Given that our knowledge of DNA repair is limited because of the complexity of the DNA system, a technique called UVA micro-irradiation has been developed that can be used to visualize the recruitment of DNA repair proteins at double-strand break (DSB) sites. Interestingly, Hoechst 33258 was used under micro-irradiation to sensitize 5-bromouracil (^BrU)-labelled DNA, causing efficient DSBs. However, the molecular basis of DSB formation under UVA micro-irradiation remains unknown. Herein, we investigated the mechanism of DSB formation under UVA micro-irradiation conditions. Our results suggest that the generation of a uracil-5-yl radical through electron transfer from Hoechst 33258 to ^BrU caused DNA cleavage preferentially at self-complementary 5′-AA^BrU^BrU-3′ sequences to induce DSB. We also investigated the DNA cleavage in the context of the nucleosome to gain a better understanding of UVA micro-irradiation in a cell-like model. We found that DNA cleavage occurred in both core and linker DNA regions although its efficiency reduced in core DNA.

鉴于我们对DNA修复的了解由于DNA系统的复杂性而受到限制，因此开发了一种称为UVA微辐照的技术，该技术可用于可视化双链断裂（DSB）位点上DNA修复蛋白的募集。有趣的是，在微辐射下使用Hoechst 33258来敏化5-溴尿嘧啶（^Br U）标记的DNA，从而产生有效的DSB。但是，在UVA微辐射下形成DSB的分子基础仍然未知。在本文中，我们研究了在UVA微辐照条件下DSB形成的机理。我们的结果表明，通过电子从Hoechst 33258转移至^Br U产生的尿嘧啶-5-基自由基优先导致DNA在自我互补5'-AA ^Br U处的裂解。^Br U-3'序列诱导DSB。我们还研究了核小体背景下的DNA裂解，以更好地了解细胞样模型中的UVA微辐射。我们发现，虽然在核心DNA中效率降低，但在核心和接头DNA区域中都发生了DNA裂解。