Lee Jaehyoun, Lee Tae-Hee
Department of Chemistry, The Pennsylvania State University , University Park, Pennsylvania 16802, United States.
Biochemistry. 2017 Feb 21;56(7):977-985. doi: 10.1021/acs.biochem.6b01252. Epub 2017 Feb 8.
Nucleosomes impose physical barriers to DNA-templated processes, playing important roles in eukaryotic gene regulation. DNA is packaged into nucleosomes by histone proteins mainly through strong electrostatic interactions that can be modulated by various post-translational histone modifications. Investigating the dynamics of histone dissociation from the nucleosome and how it is altered upon histone modifications is important for understanding eukaryotic gene regulation mechanisms. In particular, histone H2A-H2B dimer displacement in the nucleosome is one of the most important and earliest steps of histone dissociation. Two conflicting hypotheses on the requirement for dimer displacement are that nucleosomal DNA needs to be unwrapped before a dimer can displace and that a dimer can displace without DNA unwrapping. In order to test the hypotheses, we employed three-color single-molecule FRET and monitored in a time-resolved manner the early kinetics of H2A-H2B dimer dissociation triggered by high salt concentration and by histone chaperone Nap1. The results reveal that dimer displacement requires DNA unwrapping in the vast majority of the nucleosomes in the salt-induced case, while dimer displacement precedes DNA unwrapping in >60% of the nucleosomes in the Nap1-mediated case. We also found that acetylation at histone H4K16 or H3K56 affects the kinetics of Nap1-mediated dimer dissociation and facilitates the process both kinetically and thermodynamically. On the basis of these results, we suggest a mechanism by which histone chaperone facilitates H2A-H2B dimer displacement from the histone core without requiring another factor to unwrap the nucleosomal DNA.
核小体对DNA模板化过程形成物理障碍，在真核基因调控中发挥重要作用。DNA主要通过组蛋白经强烈静电相互作用包装成核小体，这种相互作用可被各种组蛋白翻译后修饰所调节。研究组蛋白从核小体上解离的动力学以及组蛋白修饰后其如何改变，对于理解真核基因调控机制至关重要。特别是，核小体中组蛋白H2A-H2B二聚体的置换是组蛋白解离最重要且最早的步骤之一。关于二聚体置换需求存在两个相互矛盾的假说：一是在二聚体能够置换之前核小体DNA需要解旋，二是二聚体可以在不解旋DNA的情况下进行置换。为了验证这些假说，我们采用三色单分子荧光共振能量转移技术，并以时间分辨的方式监测高盐浓度和组蛋白伴侣Nap1触发的H2A-H2B二聚体解离的早期动力学。结果表明，在盐诱导的情况下，绝大多数核小体中二聚体置换需要DNA解旋，而在Nap1介导的情况下，超过60%的核小体中二聚体置换先于DNA解旋。我们还发现组蛋白H4K16或H3K56的乙酰化影响Nap1介导的二聚体解离动力学，并在动力学和热力学上都促进了这一过程。基于这些结果，我们提出了一种机制，即组蛋白伴侣促进H2A-H2B二聚体从组蛋白核心上置换，而无需另一个因子来解旋核小体DNA。
