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ChIP for Identification of p53 Responsive DNA Promoters

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p53 Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 962))

Abstract

Chromatin immunoprecipitation assay (ChIP) has been frequently used to determine whether a transcriptional regulator can bind to a specific DNA element in the chromatin content of cells. Here, we describe a detailed protocol for this assay with hands-on tips based on our own experience in working on the transcriptional regulator and tumor suppressor p53.

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References

  1. Moll UM, Wolff S, Speidel D, Deppert W (2005) Transcription-independent pro-apoptotic functions of p53. Curr Opin Cell Biol 17(6):631–636

    Article  PubMed  CAS  Google Scholar 

  2. Vogelstein B, Lane D, Levine AJ (2000) Surfing the p53 network. Nature 408(6810):307–310

    Article  PubMed  CAS  Google Scholar 

  3. Marchenko ND, Moll UM (2007) The role of ubiquitination in the direct mitochondrial death program of p53. Cell Cycle 6(14):1718–1723

    Article  PubMed  CAS  Google Scholar 

  4. Vousden KH, Lane DP (2007) p53 in health and disease. Nat Rev Mol Cell Biol 8(4):275–283

    Article  PubMed  CAS  Google Scholar 

  5. He L, He X, Lim LP, de Stanchina E, Xuan Z, Liang Y, Xue W, Zender L, Magnus J, Ridzon D, Jackson AL, Linsley PS, Chen C, Lowe SW, Cleary MA, Hannon GJ (2007) A microRNA component of the p53 tumour suppressor network. Nature 447(7148):1130–1134

    Article  PubMed  CAS  Google Scholar 

  6. Orlando V, Paro R (1993) Mapping polycomb-repressed domains in the bithorax complex using in vivo formaldehyde cross-linked chromatin. Cell 75(6):1187–1198

    Article  PubMed  CAS  Google Scholar 

  7. Orlando V (2000) Mapping chromosomal proteins in vivo by formaldehyde-crosslinked-chromatin immunoprecipitation. Trends Biochem Sci 25(3):99–104

    Article  PubMed  CAS  Google Scholar 

  8. Brady CA, Jiang D, Mello SS, Johnson TM, Jarvis LA, Kozak MM, Broz DK, Basak S, Park EJ, McLaughlin ME, Karnezis AN, Attardi LD (2011) Distinct p53 transcriptional programs dictate acute DNA-damage responses and tumor suppression. Cell 145(4):571–583

    Article  PubMed  CAS  Google Scholar 

  9. Zeng SX, Dai MS, Keller DM, Lu H (2002) SSRP1 functions as a co-activator of the transcriptional activator p63. EMBO J 21(20):5487–5497

    Article  PubMed  CAS  Google Scholar 

  10. Zhang Y, Liao JM, Zeng SX, Lu H (2011) p53 downregulates Down syndrome-associated DYRK1A through miR-1246. EMBO Rep 12(8):811–817

    Article  PubMed  CAS  Google Scholar 

  11. Jin Y, Zeng SX, Dai MS, Yang XJ, Lu H (2002) MDM2 inhibits PCAF (p300/CREB-binding protein-associated factor)-mediated p53 acetylation. J Biol Chem 277(34):30838–30843

    Article  PubMed  CAS  Google Scholar 

  12. Bargonetti J, Friedman PN, Kern SE, Vogelstein B, Prives C (1991) Wild-type but not mutant p53 immunopurified proteins bind to sequences adjacent to the SV40 origin of replication. Cell 65(6):1083–1091

    Article  PubMed  CAS  Google Scholar 

  13. Kern SE, Kinzler KW, Bruskin A, Jarosz D, Friedman P, Prives C, Vogelstein B (1991) Identification of p53 as a sequence-specific DNA-binding protein. Science 252(5013):1708–1711

    Article  PubMed  CAS  Google Scholar 

  14. el-Deiry WS, Kern SE, Pietenpol JA, Kinzler KW, Vogelstein B (1992) Definition of a consensus binding site for p53. Nat Genet 1(1):45–49

