title: Characterization of an In Vivo Z-DNA Detection Probe Based on a Cell Nucleus Accumulating Intrabody.
abstract: Left-handed Z-DNA is a physiologically unstable DNA conformation, and its existence in vivo can be attributed to localized torsional distress. Despite evidence for the existence of Z-DNA in vivo, its precise role in the control of gene expression is not fully understood. Here, an in vivo probe based on an anti-Z-DNA intrabody is proposed for native Z-DNA detection. The probe was used for chromatin immunoprecipitation of potential Z-DNA-forming sequences in the human genome. One of the isolated putative Z-DNA-forming sequences was cloned upstream of a reporter gene expression cassette under control of the CMV promoter. The reporter gene encoded an antibody fragment fused to GFP. Transient co-transfection of this vector along with the Z-probe coding vector improved reporter gene expression. This improvement was demonstrated by measuring reporter gene mRNA and protein levels and the amount of fluorescence in co-transfected CHO-K1 cells. These results suggest that the presence of the anti-Z-DNA intrabody can interfere with a Z-DNA-containing reporter gene expression. Therefore, this in vivo probe for the detection of Z-DNA could be used for global correlation of Z-DNA-forming sequences and gene expression regulation.
authors.affiliation: Department of Cell Biology, Institute of Biological Sciences, University of Brasilia, Brasilia, Brazil.
authors.forename: Galina
authors.initials: G
authors.lastname: Gulis
authors.affiliation: Molecular Pathology Graduation Program, Medicine Faculty, University of Brasilia, Brasilia, Brazil.|Faculty of Ceilandia, University of Brasilia, Brasilia, Brazil.
authors.forename: Izabel Cristina Rodrigues
authors.initials: IC
authors.lastname: Silva
authors.affiliation: Molecular Pathology Graduation Program, Medicine Faculty, University of Brasilia, Brasilia, Brazil.
authors.forename: Herdson Renney
authors.initials: HR
authors.lastname: Sousa
authors.affiliation: Molecular Pathology Graduation Program, Medicine Faculty, University of Brasilia, Brasilia, Brazil.
authors.forename: Isabel Garcia
authors.initials: IG
authors.lastname: Sousa
authors.affiliation: Molecular Biology Graduation Program, Institute of Biological Sciences, University of Brasilia, Brasilia, Brazil.
authors.forename: Maryani Andressa Gomes
authors.initials: MA
authors.lastname: Bezerra
authors.affiliation: Molecular Biology Graduation Program, Institute of Biological Sciences, University of Brasilia, Brasilia, Brazil.
authors.forename: Luana Salgado
authors.initials: LS
authors.lastname: Quilici
authors.affiliation: Department of Cell Biology, Institute of Biological Sciences, University of Brasilia, Brasilia, Brazil.
authors.forename: Andrea Queiroz
authors.initials: AQ
authors.lastname: Maranhao
authors.affiliation: Department of Cell Biology, Institute of Biological Sciences, University of Brasilia, Brasilia, Brazil. brigido@unb.br.
authors.forename: Marcelo Macedo
authors.initials: MM
authors.lastname: Brigido
mesh_terms: D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D002467:Cell Nucleus / Q000235:genetics*; D047369:Chromatin Immunoprecipitation; D006224:Cricetinae; D003412:Cricetulus; D015342:DNA Probes / Q000378:metabolism*; D043542:DNA, Z-Form / Q000302:isolation & purification* / Q000378:metabolism; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006801:Humans; D061986:MCF-7 Cells
chemical_list: D015342:DNA Probes; D043542:DNA, Z-Form
keywords: Anti-Z-DNA; Chromatin immunoprecipitation; Co-transfection; Intrabody; Z-DNA; Z-probe