Structure & function of the p53 tumor suppressor protein.
Mutation of the p53 tumor suppressor gene is the most frequent lesion detected in cancer. Consequently, understanding the structure and function of the normal p53 protein and how it differs from the mutant p53 proteins that are commonly found in cancer patients' tumors should provide valuable information. P53 sits at the nexus of a complex network of signals from agents of stress such as DNA damage or hypoxia and many others. P53 transmits these signals to genes that control cell cycle arrest, programmed cell death, senescence, angiogenesis and additional processes. It is well established that p53 functions as a DNA binding protein that can activate or repress transcription from numerous genes involved in the above processes. We have analyzed in detail the functional domains of p53 and how they are regulated by both covalent and non-covalent modifiers. The most well studied regulator of p53 is the Mdm2 protein that is itself a transcriptional target of p53. Interestingly, two related genes, p63 and p73, share structural and functional similarities with p53 yet their roles as tumor suppressors are unclear. Our laboratory is currently addressing the following questions about these important proteins:
How does p53 regulate transcription of its target genes? We have had a long-standing interest in understanding the mechanism by which p53 recognizes its binding sites in the promoters of its target genes and how it then works to recruit or work with transcriptional co-factors. Our findings have revealed that p53 can slide on DNA and that its highly basic C-terminus is required for this activity. We found that p53, p63 and p73 share similarities but also there are differences in the ways that they bind to their sites in DNA that may relate to the unique properties of the p53 C-terminus. Our current goal is to deconstruct the modes by which p53 recognizes its sites in nucleosomal and genomic DNA. We then wish to examine how p53 selects its various target genes to regulate different cellular outcomes such as arrest or apoptosis. We discovered hCAS/Cse1L as a novel co-regulator of a sub-set of p53 target genes and cell death. We have also found that when DNA replication is blocked, p53 can facilitate the initial stages of transcription of its target genes, but that in some cases elongation of the mRNA is inhibited. Finally, we have identified interesting mutant forms of p53 that can selectively activate some p53 targets but not others. The goal is to elucidate the mechanism of this p53 target gene selectivity in future studies.
How is the p53/Mdm2 circuit regulated? One of the key interactors with p53 is the Mdm2 protein that inhibits p53 by multiple mechanisms. Mdm2 both directly represses p53’s transcriptional activity and, through its ability to serve as an E3 ubiquitin ligase, can actively degrade p53 in unstressed cells. Various stress signals interfere with the ability of Mdm2 to inhibit p53. Understanding the relationship between Mdm2 and p53 is currently being actively studied in our laboratory. Interestingly Mdm2 |is itself extensively regulated by stress and other signals in cells. We have found that cyclin G1 can regulate this circuit and are examining how cyclin G1 itself is controlled in cells. An important factor in regulation of p53 is a protein closely related to Mdm2, namely MdmX. Mdm2 and MdmX can associate with themselves and with each other. Such complex formation is required for the E3 ligase activity of Mdm2. We are using biochemical and cell based assays to understand how Mdm2 and MdmX together and separately regulate p53 and each other.
How do cells transmit signals from genotoxic stress to regulate p53 and its homologues p63 and p73? Our experiments have focused on the checkpoint kinases Chk1 and Chk2 and their ability to regulate p53 and p73. In some human cells, however, p53 is induced in the absence of Chk1or Chk2, whilst in others Chk2 appears to be important for p53 activation. We would like to identify factors and kinases that regulate p53 when Chk2 is not involved. We are also studying the mechanisms by which the turnover of different p63 isoforms is regulated after DNA damage.
What are the roles of the p63 and p73 genes and how are their protein products regulated? We found that a subset of tumor-derived mutant forms of p53 can down-regulate the normally active forms of p63 and p73. This may partially explain why in some cases mutant forms of p53 appear to serve as pro-oncogenic factors. We seek to clarify when and how mutant p53 proteins regulate p63/p73 proteins and whether this is important to tumorigenesis. We are also in the process of identifying cellular proteins that interact specifically with p63 and p73 and how these interactions affect their functions.
