Abstract

Abberant hypomethylation in DNA regions with noncanonical folding potential (ncDNA motifs) is believed to predetermine tumor development - presumably, by facilitating G-quadruplex (G4) and/or i-motif (IM) formation via altering nucleosome positioning (stable G4s induce subsequent genomic rearrangements). We questioned whether CpG methylation per se affects the dsDNA-ncDNA equilibrium. Thermodynamic studies of genomic and model oligonucleotides with methylated CpG sites at different positions are reported. The genomic oligonucleotides analyzed in this work are DNA fragments with reportedly different methylation statuses in colorectal cancer and normal cells. Free energies of duplex, ncDNA formation from single strands were calculated based on melting curve analyses. Polyethylenglycole was used to imitate crowding effect. Our results suggest that CpG methylation may alter the energetic barrier for dsDNA-IM transitions.

Keywords: DNA secondary structures; Methylation; G-quadruplexes; i-motifs; Thermodynamic stability; Molecular crowding

Abbreviations: CD: Circular Dichroism; CGI: Cpg Islands; Etbr: Ethidium Bromide; G4: G-Quadruplex; HPLC: High Performance Liquid Chromatography; IM: Intercalated Motif (I-Motif); MALDI-TOF; MS: Matrix Assisted Laser Desorption Ionization: Time of Flight Mass Spectrometry; Ncdna: Noncanonical DNA Structures; ON: Oligonucleotide; PEG: Polyethylene Glycol; RRT: Rotational Relaxation Time; TDS: Thermal Difference Spectrum