A mutation in the p53 tumor suppressor gene of AHH-1 [formula omitted] human lymphoblastoid cells

Abstract

Loss-of-function mutations in the p53 tumor suppressor gene result in an altered response to DNA-damaging agents. Included in the mutant p53 phenotype are the loss of the G 1 checkpoint and delayed apoptotic cell death, characteristics we have consistently observed in the the AHH-1 tk + − cell line following exposure to DNA-damaging agents. In order to determine the functional status of p53 in the AHH-1 tk + − cell line, molecular analysis (single-strand conformational polymorphism [SSCP] and sequence analysis) was performed on exons 5–9 of the p53 gene. In addition, the status of the p53 gene in the closely related lymphoblast line, MCL-5, which, in our hands, has a much higher spontaneous rate of apoptosis than AHH-1 tk + − , was also determined by molecular analysis. Initial SSCP analysis of AHH-1 tk + − revealed an abnormal migration pattern of exon 8 when compared to a wild-type control. Subsequent sequence analysis indicated that a base-pair substitution ( C GG → T GG) mutation had occurred at codon 282, a reported ‘hot spot’ for 5-methylcytosine mutations in the human p53 gene. Neither SSCP nor sequence analysis of exons 5–9 of MCL-5 indicated any differences from wild-type DNA. These results suggest that the lack of a G 1 arrest and the delayed entrance into apoptosis observed in chemically-exposed AHH-1 tk + − cells are, at least partially, accounted for by a loss-of-function mutation in the p53 gene.