Differential Transactivation by the p53 Transcription Factor Is Highly Dependent on p53 Level and Promoter Target Sequence

Abstract

Little is known about the mechanisms that regulate differential transactivation by p53. We developed a system in the yeast Saccharomyces cerevisiae that addresses p53 transactivation capacity from 26 different p53 response elements (REs) under conditions where all other factors, such as chromatin, are kept constant. The system relies on a tightly regulated promoter (rheostatable) that can provide for a broad range of p53 expression. The p53 transactivation capacity toward each 20- to 22-bp-long RE could be ranked by using a simple phenotypic assay. Surprisingly, there was as much as a 1,000-fold difference in transactivation. There was no correlation between the functional rank and statistical predictions of binding energy of the REs. Instead we found that the central sequence element in an RE greatly affects p53 transactivation capacity, possibly because of DNA structural properties. Our results suggest that intrinsic DNA binding affinity and p53 protein levels are important contributors to p53-induced differential transactivation. These results are also relevant to understanding the regulation by other families of transcription factors that recognize several sequence-related response elements and/or have tightly regulated expression. We found that p53 had weak activity towards half the apoptotic REs. In addition, p53 alleles associated with familial breast cancer, previously classified as wild type, showed subtle differences in transactivation capacity towards several REs.

Our thanks go to Richard Iggo for the generous gifts of the yIG397 yeast strain and p53 expression vectors; to Gilberto Fronza for the p53 mutant alleles; to Chris Miller, Dmitry Gordenin, Kirill Lobachev, Yong Hwang Jin, and Karthikeyan Gopalakrishnan for advice, helpful discussions, and comments on the manuscript; and to Trevor Archer and Alex Merrick for critical review.

This work was partially supported by DOD grant DAMD 17-011-0579 to M.A.R.