Polach K J, Lowary P T, Widom J
Department of Biochemistry Molecular Biology, and Cell Biology, Northwestern University, Evanston, IL 60208-3500, USA.
J Mol Biol. 2000 Apr 28;298(2):211-23. doi: 10.1006/jmbi.2000.3644.
The N and C-terminal tail domains of the core histones play important roles in gene regulation, but the mechanisms through which they act are not known. These tail domains are highly positively charged and are the sites of numerous post-translational modifications, including many sites for lysine acetylation. Nucleosomes in which these tail domains have been removed by trypsin remain otherwise intact, and are used by many laboratories as a model system for highly acetylated nucleosomes. Here, we test the hypothesis that one role of the tail domains is to directly regulate the accessibility of nucleosomal DNA to other DNA-binding proteins. Three assays are used: equilibrium binding by a site-specific, DNA-binding protein, and dynamic accessibility to restriction enzymes or to a non-specific exonuclease. The effects of removal of the tail domains as monitored by each of these assays can be understood within the framework of the site exposure model for the dynamic equilibrium accessibility of target sites located within the nucleosomal DNA. Removal of the tail domains leads to a 1.5 to 14-fold increase in position-dependent equilibrium constants for site exposure. The smallness of the effect weighs against models for gene activation in which histone acetylation is a mandatory initial event, required to facilitate subsequent access of regulatory proteins to nucleosomal DNA target sites. Alternative roles for histone acetylation in gene regulation are discussed.
核心组蛋白的N端和C端尾部结构域在基因调控中发挥着重要作用,但其作用机制尚不清楚。这些尾部结构域带有大量正电荷,是众多翻译后修饰的位点,包括许多赖氨酸乙酰化位点。通过胰蛋白酶去除这些尾部结构域的核小体在其他方面保持完整,许多实验室将其用作高度乙酰化核小体的模型系统。在此,我们检验了一个假说,即尾部结构域的一个作用是直接调节核小体DNA对其他DNA结合蛋白的可及性。我们使用了三种测定方法:位点特异性DNA结合蛋白的平衡结合,以及对限制性内切酶或非特异性核酸外切酶的动态可及性。通过这些测定方法监测到的去除尾部结构域的影响,可以在核小体DNA内靶位点动态平衡可及性的位点暴露模型框架内得到理解。去除尾部结构域会导致位点暴露的位置依赖性平衡常数增加1.5至14倍。这种影响较小,不利于将组蛋白乙酰化作为促进调节蛋白随后进入核小体DNA靶位点所需的强制性初始事件的基因激活模型。文中还讨论了组蛋白乙酰化在基因调控中的其他作用。