title: Minor groove contacts are essential for an interaction of the human cytomegalovirus IE2 protein with its DNA target.
abstract: The 86 kDa immediate early-2 protein (IE2, IE86) of human cytomegalovirus (HCMV) is a multifunctional polypeptide that can regulate gene expression both positively and negatively. In particular, it represses its own mRNA synthesis by binding directly to a sequence element, termed cis repression signal (CRS), that is located between the TATA box and the transcriptional start site of the major IE enhancer/promoter of HCMV. Here, we provide evidence that IE86, unlike most sequence-specific DNA-binding proteins, interacts primarily within the minor groove of the DNA helix. This was shown by hydroxyl radical and methylation interference assays. In addition, binding studies with inosine-substituted oligonucleotides which have an altered major groove morphology without changing the surface of the minor groove, confirmed the results obtained in interference analyses. This establishes IE86 as a member of a small group of DNA binding proteins that interact with A - T rich sequences within the minor groove and which also includes the TATA-box binding protein TBP. Remarkably, IE86 and TBP are able to bind simultaneously in an immediate vicinity at the major IE enhancer/promoter of HCMV. As minor groove binding proteins are known to bend DNA heavily this could contribute to the observed negative regulation of transcription by IE86.
authors.affiliation: Institut für Klinische und Molekulare Virologie der Universität Erlangen-Nürnberg, Germany.
authors.forename: D
authors.initials: D
authors.lastname: Lang
authors.forename: T
authors.initials: T
authors.lastname: Stamminger
mesh_terms: D000956:Antigens, Viral / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D003300:Copper; D003587:Cytomegalovirus / Q000235:genetics; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism; D003850:Deoxyribonuclease I; D005786:Gene Expression Regulation; D017665:Hydroxyl Radical / Q000378:metabolism; D017874:Immediate-Early Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008745:Methylation; D010618:Phenanthrolines; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins; D012091:Repetitive Sequences, Nucleic Acid; D016385:TATA Box
chemical_list: D000956:Antigens, Viral; D004279:DNA, Viral; D017874:Immediate-Early Proteins; D010618:Phenanthrolines; D011994:Recombinant Proteins; C044582:immediate-early proteins, cytomegalovirus; D017665:Hydroxyl Radical; D003300:Copper; D004247:DNA; D003850:Deoxyribonuclease I; C025205:1,10-phenanthroline