AGG Interruptions in (CGG)n DNA Repeat Tracts Modulate the Structure and Thermodynamics of Non-B Conformations in Vitro

Abstract

The trinucleotide repeat sequence CGG/CCG is known to expand in the human genome. This expansion is the primary pathogenic signature of fragile X syndrome, which is the most common form of inherited mental retardation. It has been proposed that formation of non-B conformations by the repetitive sequence contributes to the expansion mechanism. It is also known that the CGG/CCG repeat sequence of healthy individuals, which is not prone to expansion, contains AGG/CCT interruptions every 8-11 CGG/CCG repeats. Using DNA containing 19 or 39 CGG repeats, we have found that both the position and number of interruptions modulate the non-B conformation adopted by the repeat sequence. Analysis by chemical probes revealed larger loops and the presence of bulges for sequences containing interruptions. Additionally, using optical analysis and calorimetry, the effect of these structural changes on the thermodynamic stability of the conformation has been quantified. Notably, changing even one nucleotide, as occurs when CGG is replaced with an AGG interruption, causes a measurable decrease in the stability of the conformation adopted by the repeat sequence. These results provide insight into the role interruptions may play in preventing expansion in vivo and also contribute to our understanding of the relationship between non-B conformations and trinucleotide repeat expansion.