UV Damage in DNA Promotes Nucleosome Unwrapping

Biochemistry and Biophysics, School of Molecular Biosciences, Washington State University, Pullman, Washington 99164-7520, USA.
Journal of Biological Chemistry (Impact Factor: 4.6). 08/2010; 285(34):26295-303. DOI: 10.1074/jbc.M110.140087
Source: PubMed

ABSTRACT The association of DNA with histones in chromatin impedes DNA repair enzymes from accessing DNA lesions. Nucleosomes exist in a dynamic equilibrium in which portions of the DNA molecule spontaneously unwrap, transiently exposing buried DNA sites. Thus, nucleosome dynamics in certain regions of chromatin may provide the exposure time and space needed for efficient repair of buried DNA lesions. We have used FRET and restriction enzyme accessibility to study nucleosome dynamics following DNA damage by UV radiation. We find that FRET efficiency is reduced in a dose-dependent manner, showing that the presence of UV photoproducts enhances spontaneous unwrapping of DNA from histones. Furthermore, this UV-induced shift in unwrapping dynamics is associated with increased restriction enzyme accessibility of histone-bound DNA after UV treatment. Surprisingly, the increased unwrapping dynamics is even observed in nucleosome core particles containing a single UV lesion at a specific site. These results highlight the potential for increased "intrinsic exposure" of nucleosome-associated DNA lesions in chromatin to repair proteins.

1 Follower
  • [Show abstract] [Hide abstract]
    ABSTRACT: Sequence-specific DNA-binding modules, N-methylpyrrole (Py)-N-methylimidazole-(Im) polyamides have been recently conjugated with fluorophores, and some of these conjugates could be used for the detection of specific DNA sequences. In this study, we synthesized two Py-Im polyamides 1 and 2, which interact with the 145-bp nucleosome positioning sequence 601. We conjugated the cyanine dye Cy3 or Cy5 with 1 or 2. In the absence of target DNA, the fluorescent conjugate of a Py-Im polyamide had lower fluorescence intensity compared with Cy3 or Cy5 alone. In the presence of either the target DNA or the nucleosome, the fluorescence intensity of the conjugates increased. Furthermore, we observed a Förster resonance energy transfer between the Cy3-Py-Im polyamide and the Cy5-Py-Im polyamide on the nucleosome. These results open up the possibilities that fluorescent conjugates of Py-Im polyamides can be used for characterization of the dynamic interactions within protein-DNA complexes.
    01/2014; 2(3):297. DOI:10.1039/c3bm60202h
  • [Show abstract] [Hide abstract]
    ABSTRACT: Nucleosomes are extremely stable histone-DNA complexes that form the building blocks of chromatin, which accommodates genomic DNA within the nucleus. The dynamic properties of chromatin play essential roles in regulating genomic DNA functions, such as DNA replication, recombination, repair, and transcription. Histones are the protein components of nucleosomes, and their diverse modifications and variants increase the versatility of nucleosome structures and their dynamics in chromatin. Therefore, a technique to evaluate the physical properties of nucleosomes would facilitate functional studies of the various nucleosomes. In this report, we describe a convenient assay for evaluating the thermal stability of nucleosomes in vitro.
    Methods 09/2014; 70(2-3). DOI:10.1016/j.ymeth.2014.08.019 · 3.22 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Nucleotide excision repair (NER) is a key component of the DNA damage response (DDR) and it is essential to safeguard genome integrity against genotoxic insults. The regulation of NER is primarily mediated by protein post-translational modifications (PTMs). The NER machinery removes a wide spectrum of DNA helix distorting lesions, including those induced by solar radiation, through two sub-pathways: global genome nucleotide excision repair (GG-NER) and transcription coupled nucleotide excision repair (TC-NER). Severe clinical consequences associated with inherited NER defects, including premature ageing, neurodegeneration and extreme cancer-susceptibility, underscore the biological relevance of NER. In the last two decades most of the core NER machinery has been elaborately described, shifting attention to molecular mechanisms that either facilitate NER in the context of chromatin or promote the timely and accurate interplay between NER factors and various post-translational modifications. In this review, we summarize and discuss the latest findings in NER. In particular, we focus on emerging factors and novel molecular mechanisms by which NER is regulated.
    Experimental Cell Research 08/2014; 329(1). DOI:10.1016/j.yexcr.2014.08.010 · 3.37 Impact Factor