How does p53 promote apoptosis? In some tumor cells p53 will not promote apoptosis unless cells are treated with DNA damaging agents and we hope to determine the mechanism by which such agents facilitate cell death mediated by p53. We have also studied apoptosis caused by a transcriptionally impaired mutant form of p53 and found that there are significant differences as well as similarities when compared to apoptosis caused by wild-type p53.
Finally, with respect to the central role that p53 plays in human cancer, can we use information derived from the basic research on this protein to develop p53-based cancer therapeutics? The search for answers to these and other questions are the basis for much of the work currently going on in our laboratory.
- Moon S-H*, Huang C-H*, Houlihan, SL *, Regunath K, Freed-Pastor WA, Morris IV, JP, Tschaharganeh DF, A, Kastenhuber ER, Barsotti, AM., Culp-Hill, R. Xue W., Ho, Y-J., Baslan, T. Li, X., Mayle, A., de Stanchina., E., Zender L, Tong, DR., D’Alessandro, A., Lowe SW*, and C Prives*.(2019) p53 Represses the Mevalonate Pathway to Mediate Tumor Suppression. Cell. 176(3):564-580.
- Katz C, Low Calle, A., Choe, J., Laptenko O Joseph J S, Garofalo F, Zhu Y, Friedler A, Prives C (2018): Mdm2 preferentially degrades dimer forms of wild-type and mutant p53. Genes & Dev. 32(5-6):430-447.
- Lessel D*, Wu D*, Trujillo C, Ramezani T, Alwasiyah MK, Saha B, Hisama FM, Rading K, Goebel I, Schütz P, Speit G, Högel J, Thiele H, Nürnberg G, Nürnberg P, Hammerschmidt M, Katz C, Martin GM, Oshima J, Prives C*, and C Kubisch* 2017. Dysfunction of the Mdm2/p53 axis is linked to premature aging. J. Clin. Invest. 127: 3598-3608.
- Pfister, N.T., Fomin, V., Regunath, K , Zhou, W., Silwal-Pandit, L., Zhou, J.Y.,., Freed-Pastor,W.A., Laptenko, O., Neo, S. P., Bargonetti, J., Hoque, M., Tian,B., Gunaratne, J., Engebraaten, O., Manley, J.l. Børresen-Dale, A-L., Neilsen, P.M., and C. Prives. 2015 Mutant p53 Cooperates with the SWI/SNF Chromatin Remodeling Complex to Regulate VEGFR2 in Breast Cancer Cells. Genes Dev 29: 1298-315
- Laptenko, O., Shiff, I., Freed-Pastor, W., Zupnik, A., Mattia, M., Freulich1, E., Shamir,I., Kadouri, N., Kahn, T., Manfredi, J., Simon , I., and C. Prives. 2015. The p53 C-Terminus Controls Site-Specific DNA Binding and Promotes Structural Changes within the Central DNA Binding Domain. Mol Cell. 57: 1034-1046.
- Zhu Y, Regunath K, Jacq X, Prives C. 2013. Cisplatin causes cell death via TAB1 regulation of p53/MDM2/MDMX circuitry. Genes Dev. 27: 1739-51.
- Rokudai, S., Laptanko, O., Arnal, S. M., Taya., Y., Kitabayashi, I., and C. Prives. 2013. MOZ increases p53 acetylation and premature senescence through its complex formation with PML. Proc Natl Acad Sci U S A. 110: 3895-3900.
- Singer, SR, Zhao, R,Barsotti, AM, Ouwehand, A, Fazollahi, M, Coutavas, E, Breuhahn, K, Neumann , O, Longerich,T, Pusterla, T, Powers, MA, Giles, KM, Leedman, PJ, Hess,J., Grunwald, D, Bussemaker, HJ, Singer, RH, Schirmacher, P, and C. Prives. 2012 Nuclear pore component Nup98 is a potential tumor suppressor and regulates post-transcriptional expression of select p53 target genes. Mol Cell 48: 799-810.
