Abstract
During apoptosis Bcl-2 proteins control permeabilization of the mitochondrial outer membrane leading to the release of cytochrome c. Essential gatekeepers for cytochrome c release are the proapoptotic multidomain proteins, Bax, and Bak. The expression of Bax is upregulated upon cellular stress by the tumor suppressor p53. Despite the high functional homology of Bax and Bak, little is known about how the bak gene is regulated. To investigate its transcriptional regulation in further detail, we have analyzed a region spanning 8200 bp upstream of the bak start codon (within exon 2) for transcription factor-binding sites, and identified three p53 consensus sites (BS1–3). Reporter gene assays in combination with site-directed mutagenesis revealed that only one putative p53-binding site (BS3) is necessary and sufficient for induction of reporter gene expression by p53. Consistently, p53 induces expression of endogenous Bak. At the mRNA level, induction of Bak expression is weaker than induction of Puma and p21. Interestingly, Bak expression can also be induced by p73 that binds however to each of the three p53-binding sites within the bak promoter region. Our data suggest that expression of Bak can be induced by both, p53 and p73 utilizing different binding sites within the bak promoter.
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
Abbreviations
- BS:
-
binding site
- CDK:
-
cyclin-dependent kinase
- EMSA:
-
electrophoretic mobility shift assay
- qRT-PCR:
-
quantitative real-time PCR
References
Danial NN, Korsmeyer SJ . Cell death: critical control points. Cell 2004; 116: 205–219.
Schwerk C, Schulze-Osthoff K . Regulation of apoptosis by alternative pre-mRNA splicing. Mol Cell 2005; 19: 1–13.
Galonek HL, Hardwick JM . Upgrading the BCL-2 network. Nat Cell Biol 2006; 8: 1317–1319.
Kim H, Rafiuddin-Shah M, Tu HC, Jeffers JR, Zambetti GP, Hsieh JJ et al. Hierarchical regulation of mitochondrion-dependent apoptosis by BCL-2 subfamilies. Nat Cell Biol 2006; 8: 1348–1358.
Willis SN, Chen L, Dewson G, Wei A, Naik E, Fletcher JI et al. Proapoptotic Bak is sequestered by Mcl-1 and Bcl-xL, but not Bcl-2, until displaced by BH3-only proteins. Genes Dev 2005; 19: 1294–1305.
Wei MC, Zong WX, Cheng EH, Lindsten T, Panoutsakopoulou V, Ross AJ et al. Proapoptotic Bax and Bak: a requisite gateway to mitochondrial dysfunction and death. Science 2001; 292: 727–730.
Willis SN, Fletcher JI, Kaufmann T, van Delft MF, Chen L, Czabotar PE et al. Apoptosis initiated when BH3 ligands engage multiple Bcl-2 homologs, not Bax or Bak. Science 2007; 315: 856–859.
Gillissen B, Essmann F, Hemmati PG, Richter A, Richter A, Oztop I et al. Mcl-1 determines the Bax dependency of Nbk/Bik-induced apoptosis. J Cell Biol 2007; 179: 701–715.
Nakano K, Vousden KH . PUMA, a novel proapoptotic gene, is induced by p53. Mol Cell 2001; 7: 683–694.
Oda E, Ohki R, Murasawa H, Nemoto J, Shibue T, Yamashita T et al. Noxa, a BH3-only member of the Bcl-2 family and candidate mediator of p53-induced apoptosis. Science 2000; 288: 1053–1058.
Miyashita T, Reed JC . Tumor suppressor p53 is a direct transcriptional activator of the human bax gene. Cell 1995; 80: 293–299.
Miyashita T, Krajewski S, Krajewska M, Wang HG, Lin HK, Liebermann DA et al. Tumor suppressor p53 is a regulator of bcl-2 and bax gene expression in vitro and in vivo. Oncogene 1994; 9: 1799–1805.
Bourgarel-Rey V, Savry A, Hua G, Carre M, Bressin C, Chacon C et al. Transcriptional down-regulation of Bcl-2 by vinorelbine: identification of a novel binding site of p53 on Bcl-2 promoter. Biochem Pharmacol 2009; 78: 1148–1156.
