Effect of the Presence of G-quadruplex Prone DNA Sequences on Transcription Induced by Cancer-relevant P53 Tumor Suppressor Mutants

abstract

English

The p53 protein plays an indispensable role in cells as a guardian of the genome and tumor suppressor. Loss or mutation of the TP53 gene leads to reduced production of wild-type protein or formation of its mutant forms. Reduced function of P53 or complete loss of the gene may contribute to genomic instability and promote tumorigenesis, proliferation of tumor cells, and generation of resistance to commonly used medical treatments. Similarly, G-quadruplex motifs (G4s) forming at the wrong time or place in the genome may also contribute to genomic instability. G4s are secondary non canonical DNA (or RNA) structures. The presence of G4s has been localized in important regions of the genome, such as telomeres and promoters of oncogenes and the genomes of many viruses and pathogens. The involvement of G4s in the regulation of gene expression and its localization in important regions of the genome make G4s a promising target in cancer therapy. In this study, the effect of G4 prone DNA sequences on the transactivation properties of cancer-relevant P53 partial loss-of-function mutants in the yeast one-hybrid expression system (Y1H) was investigated. Seven isogenic Saccharomyces cerevisiae yLFM strains containing the identified KSHV-derived G4 prone DNA sequences placed downstream of the p53 responsive element of the BBC3 gene (PUMA) were used. The activity of the mutant P53s and P53 wild-type proteins was monitored under an inducible promoter to obtain moderate or high levels of protein expression. G4 prone DNA sequences were themselves biophysically characterized by ThT assay and CD spectroscopy. The results of functional assays in Y1H showed that basal reporter transcription in the absence of specific transcription factors is stimulated by the presence of G4, correlating proportionally with the propensity to form G4 structures. In association with the higher constitutive activity of the yeast minimal promoter containing G4, the transcriptional activity of mutant P53 proteins (A161T, R213L, N235S, V272L, R282W, R283C, R337C, R337H, and G360V) was investigated. Our results demonstrated that the G4 forming sequences presence increase the transactivation ability of partial function P53 family proteins. These observations are pointing to the importance of DNA structural characteristics for accurate classification of P53 protein functionality in the context of the wide variety of TP53 germline and somatic mutations.

P53 mutant proteins, G-quaduplexes, transcription activity

KRATOCHVILOVÁ, L.; VOJSOVIČ, M.; VALKOVÁ, N.; BRÁZDA, V.

23. 4. 2024

Czech Chemical Society

2336-7210

Czech Chemical Society Symposium Series

22

1

Czech Republic

23

24

2

http://www.ccsss.cz/index.php/ccsss/issue/view/43/77