title: Cockayne Syndrome group B protein stimulates NEIL2 DNA glycosylase activity.
abstract: Cockayne Syndrome is a segmental premature aging syndrome, which can be caused by loss of function of the CSB protein. CSB is essential for genome maintenance and has numerous interaction partners with established roles in different DNA repair pathways including transcription coupled nucleotide excision repair and base excision repair. Here, we describe a new interaction partner for CSB, the DNA glycosylase NEIL2. Using both cell extracts and recombinant proteins, CSB and NEIL2 were found to physically interact independently of DNA. We further found that CSB is able to stimulate NEIL2 glycosylase activity on a 5-hydroxyl uracil lesion in a DNA bubble structure substrate in vitro. A novel 4,6-diamino-5-formamidopyrimidine (FapyA) specific incision activity of NEIL2 was also stimulated by CSB. To further elucidate the biological role of the interaction, immunofluorescence studies were performed, showing an increase in cytoplasmic CSB and NEIL2 co-localization after oxidative stress. Additionally, stalling of the progression of the transcription bubble with α-amanitin resulted in increased co-localization of CSB and NEIL2. Finally, CSB knockdown resulted in reduced incision of 8-hydroxyguanine in a DNA bubble structure using whole cell extracts. Taken together, our data supports a biological role for CSB and NEIL2 in transcription associated base excision repair.
authors.affiliation: Danish Center for Molecular Gerontology and Danish Aging Research Center, Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark.
authors.forename: Maria D
authors.initials: MD
authors.lastname: Aamann
authors.affiliation: Danish Center for Molecular Gerontology and Danish Aging Research Center, Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark.
authors.forename: Christina
authors.initials: C
authors.lastname: Hvitby
authors.affiliation: Laboratory of Molecular Gerontology, National Institute on Aging, National Institutes of Health, Baltimore, MD, USA.
authors.forename: Venkateswarlu
authors.initials: V
authors.lastname: Popuri
authors.affiliation: Center for Healthy Aging, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Meltem
authors.initials: M
authors.lastname: Muftuoglu
authors.affiliation: Danish Center for Molecular Gerontology and Danish Aging Research Center, Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark.
authors.forename: Lasse
authors.initials: L
authors.lastname: Lemminger
authors.affiliation: Danish Center for Molecular Gerontology and Danish Aging Research Center, Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark.
authors.forename: Cecilie K
authors.initials: CK
authors.lastname: Skeby
authors.affiliation: Danish Center for Molecular Gerontology and Danish Aging Research Center, Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark; Center for Healthy Aging, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Guido
authors.initials: G
authors.lastname: Keijzers
authors.affiliation: Danish Center for Molecular Gerontology and Danish Aging Research Center, Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark; University of Ulsan, Republic of Korea.
authors.forename: Byungchan
authors.initials: B
authors.lastname: Ahn
authors.affiliation: Laboratory for Molecular Biology, Center for Molecular Biology and Neuroscience, Oslo University Hospital, Oslo, Norway.
authors.forename: Magnar
authors.initials: M
authors.lastname: Bjørås
authors.affiliation: Laboratory of Molecular Gerontology, National Institute on Aging, National Institutes of Health, Baltimore, MD, USA.
authors.forename: Vilhelm A
authors.initials: VA
authors.lastname: Bohr
authors.affiliation: Danish Center for Molecular Gerontology and Danish Aging Research Center, Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark. Electronic address: tvs@mb.au.dk.
authors.forename: Tinna
authors.initials: T
authors.lastname: Stevnsner
mesh_terms: D003593:Cytoplasm / Q000378:metabolism; D004247:DNA / Q000737:chemistry; D045647:DNA Glycosylases / Q000378:metabolism*; D004265:DNA Helicases / Q000378:metabolism*; D004260:DNA Repair; D045643:DNA Repair Enzymes / Q000378:metabolism*; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D006147:Guanine / Q000031:analogs & derivatives / Q000737:chemistry; D006367:HeLa Cells; D006801:Humans; D008856:Microscopy, Fluorescence; D018384:Oxidative Stress; D000075223:Poly-ADP-Ribose Binding Proteins; D011994:Recombinant Proteins / Q000378:metabolism; D014158:Transcription, Genetic; D024483:Vitamin K 3 / Q000737:chemistry
chemical_list: D000075223:Poly-ADP-Ribose Binding Proteins; D011994:Recombinant Proteins; C024829:8-hydroxyguanine; D006147:Guanine; D024483:Vitamin K 3; D004247:DNA; D045647:DNA Glycosylases; D004265:DNA Helicases; C078673:ERCC6 protein, human; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; C473642:NEIL2 protein, human; D045643:DNA Repair Enzymes
keywords: Base excision repair; CSB; Cockayne Syndrome; NEIL2; Oxidative damage