Sohn D, Essmann F, Schulze-Osthoff K, Janicke RU . p21 blocks irradiation-induced apoptosis downstream of mitochondria by inhibition of cyclin-dependent kinase-mediated caspase-9 activation. Cancer Res 2006; 66: 11254–11262.
el-Deiry WS, Kern SE, Pietenpol JA, Kinzler KW, Vogelstein B . Definition of a consensus binding site for p53. Nat Genet 1992; 1: 45–49.
el-Deiry WS, Tokino T, Velculescu VE, Levy DB, Parsons R, Trent JM et al. WAF1, a potential mediator of p53 tumor suppression. Cell 1993; 75: 817–825.
Yu J, Zhang L, Hwang PM, Kinzler KW, Vogelstein B . PUMA induces the rapid apoptosis of colorectal cancer cells. Mol Cell 2001; 7: 673–682.
Melino G, Bernassola F, Ranalli M, Yee K, Zong WX, Corazzari M et al. p73 Induces apoptosis via PUMA transactivation and Bax mitochondrial translocation. J Biol Chem 2004; 279: 8076–8083.
Harms KL, Chen X . The functional domains in p53 family proteins exhibit both common and distinct properties. Cell Death Differ 2006; 13: 890–897.
Kastan MB, Zhan Q, el-Deiry WS, Carrier F, Jacks T, Walsh WV et al. A mammalian cell cycle checkpoint pathway utilizing p53 and GADD45 is defective in ataxia-telangiectasia. Cell 1992; 71: 587–597.
Zauberman A, Flusberg D, Haupt Y, Barak Y, Oren M . A functional p53-responsive intronic promoter is contained within the human mdm2 gene. Nucleic Acids Res 1995; 23: 2584–2592.
Takimoto R, El-Deiry WS . Wild-type p53 transactivates the KILLER/DR5 gene through an intronic sequence-specific DNA-binding site. Oncogene 2000; 19: 1735–1743.
Weinberg RL, Veprintsev DB, Bycroft M, Fersht AR . Comparative binding of p53 to its promoter and DNA recognition elements. J Mol Biol 2005; 348: 589–596.
Riley T, Sontag E, Chen P, Levine A . Transcriptional control of human p53-regulated genes. Nat Rev Mol Cell Biol 2008; 9: 402–412.
Osada M, Park HL, Nagakawa Y, Begum S, Yamashita K, Wu G et al. A novel response element confers p63- and p73-specific activation of the WNT4 promoter. Biochem Biophys Res Commun 2006; 339: 1120–1128.
You H, Pellegrini M, Tsuchihara K, Yamamoto K, Hacker G, Erlacher M et al. FOXO3a-dependent regulation of Puma in response to cytokine/growth factor withdrawal. J Exp Med 2006; 203: 1657–1663.
Ming L, Sakaida T, Yue W, Jha A, Zhang L, Yu J . Sp1 and p73 activate PUMA following serum starvation. Carcinogenesis 2008; 29: 1878–1884.
Chirakkal H, Leech SH, Brookes KE, Prais AL, Waby JS, Corfe BM . Upregulation of BAK by butyrate in the colon is associated with increased Sp3 binding. Oncogene 2006; 25: 7192–7200.
Wang P, Qiu W, Dudgeon C, Liu H, Huang C, Zambetti GP et al. PUMA is directly activated by NF-kappaB and contributes to TNF-alpha-induced apoptosis. Cell Death Differ 2009; 16: 1192–1202.
Morrison DJ, English MA, Licht JD . WT1 induces apoptosis through transcriptional regulation of the proapoptotic Bcl-2 family member Bak. Cancer Res 2005; 65: 8174–8182.
Zhang L, Yu J, Park BH, Kinzler KW, Vogelstein B . Role of BAX in the apoptotic response to anticancer agents. Science 2000; 290: 989–992.
Hemmati PG, Guner D, Gillissen B, Wendt J, von Haefen C, Chinnadurai G et al. Bak functionally complements for loss of Bax during p14ARF-induced mitochondrial apoptosis in human cancer cells. Oncogene 2006; 25: 6582–6594.
