Influence of hairpins on template reannealing at trinucleotide repeat duplexes: A model for slipped DNA

Hairpin stabilization of polymerase slippage has been proposed as part of the mechanism for large-scale expansion of CG-rich (CNG, where N = A, T, G, or C) trinucleotide repeats. However, hairpin formation does not entirely account for why long repeats but not short repeats or palindromes expand. Using ultraviolet spectroscopic methods, we examine the thermodynamic and kinetic properties of repeating trinucleotides to evaluate their behavior at a slippage site. We find that CNC trinucleotide repeats associated with expansion form stable hairpins whether they are short (with as few as 10 repeats) or long. However, long repeating stretches exist as single strands up to 2 orders of magnitude longer than sequences with either short repeats or random DNA. Thus, long hairpins have long lifetimes even in the presence of their complementary strands and inhibit duplex reannealing at a slippage site. The kinetic properties explain why expansion occurs with high frequency at long repeats but not at short repeats or palindromes.