Preferential localization of DNA damage induced by depurination and bleomycin in a plasmid containing a scaffold-associated region.
Recent evidence suggests that DNA damage of various origins is not randomly distributed in the genome but appears to be clustered in unidentified hypersensitive regions of the chromatin. A model was proposed that stipulates that unpaired DNA stretches, such as those found in scaffold- (or matrix)-associated regions (SARs) under torsional strain, are candidate regions of hypersensitivity to DNA damage in vivo. In this study, we assessed in vitro the relative susceptibility of supercoiled plasmids containing a SAR or chromatin loop DNA segment to DNA damage induced by acid-catalyzed depurination or FeIII-bleomycin. Single-strand specific S1 nuclease was used in combination with 3'-end-labeling to detect single-strand breaks or gaps, after cleavage of abasic sites or removal of 3'-phosphoglycolates by Escherichia coli endonuclease IV. The optimal conditions of DNA cleavage specificity by S1 nuclease were determined. Using these conditions, the DNA cleavage patterns obtained showed (i) a preferential localization of S1 hypersensitive sites in the SAR DNA as compared with plasmid or chromatin loop DNA and (ii) a strikingly similar localization of DNA damage with the two clastogenic treatments.
Legault
J
J
Unit of Health and Environment, CHUL Research Center and Laval University, Ste-Foy, QC, Canada.
Tremblay
A
A
Mirault
M E
ME
D001482:Base Composition; D001761:Bleomycin / Q000031:analogs & derivatives / Q000494:pharmacology*; D002843:Chromatin / Q000235:genetics; D004249:DNA Damage / Q000187:drug effects*; D004278:DNA, Superhelical / Q000737:chemistry / Q000187:drug effects / Q000502:physiology; D003850:Deoxyribonuclease I; D015530:Nuclear Matrix / Q000235:genetics*; D009690:Nucleic Acid Conformation / Q000187:drug effects; D010957:Plasmids / Q000187:drug effects / Q000378:metabolism*; D011687:Purines / Q000378:metabolism*; D015719:Single-Strand Specific DNA and RNA Endonucleases
D002843:Chromatin; D004278:DNA, Superhelical; D011687:Purines; C023503:iron bleomycin; D001761:Bleomycin; D003850:Deoxyribonuclease I; D015719:Single-Strand Specific DNA and RNA Endonucleases