Identification of p53 as a Sequence-Specific DNA-Binding Protein

Abstract

The tumor-suppressor gene p53 is altered by missense mutation in numerous human malignancies. However, the biochemical properties of p53 and the effect of mutation on these properties are unclear. A human DNA sequence was identified that binds specifically to wild-type human p53 protein in vitro. As few as 33 base pairs were sufficient to confer specific binding. Certain guanines within this 33-base pair region were critical, as methylation of these guanines or their substitution with thymine-abrogated binding. Human p53 proteins containing either of two missense mutations commonly found in human tumors were unable to bind significantly to this sequence. These data suggest that a function of p53 may be mediated by its ability to bind to specific DNA sequences in the human genome, and that this activity is altered by mutations that occur in human tumors.

Targeting gene expression to tumor cells with loss of wild-type p53 function

“…13,[27][28][29] The relevance of this transactivating ability to the tumor suppressor function of p53 is reflected by the fact that ϳ80% of natural genetic changes in TP53 are missense mutations, which are largely confined to its evolutionarily well-conserved and sequence-specific DNA binding central region. 30,31 Furthermore, ϳ30% of mutations are concentrated at several hot spots (e.g., 175r3h, 248r3w, 249r3s, and 273r3h), the original residues of which either directly contact the DNA or preserve the structural integrity of the domain required for DNA binding.…”

“…30,31 Furthermore, ϳ30% of mutations are concentrated at several hot spots (e.g., 175r3h, 248r3w, 249r3s, and 273r3h), the original residues of which either directly contact the DNA or preserve the structural integrity of the domain required for DNA binding. 31 The overwhelming majority of mtp53 molecules have lost the DNA sequence-specific binding capacity necessary for p53 transactivation of gene expression, 27,28,32 demonstrating the importance of this property for tumor suppressor function. 33 p53 also suppresses the transcription of a number of cellular and viral genes with promoters that are devoid of the consensus sequence recognized by p53, including c-fos, c-jun, interleukin-6, 34 Hsp70, 14 proliferating cell nuclear antigen, 35 multidrug resistance gene (MDR1) 36 topoisomerase IIa, 37 and Bcl-2.…”

Additive effect between NF-κB subunits and p53 protein for transcriptional activation of human p53 promoter

“…Moreover, the level of p53 can also be modulated at the transcriptional level after mitogenic stimulation, di erentiation induction and genotoxic stress (Reich and Levine, 1984;Sun et al, 1995;Balint and Reisman, 1996;Kirch et al, 1999). The WT-activated p53 acts as a transcriptional factor and regulates the expression of target genes including p21(WAF1/CIP1), gadd45, bax or bcl2, which in turn control cell cycle checkpoints, DNA repair and apoptosis (Kern et al, 1991;Farmer et al, 1992;Miyashita et al, 1994).…”

p53 protein is activated during muscle differentiation and participates with MyoD in the transcription of muscle creatine kinase gene

“…p53 protein functions as a transcription factor that binds to speci®c DNA sequences and activates gene transcription (Kern et al, 1991;Farmer et al, 1992). Between the genes that have been identi®ed as speci®c targets for p53 protein are p21/WAF, MDM2, GADD45, cyclin G and bax, whose products function as regulators of cell growth (reviewed by Ko and Prives, 1996;Levine, 1997).…”