- Freed-Pastor, WA., Mizuno,H., Zhao, X., Langerød, A., Moon, S-H., Rodriguez- Barrueco , R., Barsotti, A., Chicas\, A., Li, W., Polotskaia, A., Bissell,MJ., Osborne, TF., Tian, B., Lowe, SW., Silva, JM., Børresen-Dale, A-L., Levine, AJ., Bargonetti, J. and C. Prives. (2012) Mutant p53 Disrupts Mammary Acinar Morphogenesis via the Mevalonate Pathway. Cell 244-58: 148.
- Laptenko, O., Beckerman, R., Freulich, E. and C. Prives (2011) p53 binding to nucleosomes within the p21 promoter in vivo leads to nucleosome loss and transcriptional activation. Proc. Natl. Acad. Sci. U.S.A 10385-90: 108.Barsotti, A.M, C. Prives (2010) p53-mediated transcriptional repression: the missing “linc”. Cell 358-60: 142.
- Poyurovsky M.V., Laptenko, O., Beckerman, R., Ahn, J., Lokshin , M., Hener-Katz, C., Mattia M., Zupnick, A., Friedle, A., and C. Prives (2010) Regulation of the p53-Mdm2 Complex via Direct Association of the p53 C-terminus with the N-terminal domain of Mdm2. Nature Structure Molecular Biology 982-9: 8.
- Barsotti, A. M. and C. Prives (2009) Pro-Proliferative FoxM1 is a target of p53- mediated repression Oncogene 4295-4305: 28.
- Zhu, Y., Poyurovsky, M.V., Li, Y., Biderman, L., Stahl J., Jacq, X., and C. Prives (2009) Ribosomal Protein S7 is both a Regulator and Substrate of MDM2. Mol. Cell 316-26: 35.
- Li, Y., Peart, M.J. and Prives (2009) Stxbp4 regulates DNp63 stability by suppression of RACK1-dependent degradation Mol. Cell. Biol 3953-63: 29.
- Beckerman, R., Donner, A.J., Mattia, M., Peart, M.J., Manley J.L., Espinosa J.M. and C. Prives (2009) A role for Chk1 in blocking transcriptional elongation of p21 RNA during the S phase checkpoint. Genes & Dev 1364-77 PMCID: PMC2701578: 11.
- Vousden, K., and C. Prives (2009) Blinded by the Light: the Growing Complexity of p53. Cell 413-431: 137.
- Li, H., Okamoto, K., Peart, M.J. and C. Prives. (2008) Lysine-Independent Turnover of Cyclin G1 can be Stabilized by B’a Subunits of Protein Phosphatase 2A. Mol Cell Biol PMCID: PMC2630686: 919-28.
- Baptiste-Okoh, N. Barsotti, A.M., and C. Prives. (2008) A role for caspase 2 in the process of p53 mediated apoptosis. Proc Natl Acad Sci. US 105: 1937-1942.
- Kass, E. M., Ahn, J. Tanaka, T., Keezer S., and C. Prives. 2007 (2007) Stability of Checkpoint Kinase 2 is Regulated via Phosphorylation at Serine 456. J. Biol. Chem 282: 30311-21.
- Tanaka, T., Ohkubo, S., and C Prives (2007) Nuclear export factor hCAS/CSE1L associates with chromatin and regulates expression of selective p53 target genes. Cell 130: 638-650.
- Karni-Schmidt, O., Friedler, A., A. Zupnick, K. McKinney, M. Sheetz., A. Fersht, and C. Prives (2007) Energy dependent nucleolar localization of p53 in vitro requires two discrete regions within the p53 carboxy terminus Oncogene 26: 3878-91.
- Mattia M., Gottifredi V., McKinney, K. and C. Prives (2007) p53-dependent p21 mRNA elongation is impaired when DNA replication is stalled. Mol. Cell. Biol 27: 1309-20.
- Lokshin, M., Li, H., Gaiddon, C. and C. Prives (2007) p53 and p73 display common and distinct requirements for sequence specific binding to DNA. Nucl. Acids Res. 35: 340-52.
- Poyurovsky, M.V., Priest, C, Kentsis, A., Borden, K.L.B., Pan Z-Q., Pavletich, N., and C. Prives. (2007) The Mdm2 RING domain C-terminus is required for supramolecular assembly and ubiquitin ligase activity EMBO. J 26: 90-101.
