title: Differential role of base excision repair proteins in mediating cisplatin cytotoxicity.
abstract: Interstrand crosslinks (ICLs) are covalent lesions formed by cisplatin. The mechanism for the processing and removal of ICLs by DNA repair proteins involves nucleotide excision repair (NER), homologous recombination (HR) and fanconi anemia (FA) pathways. In this report, we monitored the processing of a flanking uracil adjacent to a cisplatin ICL by the proteins involved in the base excision repair (BER) pathway. Using a combination of extracts, purified proteins, inhibitors, functional assays and cell culture studies, we determined the specific BER proteins required for processing a DNA substrate with a uracil adjacent to a cisplatin ICL. Uracil DNA glycosylase (UNG) is the primary glycosylase responsible for the removal of uracils adjacent to cisplatin ICLs, whereas other uracil glycosylases can process uracils in the context of undamaged DNA. Repair of the uracil adjacent to cisplatin ICLs proceeds through the classical BER pathway, highlighting the importance of specific proteins in this redundant pathway. Removal of uracil is followed by the generation of an abasic site and subsequent cleavage by AP endonuclease 1 (APE1). Inhibition of either the repair or redox domain of APE1 gives rise to cisplatin resistance. Inhibition of the lyase domain of Polymerase β (Polβ) does not influence cisplatin cytotoxicity. In addition, lack of XRCC1 leads to increased DNA damage and results in increased cisplatin cytotoxicity. Our results indicate that BER activation at cisplatin ICLs influences crosslink repair and modulates cisplatin cytotoxicity via specific UNG, APE1 and Polβ polymerase functions.
authors.affiliation: Department of Oncology, Karmanos Cancer Institute, Wayne State University, Detroit, MI 48201, United States.
authors.forename: Akshada
authors.initials: A
authors.lastname: Sawant
authors.affiliation: Department of Oncology, Karmanos Cancer Institute, Wayne State University, Detroit, MI 48201, United States.
authors.forename: Ashley M
authors.initials: AM
authors.lastname: Floyd
authors.affiliation: Department of Biochemistry and Cancer Biology, University of Toledo - Health Science Campus, Toledo, OH 43614, United States.
authors.forename: Mohan
authors.initials: M
authors.lastname: Dangeti
authors.affiliation: Department of Oncology, Karmanos Cancer Institute, Wayne State University, Detroit, MI 48201, United States.
authors.forename: Wen
authors.initials: W
authors.lastname: Lei
authors.affiliation: Department of Oncologic Sciences, Molecular & Metabolic Oncology Program, Mitchell Cancer Institute, University of South Alabama,1660 Springhill Avenue, Mobile, AL 36604, United States.
authors.forename: Robert W
authors.initials: RW
authors.lastname: Sobol
authors.affiliation: Department of Oncology, Karmanos Cancer Institute, Wayne State University, Detroit, MI 48201, United States. Electronic address: patricks@karmanos.org.
authors.forename: Steve M
authors.initials: SM
authors.lastname: Patrick
mesh_terms: D000818:Animals; D002945:Cisplatin / Q000633:toxicity*; D018736:DNA Adducts; D019951:DNA Polymerase beta / Q000378:metabolism; D004260:DNA Repair; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D006801:Humans; D051379:Mice; D013379:Substrate Specificity; D014498:Uracil / Q000378:metabolism*; D051981:Uracil-DNA Glycosidase / Q000378:metabolism*; D000076105:X-ray Repair Cross Complementing Protein 1
chemical_list: D018736:DNA Adducts; D004268:DNA-Binding Proteins; D000076105:X-ray Repair Cross Complementing Protein 1; C000619371:XRCC1 protein, human; C000619352:Xrcc1 protein, mouse; C057270:cisplatin-DNA adduct; D014498:Uracil; D019951:DNA Polymerase beta; D051981:Uracil-DNA Glycosidase; C085108:APEX1 protein, human; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D002945:Cisplatin
keywords: BER; Cisplatin; ICL; Resistance