. doi: 10.1016/0092-8674(77)90272-0. [DOI] [PubMed] [Google Scholar]
  5. Bienz-Tadmor B., Zakut-Houri R., Libresco S., Givol D., Oren M. The 5' region of the p53 gene: evolutionary conservation and evidence for a negative regulatory element. EMBO J. 1985 Dec 1;4(12):3209–3213. doi: 10.1002/j.1460-2075.1985.tb04067.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  7. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  8. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  9. Chow V., Ben-David Y., Bernstein A., Benchimol S., Mowat M. Multistage Friend erythroleukemia: independent origin of tumor clones with normal or rearranged p53 cellular oncogenes. J Virol. 1987 Sep;61(9):2777–2781. doi: 10.1128/jvi.61.9.2777-2781.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Crawford L. V., Pim D. C., Bulbrook R. D. Detection of antibodies against the cellular protein p53 in sera from patients with breast cancer. Int J Cancer. 1982 Oct 15;30(4):403–408. doi: 10.1002/ijc.2910300404. [DOI] [PubMed] [Google Scholar]
  11. Crawford L. V., Pim D. C., Lamb P. The cellular protein p53 in human tumours. Mol Biol Med. 1984 Aug;2(4):261–272. [PubMed] [Google Scholar]
  12. DeLeo A. B., Jay G., Appella E., Dubois G. C., Law L. W., Old L. J. Detection of a transformation-related antigen in chemically induced sarcomas and other transformed cells of the mouse. Proc Natl Acad Sci U S A. 1979 May;76(5):2420–2424. doi: 10.1073/pnas.76.5.2420. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dippold W. G., Jay G., DeLeo A. B., Khoury G., Old L. J. p53 transformation-related protein: detection by monoclonal antibody in mouse and human cells. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1695–1699. doi: 10.1073/pnas.78.3.1695. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dony C., Kessel M., Gruss P. Post-transcriptional control of myc and p53 expression during differentiation of the embryonal carcinoma cell line F9. Nature. 1985 Oct 17;317(6038):636–639. doi: 10.1038/317636a0. [DOI] [PubMed] [Google Scholar]
  15. Filmus J., Benchimol S., Buick R. N. Comparative analysis of the involvement of p53, c-myc and c-fos in epidermal growth factor-mediated signal transduction. Exp Cell Res. 1987 Apr;169(2):554–559. doi: 10.1016/0014-4827(87)90215-1. [DOI] [PubMed] [Google Scholar]
  16. Gorman C. M., Moffat L. F., Howard B. H. Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Mol Cell Biol. 1982 Sep;2(9):1044–1051. doi: 10.1128/mcb.2.9.1044. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Graham F. L., van der Eb A. J. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology. 1973 Apr;52(2):456–467. doi: 10.1016/0042-6822(73)90341-3. [DOI] [PubMed] [Google Scholar]
  18. Heard J. M., Herbomel P., Ott M. O., Mottura-Rollier A., Weiss M., Yaniv M. Determinants of rat albumin promoter tissue specificity analyzed by an improved transient expression system. Mol Cell Biol. 1987 Jul;7(7):2425–2434. doi: 10.1128/mcb.7.7.2425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Herbomel P., Bourachot B., Yaniv M. Two distinct enhancers with different cell specificities coexist in the regulatory region of polyoma. Cell. 1984 Dec;39(3 Pt 2):653–662. doi: 10.1016/0092-8674(84)90472-0. [DOI] [PubMed] [Google Scholar]
  20. Jay G., DeLeo A. B., Appella E., Dubois G. C., Law L. W., Khoury G., Old L. J. A common transformation-related protein in murine sarcomas and leukemias. Cold Spring Harb Symp Quant Biol. 1980;44(Pt 1):659–664. doi: 10.1101/sqb.1980.044.01.069. [DOI] [PubMed] [Google Scholar]
