Analysis of DNA methylation and chromatin conformation in G-quadruplex regions in gene c-Myc

Abstract

G-quadruplex structures (G4) have been suggested as potential epigenetic regulators. In addition, recent studies have shown that the formation and stabilization of G4 depend on the chromatin status since the presence or absence of G4 in the same genomic sequence is cell-type specific. The enrichment of G4 in the promoters of proto-oncogenes has been widely demonstrated, especially in the c-MYC promoter. C-MYC gene is unregulated in most types of tumors. This proto-oncogene is under strict transcriptional control, as is its mRNA and the MYC protein. Classically, the regulation of this gene involves many regulatory transcriptional motifs found in its proximal promoter region. Recently, great attention has also been given to a regulatory region distal to c-MYC, an enhancer. Enhancers have extensively studied in recent years as changes in their dynamics controlling gene expression have been associated with the development of diseases, especially cancer. In the later project we proposed to investigate whether G4 structures are able to change chromatin dynamics, specifically the frequency of enhancer / promoter interaction in the c-MYC gene, which is enriched by G4 in its promoter and enhancer regions. Furthermore, as G4 stabilizing drugs are considered a potentially antitumoral agent since they have the ability to inhibit proliferating tumor cells, we also proposed to investigate whether the stabilization of these structures can alter the epigenome, specifically the DNA methylation pattern, in regulatory regions. This project is part of an Auxílio Regular recently approved by Fapesp. In this Scientific Initiation project, we propose experiments and analyzes that will be part of the aforementioned project. Accordingly, this study will be composed of cell culture experiments, standardization of the drug concentration and all the experiments that involve the analysis of DNA (DNA methylation) and gene expression. This work is relevant for the field as today the 3D genome organization and its promoter-enhancer interactions have been the focus of many studies. The more we know about this mechanism of gene regulation, the easier it becomes to understand the roles of genetic polymorphisms and the epimutations present in these regions which also facilitates the use of drugs that can modulate this mechanism. In addition, as the studied gene is relevant in cancer research, understanding its regulation and possible modulation through a drug that acts on its structures is of great importance for the research.