title: Efficient processing of TFO-directed psoralen DNA interstrand crosslinks by the UvrABC nuclease.
abstract: Photoreactive psoralens can form interstrand crosslinks (ICLs) in double-stranded DNA. In eubacteria, the endonuclease UvrABC plays a key role in processing psoralen ICLs. Psoralen-modified triplex-forming oligonucleotides (TFOs) can be used to direct ICLs to specific genomic sites. Previous studies of pyrimidine-rich methoxypsoralen-modified TFOs indicated that the TFO inhibits cleavage by UvrABC. Because different chemistries may alter the processing of TFO-directed ICLs, we investigated the effect of another type of triplex formed by purine-rich TFOs on the processing of 4'-(hydroxymethyl)-4,5',8-trimethylpsoralen (HMT) ICLs by the UvrABC nuclease. Using an HMT-modified TFO to direct ICLs to a specific site, we found that UvrABC made incisions on the purine-rich strand of the duplex approximately 3 bases from the 3'-side and approximately 9 bases from the 5'-side of the ICL, within the TFO-binding region. In contrast to previous reports, the UvrABC nuclease cleaved the TFO-directed psoralen ICL with a greater efficiency than that of the psoralen ICL alone. Furthermore, the TFO was dissociated from its duplex binding site by UvrA and UvrB. As mutagenesis by TFO-directed ICLs requires nucleotide excision repair, the efficient processing of these lesions supports the use of triplex technology to direct DNA damage for genome modification.
authors.affiliation: Department of Carcinogenesis, Science Park-Research Division, University of Texas MD Anderson Cancer Center, Smithville, TX, USA.
authors.forename: Laura A
authors.initials: LA
authors.lastname: Christensen
authors.forename: Hong
authors.initials: H
authors.lastname: Wang
authors.forename: Bennett
authors.initials: B
authors.lastname: Van Houten
authors.forename: Karen M
authors.initials: KM
authors.lastname: Vasquez
mesh_terms: D003432:Cross-Linking Reagents / Q000737:chemistry*; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D009841:Oligonucleotides / Q000737:chemistry / Q000378:metabolism*; D014307:Trioxsalen / Q000031:analogs & derivatives* / Q000737:chemistry
chemical_list: D003432:Cross-Linking Reagents; D029968:Escherichia coli Proteins; D009841:Oligonucleotides; C051802:triplex DNA; C016507:hydroxymethyltrioxsalen; D004247:DNA; D004706:Endodeoxyribonucleases; C036385:endodeoxyribonuclease uvrABC; D014307:Trioxsalen