  21. Khochbin S., Principaud E., Chabanas A., Lawrence J. J. Early events in murine erythroleukemia cells induced to differentiate. Accumulation and gene expression of the transformation-associated cellular protein p53. J Mol Biol. 1988 Mar 5;200(1):55–64. doi: 10.1016/0022-2836(88)90333-6. [DOI] [PubMed] [Google Scholar]
  22. Koeffler H. P., Miller C., Nicolson M. A., Ranyard J., Bosselman R. A. Increased expression of p53 protein in human leukemia cells. Proc Natl Acad Sci U S A. 1986 Jun;83(11):4035–4039. doi: 10.1073/pnas.83.11.4035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kozak M. Leader length and secondary structure modulate mRNA function under conditions of stress. Mol Cell Biol. 1988 Jul;8(7):2737–2744. doi: 10.1128/mcb.8.7.2737. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Krieg P. A., Melton D. A. Functional messenger RNAs are produced by SP6 in vitro transcription of cloned cDNAs. Nucleic Acids Res. 1984 Sep 25;12(18):7057–7070. doi: 10.1093/nar/12.18.7057. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lamb P., Crawford L. Characterization of the human p53 gene. Mol Cell Biol. 1986 May;6(5):1379–1385. doi: 10.1128/mcb.6.5.1379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Linzer D. I., Levine A. J. Characterization of a 54K dalton cellular SV40 tumor antigen present in SV40-transformed cells and uninfected embryonal carcinoma cells. Cell. 1979 May;17(1):43–52. doi: 10.1016/0092-8674(79)90293-9. [DOI] [PubMed] [Google Scholar]
  27. Masuda H., Miller C., Koeffler H. P., Battifora H., Cline M. J. Rearrangement of the p53 gene in human osteogenic sarcomas. Proc Natl Acad Sci U S A. 1987 Nov;84(21):7716–7719. doi: 10.1073/pnas.84.21.7716. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Matlashewski G. J., Tuck S., Pim D., Lamb P., Schneider J., Crawford L. V. Primary structure polymorphism at amino acid residue 72 of human p53. Mol Cell Biol. 1987 Feb;7(2):961–963. doi: 10.1128/mcb.7.2.961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Matlashewski G., Banks L., Pim D., Crawford L. Analysis of human p53 proteins and mRNA levels in normal and transformed cells. Eur J Biochem. 1986 Feb 3;154(3):665–672. doi: 10.1111/j.1432-1033.1986.tb09449.x. [DOI] [PubMed] [Google Scholar]
  30. Matlashewski G., Lamb P., Pim D., Peacock J., Crawford L., Benchimol S. Isolation and characterization of a human p53 cDNA clone: expression of the human p53 gene. EMBO J. 1984 Dec 20;3(13):3257–3262. doi: 10.1002/j.1460-2075.1984.tb02287.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Maxam A. M., Gilbert W. A new method for sequencing DNA. Proc Natl Acad Sci U S A. 1977 Feb;74(2):560–564. doi: 10.1073/pnas.74.2.560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. McGeady M. L., Wood T. G., Maizel J. V., Vande Woude G. F. Sequences upstream from the mouse c-mos oncogene may function as a transcription termination signal. DNA. 1986 Aug;5(4):289–298. doi: 10.1089/dna.1986.5.289. [DOI] [PubMed] [Google Scholar]
  33. Milner J., McCormick F. Lymphocyte stimulation: concanavalin A induces the expression of a 53K protein. Cell Biol Int Rep. 1980 Jul;4(7):663–667. doi: 10.1016/0309-1651(80)90205-2. [DOI] [PubMed] [Google Scholar]
  34. Mowat M., Cheng A., Kimura N., Bernstein A., Benchimol S. Rearrangements of the cellular p53 gene in erythroleukaemic cells transformed by Friend virus. Nature. 1985 Apr 18;314(6012):633–636. doi: 10.1038/314633a0. [DOI] [PubMed] [Google Scholar]
