Abstract
Expansions of trinucleotide repeats (TNRs) are associated with genetic disorders such as Friedreich’s ataxia. The tumor suppressor p53 is a central regulator of cell fate in response to different types of insults. Sequence and structure-selective modes of DNA recognition are among the main attributes of p53 protein. The focus of this work was analysis of the p53 structure-selective recognition of TNRs associated with human neurodegenerative diseases. Here, we studied binding of full length p53 and several deletion variants to TNRs folded into DNA hairpins or loops. We demonstrate that p53 binds to all studied non-B DNA structures, with a preference for non-B DNA structures formed by pyrimidine (Py) rich strands. Using deletion mutants, we determined the C-terminal DNA binding domain of p53 to be crucial for recognition of such non-B DNA structures. We also observed that p53 in vitro prefers binding to the Py-rich strand over the purine (Pu) rich strand in non-B DNA substrates formed by sequence derived from the first intron of the frataxin gene. The binding of p53 to this region was confirmed using chromatin immunoprecipitation in human Friedreich’s ataxia fibroblast and adenocarcinoma cells. Altogether these observations provide further evidence that p53 binds to TNRs’ non-B DNA structures.
Highlights
Wild type p53 (p53) is an important transcription factor that plays a significant role in the prevention of tumor cells development and is widely known as “the guardian of the genome and epigenome” [1,2]
We show the binding affinity of p53 for non-B DNA structures formed by GAA, TTC, CTG and CAG trinucleotide repeats
Formation of non-B DNA structures inside trinucleotide repeats (TNRs) sequences is correlated to TNR expansion, that is in turn associated with the progression of a range of human diseases [26]
Summary
Wild type p53 (p53) is an important transcription factor that plays a significant role in the prevention of tumor cells development and is widely known as “the guardian of the genome and epigenome” [1,2]. In the case of hairpin formed by CTG-strand, p53 binding improved proportionally to the size of the repeats from one to seven repeats [19] Another example of TNR associated disease is Friedreich’s ataxia, which is connected with an unstable (GAA)n·(TTC)n trinucleotide repeat expansion in the first intron of the frataxin gene (FXN, chromosome 9q13 [12,20,21]). For the first time, we detect p53 interacting with the trinucleotide repeats region of the first intron of frataxin gene folded in vitro into non-B DNA structures and p53 binding to this region in adenocarcinoma cells (MCF7) and Friedreich’s ataxia fibroblast cells (FXN 4654) upon p53 stabilization These observations provide further evidence that it could be useful to investigate how p53 influences genetic instabilities of TNR sequences that are important for some human diseases
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