- Ohkubo, S., Taya,Y., Kitazato, K. and C. Prives (2006) Excess Mdm2 selectively impairs p53-dependent transcription and apoptosis without the degradation of p53. J. Biol Chem. 281: 16943-50.
- Urist M., Tanaka T., Poyurovsky M V., and Prives C. (2004) p73 Induction after DNA Damage is Regulated by Checkpoint Kinases, Chk1 and Chk2. Genes & Dev 18: 3041-3054.
- McKinney K. M., Mattia M., Gottifedi V., Prives C. (2004) p53 Linear Diffusion Along DNA Requires its C-terminus. Molecular Cell 16: 413-424.
- Baptiste N., Prives C. (2004) p53 in the cytoplasm: a question of overkill? Cell 116: 487-9.
- Ahn J., Urist M., Prives C. (2003) Questioning the role of checkpoint kinase 2 in the p53 DNA damage response. J Biol Chem 278(23): 20480-9. Article
- Gaiddon C., Lokshin M., Gross I., Levasseur D., Taya Y., Loeffler JP., Prives C. (2003) Cyclin-dependent Kinases Phosphorylate p73 at Threonine 86 in a Cell Cycle-dependent Manner and Negatively Regulate p73. J Biol Chem 278(30): 27421-27431. Article
- Poyurovsky M., Jacq X., Ma C., Karni-Schmidt O., Parker P J., Chalfie M., Manley J L and Prives C. (2003) Nucleotide binding by the Mdm2 RING domain facilitates ARF-independent Mdm2 nucleolar loccalization. Molecular Cell 12: 875-887.
- Ahn J., Prives C. (2002) Checkpoint kinase 2 (Chk2) monomers or dimers phosphorylate Cdc25C after DNA damage regardless of threonine 68 phosphorylation. J Biol Chem 277(50): 48418-26. Article
- McKinney K., Prives C. (2002) Efficient specific DNA binding by p53 requires both its central and C-terminal domains as revealed by studies with high-mobility group 1 protein. Mol Cell Biol 22(19): 6797-808. Article
- Urist M., Prives C. (2002) p53 leans on its siblings. Cancer Cell 1(4): 311-3. Article
- Okamoto K., Li H., Jensen MR., Zhang T., Taya Y., Thorgeirsson SS., Prives C. (2002) Cyclin G recruits PP2A to dephosphorylate Mdm2. Mol Cell 9: 761-771. Article
- Baptiste N, Friedlander P, Chen X, Prives C. (2002) The proline-rich domain of p53 is required for cooperation with anti-neoplastic agents to promote apoptosis of tumor cells. Oncogene 21(1): 9-21. Article
- Prives C, Manley J L. (2001) Why is p53 acetylated? Cell 107(7): 815-8. Article
- Ahn J, Prives C. (2001) The C-terminus of p53: the more you learn the less you know. Nat Struct Biol 8(9): 730-2. Article
- Gottifredi V, Shieh S-Y, Taya Y and Prives C. (2001) P53 Accumulates but is Functionally Impaired when DNA Syntheses is Blocked. Proc. Natl. Acad. Sci. U.S.A. 98: 1036-1041.
- Shieh S-Y, Ahn J, Tamai K, Taya Y and Prives C. (2000) The human homologues of checkpoint kinases Chk1 and Cds1 (Chk2) phosphorylate p53 at multiple DNA damage inducible sites. Genes & Dev 14: 289-300.
- Di Como C J, Gaiddon C and Prives C. (1999) p73 function is inhibited by tumor derived p53 mutants in mammalian cells. Mol Cell Biol 19: 1438-1499.
- Jayaraman L, Murthy K G K, Curran T, Xanthoudakis S and Prives C. (1997) Identification of redox/repair protein Ref-1 as an activator of p53. Genes & Dev 11: 558-570.
- Shieh S-Y, Ikeda M, Taya Y and Prives C. (1997) DNA damage-induced phosphorylation of p53 alleviates inhibition by mdm2. Cell 91: 325-334.