title: Identification of low-molecular-weight vitellogenin 1 (Vg1)-like proteins as nucleotide excision repair (NER) factors in developing zebrafish (Danio rerio) using a transcription-based DNA repair assay.
abstract: Nucleotide excision repair (NER) removes helix-distorting DNA lesions such as UV-induced pyrimidine dimers and cisplatin-induced strand crosslinking. Our earlier studies have identified low-molecular-weight proteins homologous to the 150-kDa vitellogenin 1 (Vg1) as UV-damaged DNA-binding factors expressed in developing zebrafish (Danio rerio). This present study explored if Vg1-like proteins also participated in NER in zebrafish. Immunoblot analysis of affinity-captured 12 h post-fertilization (hpf) zebrafish extract proteins showed a transient binding of a 30-kDa Vg1-like polypeptide to UV-damaged DNA. A transcription-based in vitro repair assay revealed a significant up-regulation of UVC or cisplatin-suppressed transcriptional activity of a marker cDNA driven by a SP6 RNA polymerase-regulated promotor after incubating the damaged plasmid with the extracts of 12 hpf embryos or 96 hpf larvae. The up-regulation of UV or cisplatin-suppressed transcription was abolished in the presence of a monoclonal anti-zebrafish Vg1 antibody. The differential sensitivity of UV-induced repair in 12 and 96 hpf zebrafish extracts to exogenous ATP suggested a development-dependent expression of Vg1-like NER factors. A T4endonuclease V digestion assay showed no inhibition of the anti-Vg1 antibody on the excision of UV-induced cyclobutane pyrimidine dimers. Our results identified the participation of Vg1-like factors in NER in developing zebrafish, and these factors may function at post-incison steps of NER.
authors.affiliation: Department of Internal Medicine, Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Keelung, 204, Taiwan.|Department of Bioscience and Biotechnology and Center of Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan.
authors.forename: Yung-Chi
authors.initials: YC
authors.lastname: Shen
authors.affiliation: Department of Bioscience and Biotechnology and Center of Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan. toddhsu@mail.ntou.edu.tw.
authors.forename: Todd
authors.initials: T
authors.lastname: Hsu
authors.affiliation: Department of Bioscience and Biotechnology and Center of Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan.
authors.forename: Li-Bin
authors.initials: LB
authors.lastname: Ling
authors.affiliation: Department of Bioscience and Biotechnology and Center of Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan.
authors.forename: Wen-Chian
authors.initials: WC
authors.lastname: You
authors.affiliation: Department of Bioscience and Biotechnology and Center of Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan.
authors.forename: Chia-Wei
authors.initials: CW
authors.lastname: Liu
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001681:Biological Assay; D002945:Cisplatin / Q000494:pharmacology; D004249:DNA Damage; D004260:DNA Repair; D004625:Embryo, Nonmammalian / Q000378:metabolism; D008970:Molecular Weight; D014158:Transcription, Genetic / Q000187:drug effects; D014466:Ultraviolet Rays; D014819:Vitellogenins / Q000235:genetics* / Q000378:metabolism; D015027:Zebrafish / Q000235:genetics* / Q000378:metabolism; D029961:Zebrafish Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D014819:Vitellogenins; D029961:Zebrafish Proteins; C486450:vtg1 protein, zebrafish; D000255:Adenosine Triphosphate; D002945:Cisplatin
keywords: Cisplatin; Nucleotide excision repair; Protein; UV; Vitellogenin; Zebrafish