Hemmati PG, Gillissen B, von Haefen C, Wendt J, Starck L, Guner D et al. Adenovirus-mediated overexpression of p14(ARF) induces p53 and Bax-independent apoptosis. Oncogene 2002; 21: 3149–3161.
Zender L, Kuhnel F, Kock R, Manns M, Kubicka S . VP22-mediated intercellular transport of p53 in hepatoma cells in vitro and in vivo. Cancer Gene Ther 2002; 9: 489–496.
Gillissen B, Essmann F, Graupner V, Starck L, Radetzki S, Dorken B et al. Induction of cell death by the BH3-only Bcl-2 homolog Nbk/Bik is mediated by an entirely Bax-dependent mitochondrial pathway. Embo J 2003; 22: 3580–3590.
Graupner V, Schulze-Osthoff K, Essmann F, Janicke RU . Functional characterization of p53beta and p53gamma, two isoforms of the tumor suppressor p53. Cell Cycle 2009; 8: 1238–1248.
Grob TJ, Novak U, Maisse C, Barcaroli D, Luthi AU, Pirnia F et al. Human delta Np73 regulates a dominant negative feedback loop for TAp73 and p53. Cell Death Differ 2001; 8: 1213–1223.
Essmann F, Pohlmann S, Gillissen B, Daniel PT, Schulze-Osthoff K, Jänicke RU . Irradiation-induced translocation of p53 to mitochondria in the absence of apoptosis. J Biol Chem 2005; 280: 37169–37177.
Totzke G, Essmann F, Pohlmann S, Lindenblatt C, Jänicke RU, Schulze-Osthoff K . A novel member of the IkappaB family, human IkappaB-zeta, inhibits transactivation of p65 and its DNA binding. J Biol Chem 2006; 281: 12645–12654.
Rothfuss O, Fischer H, Hasegawa T, Maisel M, Leitner P, Miesel F et al. Parkin protects mitochondrial genome integrity and supports mitochondrial DNA repair. Hum Mol Genet 2009; 18: 3832–3850.
Acknowledgements
We thank Bert Vogelstein, Martina Müller-Schilling, Stefan Kubicka and Kirsten Lauber for providing valuable materials and helpful discussion. We also thank Mohamed Hassan for support in EMSA experiments. This work was supported by the Deutsche Forschungsgemeinschaft (SFB773, SFB685, GRK1302) and the BMBF (AID-Net).
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
The authors declare no conflict of interest.
Additional information
Edited by H-U Simon
Supplementary Information accompanies the paper on Cell Death and Differentiation website
Supplementary information
Rights and permissions
About this article
Cite this article
Graupner, V., Alexander, E., Overkamp, T. et al. Differential regulation of the proapoptotic multidomain protein Bak by p53 and p73 at the promoter level. Cell Death Differ 18, 1130–1139 (2011). https://doi.org/10.1038/cdd.2010.179
Received:
Revised:
Accepted:
Published:
Issue date:
DOI: https://doi.org/10.1038/cdd.2010.179
Keywords
This article is cited by
-
Ethyl acetate extract of Elephantopus mollis Kunth induces apoptosis in human gastric cancer cells
BMC Complementary Medicine and Therapies (2021)
-
Eukaryotic initiation factor 5B (eIF5B) provides a critical cell survival switch to glioblastoma cells via regulation of apoptosis
Cell Death & Disease (2019)
-
Apoptosis resistance, mitotic catastrophe, and loss of ploidy control in Burkitt lymphoma
Journal of Molecular Medicine (2015)
-
Terfenadine induces anti-proliferative and apoptotic activities in human hormone-refractory prostate cancer through histamine receptor-independent Mcl-1 cleavage and Bak up-regulation
Naunyn-Schmiedeberg's Archives of Pharmacology (2014)
-
Levels of pro-apoptotic regulator Bad and anti-apoptotic regulator Bcl-xL determine the type of the apoptotic logic gate
BMC Systems Biology (2013)
