Slippage Reconfiguration of Trinucleotide Repeat Hairpins Revealed by Single-molecule Fluorescence Resonance Energy Transfer Spectroscopy

I-Ren Lee^1*, Cheng-Wei Ni¹, Yu-Chi Kuang¹, Si-Yu Chen¹, Yu-Jie Wei¹, Si-Yu Chen¹, Kai-Chun Cheng¹

¹Department of Chemistry, National Taiwan Normal University, Taipei, Taiwan

* Presenter:I-Ren Lee, email:irenlee@ntnu.edu.tw

Abnormal expansion of trinucleotide repeat (TNR) sequences is implicated in numerous neurodegenerative disorders. These repeats can adopt noncanonical secondary structures that promote slippage of replication, recombination, or repair machinery, ultimately leading to genomic instability. To elucidate how repeat length influences the intrinsic dynamics of TNR hairpins, we employed single-molecule FRET spectroscopy. Our results reveal that, despite their high thermodynamic stability, TNR hairpins undergo slippage reconfiguration driven by localized structural instability—a dynamic behavior that persists in disease-associated long repeats. We further found that sequence interruptions markedly slow this slippage reconfiguration, potentially preventing the abnormal expansion observed in uninterrupted TNRs. Moreover, the slippage motion impedes the ability of single-stranded DNA binding proteins to resolve these structures, highlighting the pivotal role of DNA slippage reconfiguration in maintaining genome stability.

Keywords: Trinucleotide repeat expansion , Genome stability, DNA hairpin dynamics, Single-molecule FRET, Slippage reconfiguration