Brouwer Ineke, Zhang Hongshan, Candelli Andrea, Normanno Davide, Peterman Erwin J G, Wuite Gijs J L, Modesti Mauro
Department of Physics and Astronomy and LaserLab, Vrije Universiteit Amsterdam, 1081 HV Amsterdam, the Netherlands.
Cancer Research Center of Marseille, CNRS UMR7258, Inserm U1068, Institut Paoli-Calmettes, Aix-Marseille Université UM105, 13273 Marseille, France.
Cell Rep. 2017 Mar 21;18(12):2845-2853. doi: 10.1016/j.celrep.2017.02.068.
Human RAD52 promotes annealing of complementary single-stranded DNA (ssDNA). In-depth knowledge of RAD52-DNA interaction is required to understand how its activity is integrated in DNA repair processes. Here, we visualize individual fluorescent RAD52 complexes interacting with single DNA molecules. The interaction with ssDNA is rapid, static, and tight, where ssDNA appears to wrap around RAD52 complexes that promote intra-molecular bridging. With double-stranded DNA (dsDNA), interaction is slower, weaker, and often diffusive. Interestingly, force spectroscopy experiments show that RAD52 alters the mechanics dsDNA by enhancing DNA flexibility and increasing DNA contour length, suggesting intercalation. RAD52 binding changes the nature of the overstretching transition of dsDNA and prevents DNA melting, which is advantageous for strand clamping during or after annealing. DNA-bound RAD52 is efficient at capturing ssDNA in trans. Together, these effects may help key steps in DNA repair, such as second-end capture during homologous recombination or strand annealing during RAD51-independent recombination reactions.
人类RAD52可促进互补单链DNA（ssDNA）的退火。为了解其活性如何整合到DNA修复过程中，需要深入了解RAD52与DNA的相互作用。在此，我们可视化了单个荧光RAD52复合物与单个DNA分子的相互作用。与ssDNA的相互作用迅速、稳定且紧密，其中ssDNA似乎缠绕在促进分子内桥接的RAD52复合物周围。与双链DNA（dsDNA）的相互作用较慢、较弱且通常具有扩散性。有趣的是，力谱实验表明，RAD52通过增强DNA柔韧性和增加DNA轮廓长度来改变dsDNA的力学性质，提示存在嵌入作用。RAD52结合改变了dsDNA过度拉伸转变的性质并防止DNA解链，这有利于退火过程中或退火后的链钳制。结合DNA的RAD52能够有效地反式捕获ssDNA。总之，这些效应可能有助于DNA修复中的关键步骤，例如同源重组过程中的第二末端捕获或RAD51非依赖性重组反应中的链退火。
