CHARACTERIZATION OF RARE P53 MUTANTS FROM CARCINOGEN-TREATED ALBUMIN-SIMIAN VIRUS-40 T-ANTIGEN TRANSGENIC RATS

The p53 gene has been either mutated or deleted in most human tumors e xamined to date. Mutations in the specific DNA-binding domain are the most common p53 mutations and are of interest because they may produce p53 molecules with transcriptional capabilities unlike those of the w ild-type (WT) p53 protein. Mutations in the rat p53 gene were found in hepatic neoplasms of carcinogen-treated transgenic rats that express simian virus 40 (SV40) large T-antigen (TAg). Because this result was unexpected, we examined some of the biochemical and biological propert ies of the mutant proteins. Corresponding nucleotide changes were made by site-directed mutagenesis of the rat p53 cDNA, which was then inse rted into a eukaryotic expression vector and transfected into the huma n hepatocyte cell line Hep 3B. Four of the mutant p53 molecules from r at hepatomas retained a strict WT conformation. Two others existed in both WT and mutant conformations. All of the mutant proteins were able to bind TAg as well as WT p53 did. Whereas the WT p53 protein was abl e to repress expression of a reporter gene containing a p53-response e lement (pSV(2)CAT), the missense-mutant p53 proteins induced transcrip tion of the reporter to an extent equivalent to that of TAg. The mutan t protei ns a Iso allowed TAg to induce the pSV(2)CAT reporter gene. T he mutant molecules were able to enhance survival of Hep 33 cells, per haps by preventing cell death, whereas expression of the WT p53 protei n caused a reduction in cell number to nearly 10% of control levels. T he results of these experiments suggest that the mutant p53 molecules observed in the carcinogen-treated transgenic rats may have unique pro perties that are important in carcinogenesis. (C) 1998 Wiley-Liss, Inc .