Linker Histones Stabilize the Intrinsic Salt-Dependent Folding of Nucleosomal Arrays: Mechanistic Ramifications for Higher-Order Chromatin Folding

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• 作者：Lenny M. Carruthers ; Jan Bednar ; Christopher L. Woodcock ; and Jeffrey C. Hansen
• 刊名：Biochemistry
• 出版年：1998
• 出版时间：October 20, 1998
• 年：1998
• 卷：37
• 期：42
• 页码：14776 - 14787
• 全文大小：255K
• 年卷期：v.37,no.42(October 20, 1998)
• ISSN：1520-4995

文摘

Defined nucleosomal arrays reconstituted from core histone octamers and twelve 208 bp tandemrepeats of Lytechinus 5S rDNA (208-12 nucleosomal arrays) possess the ability to form an unstable foldedspecies in MgCl₂ whose extent of compaction equals that of canonical higher-order 30 nm diameterchromatin structures [Schwarz, P. M., and Hansen, J. C. (1994) J. Biol. Chem. 269, 16284-16289]. Toaddress the mechanistic functions of linker histones in chromatin condensation, purified histone H5 hasbeen assembled with 208-12 nucleosomal arrays in 50 mM NaCl. Novel purification proceduressubsequently were developed that yielded preparations of 208-12 chromatin model systems in which amajority of the sample contained both one histone octamer per 5S rDNA repeat and one molecule ofhistone H5 per histone octamer. The integrity of the purified 208-12 chromatin has been extensivelycharacterized under low-salt conditions using analytical ultracentrifugation, quantitative agarose gelelectrophoresis, electron cryomicroscopy, and nuclease digestion. Results indicate that histone H5 bindingto 208-12 nucleosomal arrays constrains the entering and exiting linker DNA in a way that producesstructures that are indistinguishable from native chicken erythrocyte chromatin. Folding experimentsperformed in NaCl and MgCl₂ have shown that H5 binding markedly stabilizes both the intermediate andextensively folded states of nucleosomal arrays without fundamentally altering the intrinsic nucleosomalarray folding pathway. These results provide new insight into the mechanism of chromatin folding bydemonstrating for the first time that distinctly different macromolecular determinants are required forformation and stabilization of higher-order chromatin structures.