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Effect of nucleosome structure on DNA interstrand cross-linking reactions
J T Millard1, R J Spencer, P B Hopkins
1Department of Chemistry, Colby College, Waterville, Maine 04901, USA. jtmillar@colby.edu
Biochemistry
|May 16, 1998
Summary
Nitrogen mustards and mitomycin C cross-link DNA. While nitrogen mustards showed similar DNA cross-linking in nucleosomes and free DNA, mitomycin C cross-linking was inhibited by nucleosome structure.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Antitumor agents like nitrogen mustards and mitomycin C are crucial in cancer therapy.
- These agents function by forming interstrand cross-links in DNA, leading to cytotoxicity.
- Understanding their interaction with DNA, especially within the context of chromatin, is vital for drug development.
Purpose of the Study:
- To investigate the cellular mechanism of cytotoxic antitumor agents.
- To examine the DNA cross-linking activity of nitrogen mustards and mitomycin C on nucleosomal DNA.
- To determine how nucleosome structure influences the sites and efficiency of DNA cross-linking.
Main Methods:
- Cross-linking assays were performed on a nucleosomal core particle reconstituted on a Xenopus borealis 5S RNA gene fragment.
- The DNA cross-linking patterns of mechlorethamine, chlorambucil, melphalan, and mitomycin C were analyzed.
- Comparison of cross-linking sites and intensities between nucleosomal DNA and free DNA.
Main Results:
- Nitrogen mustards (mechlorethamine, chlorambucil, melphalan) exhibited similar monoalkylation and interstrand cross-linking sites in both nucleosomal and free DNA.
- Minor variations (two- to three-fold) in cross-linking intensity were observed but did not correlate with DNA positioning within the nucleosome.
- Mitomycin C cross-linking was significantly inhibited (five- to ten-fold) at the nucleosomal dyad, with reduced inhibition towards the ends.
- Nucleosome structure, specifically inward-facing sites, hindered mitomycin C cross-linking, indicating a role for rotational positioning.
- Unlike hydroxyl radical cleavage, none of the tested agents showed a 10-base pair periodicity in nucleosomal DNA.
Conclusions:
- Nucleosome structure differentially affects DNA cross-linking by antitumor agents.
- Nitrogen mustards are less affected by nucleosome incorporation compared to mitomycin C.
- Mitomycin C's efficacy may be modulated by its accessibility to DNA within the nucleosome, suggesting potential for targeted therapies.
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