  35. Oren M., Maltzman W., Levine A. J. Post-translational regulation of the 54K cellular tumor antigen in normal and transformed cells. Mol Cell Biol. 1981 Feb;1(2):101–110. doi: 10.1128/mcb.1.2.101. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Oren M., Reich N. C., Levine A. J. Regulation of the cellular p53 tumor antigen in teratocarcinoma cells and their differentiated progeny. Mol Cell Biol. 1982 Apr;2(4):443–449. doi: 10.1128/mcb.2.4.443. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Prost E., Moore D. D. CAT vectors for analysis of eukaryotic promoters and enhancers. Gene. 1986;45(1):107–111. doi: 10.1016/0378-1119(86)90138-1. [DOI] [PubMed] [Google Scholar]
  38. Reich N. C., Levine A. J. Growth regulation of a cellular tumour antigen, p53, in nontransformed cells. Nature. 1984 Mar 8;308(5955):199–201. doi: 10.1038/308199a0. [DOI] [PubMed] [Google Scholar]
  39. Reich N. C., Oren M., Levine A. J. Two distinct mechanisms regulate the levels of a cellular tumor antigen, p53. Mol Cell Biol. 1983 Dec;3(12):2143–2150. doi: 10.1128/mcb.3.12.2143. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Reisman D., Greenberg M., Rotter V. Human p53 oncogene contains one promoter upstream of exon 1 and a second, stronger promoter within intron 1. Proc Natl Acad Sci U S A. 1988 Jul;85(14):5146–5150. doi: 10.1073/pnas.85.14.5146. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Rogel A., Popliker M., Webb C. G., Oren M. p53 cellular tumor antigen: analysis of mRNA levels in normal adult tissues, embryos, and tumors. Mol Cell Biol. 1985 Oct;5(10):2851–2855. doi: 10.1128/mcb.5.10.2851. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Rotter V., Witte O. N., Coffman R., Baltimore D. Abelson murine leukemia virus-induced tumors elicit antibodies against a host cell protein, P50. J Virol. 1980 Nov;36(2):547–555. doi: 10.1128/jvi.36.2.547-555.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Rotter V. p53, a transformation-related cellular-encoded protein, can be used as a biochemical marker for the detection of primary mouse tumor cells. Proc Natl Acad Sci U S A. 1983 May;80(9):2613–2617. doi: 10.1073/pnas.80.9.2613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Ruscetti S. K., Scolnick E. M. Expression of a transformation-related protein (p53) in the malignant stage of Friend virus-induced diseases. J Virol. 1983 Jun;46(3):1022–1026. doi: 10.1128/jvi.46.3.1022-1026.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Schirm S., Jiricny J., Schaffner W. The SV40 enhancer can be dissected into multiple segments, each with a different cell type specificity. Genes Dev. 1987 Mar;1(1):65–74. doi: 10.1101/gad.1.1.65. [DOI] [PubMed] [Google Scholar]
  46. Sleigh M. J. A nonchromatographic assay for expression of the chloramphenicol acetyltransferase gene in eucaryotic cells. Anal Biochem. 1986 Jul;156(1):251–256. doi: 10.1016/0003-2697(86)90180-6. [DOI] [PubMed] [Google Scholar]
  47. Smith L. J., McCulloch E. A., Benchimol S. Expression of the p53 oncogene in acute myeloblastic leukemia. J Exp Med. 1986 Sep 1;164(3):751–761. doi: 10.1084/jem.164.3.751. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Todaro G. J., Green H., Swift M. R. Susceptibility of human diploid fibroblast strains to transformation by SV40 virus. Science. 1966 Sep 9;153(3741):1252–1254. doi: 10.1126/science.153.3741.1252. [DOI] [PubMed] [Google Scholar]
  49. Wigler M., Pellicer A., Silverstein S., Axel R. Biochemical transfer of single-copy eucaryotic genes using total cellular DNA as donor. Cell. 1978 Jul;14(3):725–731. doi: 10.1016/0092-8674(78)90254-4. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES