DNA damage induces long range changes to duplex structure - a non-protein start to damage detection?

Abstract

DNA-binding proteins must quickly locate specific sites on DNA to enable replication, repair, and transcription. While sequence-specific recognition is well understood, the physical basis of structure-specific recognition remains unclear, limiting our understanding of DNA damage repair. Proteins must distinguish damaged sites within largely undamaged DNA; however, studying this is challenging due to DNAs dynamic nature. We hypothesised that DNA damage causes changes in DNA structure, signalling protein recruitment. Using confocal single-molecule FRET, we analysed seven DNA duplexes containing modifications such as ribonucleotide, 8-oxoguanine (8-oxoG), abasic sites, nicks, and gaps, which are all involved in the base excision repair (BER) pathway. Each construct was measured with nine dye pairs in triplicate to capture changes in bending, twisting, and stretching. An automated analysis pipeline processed 162 measurements, enabling rigorous statistical comparisons. All modifications altered FRET efficiencies compared to undamaged DNA, including the subtlest change: a single oxygen difference (ribo-vs deoxyribonucleotide). Abasic sites, nicks, and gaps had the greatest effects. These findings provide direct evidence that DNA damage affects duplex structure and dynamics beyond the lesion site, suggesting DNA flexibility changes may act as an early signal for repair protein recruitment. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=102 SRC="FIGDIR/small/709887v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@a85839org.highwire.dtl.DTLVardef@3813dborg.highwire.dtl.DTLVardef@19fa06aorg.highwire.dtl.DTLVardef@dc9729_HPS_FORMAT_FIGEXP M_FIG C_FIG