|Budget Amount *help
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2002: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2001: ¥2,700,000 (Direct Cost: ¥2,700,000)
The yeast library (mating type a), containing p53 mutant protein expression plasmid accompanied by all the 1 amino-acid substitution resulting from 1 nucleotide substitution, and the yeast (mating type α), containing GFP or the DsRed reporter plasmid which has WAF1, MDM2, BAX, 14-3-3σ, p53AlPl, GADD45, NOXA, or p53R2 promoter, were mated and the transcriptional activation ability of each p53 mutant protein towards each promoter was measured by quantified fluorescence intensity. As a result, 2,314 of p53 mutants were classified that as follows;
(1) mutants which have fufl transcriptional activities towards all examined 8 promoters equivalent to wild type p53 (1,278),
(2) mutants which have no transcriptional activities towards all promoters equivalent to common p53 mutants (400),
(3) mutants which have variation activities depends on the promoters (636).
Comparing the obtained results with p53 mutation data base of International Agency for Research on Cancer (IARC), among the mutants classified (1), 373 of p53 mutants were reported in clinical neoplasm samples. However, reported frequency of these mutants was low (1-17 times), it suggests that there are other target genes important for neoplasm formation or these mutations are accidentally reported. On the other hand, among the mutants classified (2), 39 of p53 mutants were not reported in clinical materials. These data suggest that the DNA damaging stress influences differently on p53 genes. Finally some mutants, which classified (3), were introduced into SaOs2 cell with luciferase plasmids containing WAF1, MDM2, BAK, 14-3-3σ , GADD45, or p53R2 promoter. The promoter selectivity of p53 mutants did not always coincident with the data observed in yeast system, but indeed the promoter selectivity was observed in mammalian cell. It is suggested that the promoter selectivity comes from changing higher structure of p53 proteins due to point mutation.