Accommodation and repair of a UV photoproduct in DNA at different rotational settings on the nucleosome surface.
Cyclobutane-thymine dimers (CTDs), the most common DNA lesion induced by UV radiation, cause 30 degrees bending and 9 degrees unwinding of the DNA helix. We prepared site-specific CTDs within a short sequence bracketed by strong nucleosome-positioning sequences. The rotational setting of CTDs over one turn of the helix near the dyad center on the histone surface was analyzed by hydroxyl radical footprinting. Surprisingly, the position of CTDs over one turn of the helix does not affect the rotational setting of DNA on the nucleosome surface. Gel-shift analysis indicates that one CTD destabilizes histone-DNA interactions by 0.6 or 1.1 kJ/mol when facing away or toward the histone surface, respectively. Thus, 0.5 kJ/mol energy penalty for a buried CTD is not enough to change the rotational setting of sequences with strong rotational preference. The effect of rotational setting on CTD removal by nucleotide excision repair (NER) was examined using Xenopus oocyte nuclear extracts. The NER rates are only 2-3 times lower in nucleosomes and change by only 1.5-fold when CTDs face away or toward the histone surface. Therefore, in Xenopus nuclear extracts, the rotational orientation of CTDs on nucleosomes has surprisingly little effect on rates of repair. These results indicate that nucleosome dynamics and/or chromatin remodeling may facilitate NER in gaining access to DNA damage in nucleosomes.
Svedruzić
Zeljko M
ZM
Department of Biochemistry and Biophysics, School of Molecular Biosciences, Washington State University, Pullman, Washington 99164-4660, USA.
Wang
Chenbo
C
Kosmoski
Joseph V
JV
Smerdon
Michael J
MJ
D000818:Animals; D001667:Binding, Competitive; D002467:Cell Nucleus / Q000378:metabolism; D002843:Chromatin / Q000737:chemistry; D004247:DNA / Q000737:chemistry*; D004249:DNA Damage; D004260:DNA Repair; D019281:Dimerization; D006657:Histones / Q000737:chemistry; D017665:Hydroxyl Radical; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D009707:Nucleosomes / Q000737:chemistry* / Q000378:metabolism; D009865:Oocytes / Q000378:metabolism; D011487:Protein Conformation; D011740:Pyrimidine Dimers / Q000737:chemistry; D013816:Thermodynamics; D013997:Time Factors; D014466:Ultraviolet Rays; D014982:Xenopus laevis
D002843:Chromatin; D006657:Histones; D009707:Nucleosomes; D011740:Pyrimidine Dimers; D017665:Hydroxyl Radical; D004247:DNA