Analysis of the DNA-binding activity of p53 mutants using functional protein microarrays and its relationship to transcriptional activation

作者：Malcikova Jitka; Tichy Boris; Damborsky Jiri; Kabathova Jitka; Trbusek Martin; Mayer Jiri; Pospisilova Sarka^*

来源：Biological Chemistry, 2010, 391(2-3): 197-205.

DOI：10.1515/BC.2010.027

• R337C
• real-time PCR
• reporter gene assay
• transactivation

Sequence-specific DNA binding is the key function through which tumor suppressor p53 exerts transactivation of the downstream target genes, often being impaired in cancer cells by mutations in the TP53 gene. Functional protein microarray technology enables a high-throughput parallel analysis of protein properties within one experiment under the same conditions. Using an array approach, we analyzed the DNA binding activity of wild type p53 protein and of 49 variants. Our results show significant differences in the binding properties between the p53 mutants. The C-terminal mutant R337C displayed the highest DNA binding activity on the array. However, the same mutant showed only a partial activation in the reporter gene assay and almost no activation of downstream target genes after transfection of expression vector into cells lacking endogenous p53. These observations demonstrate that DNA binding itself is not sufficient for activating the p53 target genes in at least some of the p53 mutants and, therefore, in vitro studies might not always reflect in vivo conditions.

• 出版日期2010-3