Elbel Tabea, Langowski Jörg
German Cancer Research Center (DKFZ), Biophysics of Macromolecules (B040), Im Neuenheimer Feld 580, D-69120 Heidelberg, Germany.
J Phys Condens Matter. 2015 Feb 18;27(6):064105. doi: 10.1088/0953-8984/27/6/064105. Epub 2015 Jan 7.
Nucleosomes have to open to allow access to DNA in transcription, replication, and DNA damage repair. Changes in DNA torsional strain (e.g. during transcription elongation) influence the accessibility of nucleosomal DNA. Here we investigated the effect of DNA supercoiling-induced torsional strain on nucleosome structure and stability by scanning force microscopy (SFM) and fluorescence correlation spectroscopy (FCS). Nucleosomes were reconstituted onto 2.7 kb DNA plasmids with varying superhelical densities. The SFM results show a clear dependence of the amount of DNA wrapped around the nucleosome core on the strength and type of supercoiling. Negative supercoiling led to smaller nucleosome opening angles as compared to relaxed or positively supercoiled DNA. FCS experiments show that nucleosomes reconstituted on negatively superhelical DNA are more resistant to salt-induced destabilization, as seen by reduced H2A-H2B dimer eviction from the nucleosome. Our results show that changes in DNA topology, e.g. during transcription elongation, affect the accessibility of nucleosomal DNA.
核小体必须打开以允许在转录、复制和DNA损伤修复过程中接触DNA。DNA扭转应变的变化（例如在转录延伸过程中）会影响核小体DNA的可及性。在这里，我们通过扫描力显微镜（SFM）和荧光相关光谱（FCS）研究了DNA超螺旋诱导的扭转应变对核小体结构和稳定性的影响。将核小体重组到具有不同超螺旋密度的2.7 kb DNA质粒上。SFM结果表明，缠绕在核小体核心周围的DNA量明显依赖于超螺旋的强度和类型。与松弛或正超螺旋DNA相比，负超螺旋导致核小体开口角度更小。FCS实验表明，重组在负超螺旋DNA上的核小体对盐诱导的去稳定作用更具抗性，这表现为从核小体中逐出的H2A-H2B二聚体减少。我们的结果表明，DNA拓扑结构的变化，例如在转录延伸过程中，会影响核小体DNA的可及性。