    Article  PubMed  CAS  Google Scholar 

  15. Wei CL, Wu Q, Vega VB, Chiu KP, Ng P, Zhang T, Shahab A, Yong HC, Fu Y, Weng Z, Liu J, Zhao XD, Chew JL, Lee YL, Kuznetsov VA, Sung WK, Miller LD, Lim B, Liu ET, Yu Q, Ng HH, Ruan Y (2006) A global map of p53 transcription-factor binding sites in the human genome. Cell 124(1):207–219

    Article  PubMed  CAS  Google Scholar 

  16. Cawley S, Bekiranov S, Ng HH, Kapranov P, Sekinger EA, Kampa D, Piccolboni A, Sementchenko V, Cheng J, Williams AJ, Wheeler R, Wong B, Drenkow J, Yamanaka M, Patel S, Brubaker S, Tammana H, Helt G, Struhl K, Gingeras TR (2004) Unbiased mapping of transcription factor binding sites along human chromosomes 21 and 22 points to widespread regulation of noncoding RNAs. Cell 116(4):499–509

    Article  PubMed  CAS  Google Scholar 

  17. Liu Y, Lagowski JP, Vanderbeek GE, Kulesz-Martin MF (2004) Facilitated search for specific genomic targets by p53 C-terminal basic DNA binding domain. Cancer Biol Ther 3(11):1102–1108

    Article  PubMed  CAS  Google Scholar 

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

    Article  PubMed  CAS  Google Scholar 

  19. el-Deiry 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 tumor suppression. Cell 75(4):817–825

    Article  PubMed  CAS  Google Scholar 

  20. Harper JW, Adami GR, Wei N, Keyomarsi K, Elledge SJ (1993) The p21 Cdk-interacting protein Cip1 is a potent inhibitor of G1 cyclin-dependent kinases. Cell 75(4):805–816

    Article  PubMed  CAS  Google Scholar 

  21. Barak Y, Juven T, Haffner R, Oren M (1993) mdm2 expression is induced by wild type p53 activity. EMBO J 12(2):461–468

    PubMed  CAS  Google Scholar 

  22. Zauberman A, Barak Y, Ragimov N, Levy N, Oren M (1993) Sequence-specific DNA binding by p53: identification of target sites and lack of binding to p53 – MDM2 complexes. EMBO J 12(7):2799–2808

    PubMed  CAS  Google Scholar 

  23. Wu X, Bayle JH, Olson D, Levine AJ (1993) The p53-mdm-2 autoregulatory feedback loop. Genes Dev 7(7A):1126–1132

    Article  PubMed  CAS  Google Scholar 

  24. Szak ST, Mays D, Pietenpol JA (2001) Kinetics of p53 binding to promoter sites in vivo. Mol Cell Biol 21(10):3375–3386

    Article  PubMed  CAS  Google Scholar 

  25. Kaeser MD, Iggo RD (2002) Chromatin immunoprecipitation analysis fails to support the latency model for regulation of p53 DNA binding activity in vivo. Proc Natl Acad Sci USA 99(1):95–100

    Article  PubMed  CAS  Google Scholar 

  26. Nelson JD, Denisenko O, Bomsztyk K (2006) Protocol for the fast chromatin immunoprecipitation (ChIP) method. Nat Protoc 1(1):179–185

    Article  PubMed  CAS  Google Scholar 

  27. Hoh J, Jin S, Parrado T, Edington J, Levine AJ, Ott J (2002) The p53MH algorithm and its application in detecting p53-responsive genes. Proc Natl Acad Sci USA 99(13):8467–8472

    Article  PubMed  CAS  Google Scholar 

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Acknowledgment

H.L. was supported by NIH-NCI grants CA127724, CA095441, and CA129828.

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Correspondence to Hua Lu .

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© 2013 Springer Science+Business Media New York

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Liao, JM., Lu, H. (2013). ChIP for Identification of p53 Responsive DNA Promoters. 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_17

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  • DOI: https://doi.org/10.1007/978-1-62703-236-0_17

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-235-3

  • Online ISBN: 978-1-62703-236-0

  • eBook Packages: Springer Protocols

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