Wild-Type p53 Protein

Normal p53 gene is a critical controller of normal growth and homeostasis of cells and tissues. It acts as a guardian of the genome by preventing the proliferation of cells with 

FIGURE 10.6. Diagram of the binding sites of the antibodies used for p53 immunohistochemistry. Reproduced, with permission, from Bass etal. (1994). Copyright 1994 John Wiley Sons Limited. 

The p53 gene product is not the only factor that induces cell cycle arrest or programmed cell death (apoptosis). Two other genes, p73 and p63, encode proteins with transactivation, DNA-binding, and tetramerization domains, and they share considerable homology with p53. Like p53, these proteins also induce cell cycle arrest and apoptosis. Each of these proteins is comprised of several isoforms. The p73 protein is a structural and functional homologue of the p53 protein. 

The above findings define a proapoptotic signaling pathway involving p73 and c-Abl. Unlike p53, p73 protein levels do not increase following genotoxic stress. Moreover, although c-Abl interacts with p53 in an irradiated cell, it does not phosphorylate p53 but still contributes to radiation-induced G arrest by a p53-dependent mechanism (Yuan et al., 1999). Although p73 is related to p53, p53 alone is the tumor suppressor. p73 protein as yet has not been localized immunohistochemically. 

TABLE 10.4. p53 Amino Acid Sequence Specificity to Antibodies  

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Kraus, A., et ah. Expression of alternatively spliced mdm2 transcripts correlates with stabilized wild-type p53 protein in human glioblastoma cells. IntJ Cancer, 1999, 80(6), 930-4. 

Keleti, J., Quezado, M., Abaza, M., Raffeld, M., and Tsokos, M. (1996). The Mdm2 oncoprotein is overexpressed in rhabdomyosarcoma cell lines and stabilizes wild type p53 protein, Amer J Pathol 149,143-151. 

Monoclonal antibody PAb 240 (Oncogene Science, Uniondale, NY) recognizes a linear epitope clearly defined as being within the central core of the mutant p53 molecule (Gannon et al., 1990). The epitope for this antibody is cryptic in the active DNA binding form of wild-type p53 but is exposed at the surface of many mutant p53 proteins as well as of denatured wild-type p53 protein. 

Wild-type p53 protein is difficult to detect immunohistochemically in normal cells because of its very short half-life (—20 min) and its presence in minute amounts. However, wild-type p53 protein accumulation can be detected by using antigen retrieval techniques (Dowel and Ogden, 1996 Hall and Lane, 1994). On the other hand, because mutant p53... 

Landers, J.E., Cassel, S.L. and George, D.L. (1997) Translational enhancement of MDM2 oncogene expression in human tumor cells containing a stablized wild-type p53 protein. Cancer Res., 57, 3562-3568. 

Moll UM, LaQuaglia M, Benard J, Riou G. 1995. Wild-type p53 protein undergoes cytoplasmic sequestration in undifferentiated neuroblastomas but not in differentiated tumors. Proc Natl Acad Sci USA 92(10) 4407 4411. 

Like sulforaphane, however, PEITC-NAC (N-acetylcysteine) conjugate appears to block in Gl. Studies by Lund et al. show that in the case of the colorectal cell line HT29, which lacks wild type p53, treatment with AITC causes the cells to detach from the substratum but, at least in the short-term, they do not then enter apoptosis . Where it does occur, induction of apoptosis by isothiocyanate appears to be a p53-dependent process. However, this statement must also be qualified, because the effect appears to depend on which metabolite is considered. For example, sulforaphane does appear to be able to induce apoptosis in HT29 cells, which express a mutated form of the protein. 

Wild-type p53 will downregulate the levels of microtubule associated protein 4 (MAIM) so that, when p53 function is absent, the high levels of MAP-4 will stabilize microtubules and sensitize cells to paclitaxel therapy. 

Yen N, Ioannides CG, Xu K, et al. Cellular and humoral immune responses to adenovirus and p53 protein antigens in patients following intratumor injection of an adenovirus vector expressing wild-type p53 (Ad-p53). Cancer Gene Ther 2000 7 530-536. 

Normal p53 protein binds DNA in a sequence-specific manner and thus most likely regulates gene transcription. Co-transfection experiments show that wild-type p53 activates the expression of genes adjacent to a p53 DNA-bind-ing site. Cells bearing oncogenic forms of p53 have lost this activity. 

Mack, D. H., J. Vartikar, J. M. Pipas, and L. A. Laimins, Specific repression of TATA-mediated but not initiator-mediated transcription by wild-type p53. Nature 363 281-283, 1993. The p53 protein may repress the activity of certain promoters by direct interaction with TATA box-dependent transcription machinery. 

Clinical studies on breast cancer patients have disclosed that an increased p53 protein expression or mutant p53 seem to be associated with an improved effect by adjuvant postoperative radiotherapy (12,15). The latter clinical findings are partly supported by radiotherapy studies on lymphoblastoid human cell lines (46). In this study radiation was capable of inducing apoptosis at an equivalent frequency in both mutant and wild-type p53, but with delayed kinetics (46). 

The DNA tumor virus adenovirus produces a 55-kDa protein from the ElB region of its genome, which binds and inactivates p53. It was hypothesized that an adenovirus lacking ElB would not be able to replicate in normal cells but would in cancer cells lacking p53 function. For this reason, ONYX-015, an ElB gene-attenuated adenovirus was compared with normal adenovirus in human and colonic cancer cell lines with and without p53 function. As expected, the ONYX-015 virus replicated as efficiently as the normal virus in the cell line lacking wild-type p53, but not in the line with normal p53 function. This vector is in early clinical trials. 

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