Structural and functional analysis of chromatin assembled from defined histones.
In this review we describe how the extract-mediated chromatin assembly system derived from preblastoderm Drosophila embryos can be modified to assemble chromatin from defined histones. This approach combines the advantages of assembling (i) chromatin templates from homogeneous histones with (ii) an assembly system that generates chromatin with physiological nucleosome spacing and density and that contains the biological complexity of in vivo chromatin. We have used this technique to assemble nonacetylated and hyperacetylated histones into chromatin (K. P. Nightingale, R. Wellinger, J. Sogo, and P. B. Becker, 1998, EMBO J. 17, 2865-2876; W. A. Krajewski and P. B. Becker, 1998, Proc. Natl. Acad. Sci. USA 95, 1540-1545), and use this as an example to detail the structural and transcriptional assays used to compare and characterize these chromatin templates. The application of this procedure to assemble chromatin from recombinant histones should facilitate a wide variety of studies on the role(s) of histone mutants and variants.
Nightingale
K P
KP
Gene Expression Programme, European Molecular Biology Laboratory, Heidelberg, Germany.
Becker
P B
PB
D000818:Animals; D001671:Biochemistry / Q000379:methods*; D001756:Blastoderm / Q000737:chemistry; D002457:Cell Extracts; D002843:Chromatin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism; D004330:Drosophila / Q000235:genetics; D005821:Genetic Techniques; D006657:Histones / Q000302:isolation & purification* / Q000378:metabolism; D013698:Templates, Genetic; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic
D002457:Cell Extracts; D002843:Chromatin; D006657:Histones; D014157:Transcription Factors; D003850:Deoxyribonuclease I