A method for genetically installing site-specific acetylation in recombinant histones defines the effects of H3 K56 acetylation.
Lysine acetylation of histones defines the epigenetic status of human embryonic stem cells and orchestrates DNA replication, chromosome condensation, transcription, telomeric silencing, and DNA repair. A detailed mechanistic explanation of these phenomena is impeded by the limited availability of homogeneously acetylated histones. We report a general method for the production of homogeneously and site-specifically acetylated recombinant histones by genetically encoding acetyl-lysine. We reconstitute histone octamers, nucleosomes, and nucleosomal arrays bearing defined acetylated lysine residues. With these designer nucleosomes, we demonstrate that, in contrast to the prevailing dogma, acetylation of H3 K56 does not directly affect the compaction of chromatin and has modest effects on remodeling by SWI/SNF and RSC. Single-molecule FRET experiments reveal that H3 K56 acetylation increases DNA breathing 7-fold. Our results provide a molecular and mechanistic underpinning for cellular phenomena that have been linked with K56 acetylation.
Neumann
Heinz
H
Medical Research Council Laboratory of Molecular Biology, Hills Road, Cambridge CB2 0QH, England, UK.
Hancock
Susan M
SM
Buning
Ruth
R
Routh
Andrew
A
Chapman
Lynda
L
Somers
Joanna
J
Owen-Hughes
Tom
T
van Noort
John
J
Rhodes
Daniela
D
Chin
Jason W
JW
D000107:Acetylation; D019943:Amino Acid Substitution / Q000502:physiology; D000604:Amino Acyl-tRNA Synthetases / Q000235:genetics; D042002:Chromatin Assembly and Disassembly / Q000187:drug effects / Q000502:physiology; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D006657:Histones / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008239:Lysine / Q000031:analogs & derivatives / Q000235:genetics / Q000378:metabolism*; D009707:Nucleosomes / Q000187:drug effects / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D012965:Sodium Chloride / Q000494:pharmacology; D014157:Transcription Factors / Q000378:metabolism
D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; D006657:Histones; D009707:Nucleosomes; C103337:RSC complex, S cerevisiae; D011994:Recombinant Proteins; C445179:SWI-SNF-B chromatin-remodeling complex; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; C002315:N(alpha)-acetyllysine; D012965:Sodium Chloride; D000604:Amino Acyl-tRNA Synthetases; D008239:Lysine