title: Characterization and localization of the equine herpesvirus 1 major DNA binding protein.
abstract: In previous studies of equine herpesvirus 1 (EHV-1) gene regulation, we observed an abundant early infected cell polypeptide (ICP), designated ICP130, which appeared in reduced amounts in cells infected with defective interfering particle-rich EHV-1 stocks compared to standard EHV-1-infected cells. To characterize this ICP further, a monoclonal antibody (MAb) was developed to EHV-1 ICP130 and used to (1) affinity purify ICP130, (2) examine ICP130's ability to bind DNA, and (3) define the synthesis and intracellular localization of ICP130 during productive EHV-1 infections. Although anti-ICP130 MAbs did not crossreact with any HSV-1 protein in immunoblots, a polyclonal antiserum against HSV-2 major DNA binding protein (ICSP11,12) did react with purified EHV-1 ICP130. DNA band shift assays indicated that (1) the mobility of shifted bands representing DNA/EHV-1-infected cell protein complexes was further decreased by the addition of either anti-ICP130 MAbs or anti-ICSP11,12, but not by the addition of irrelevant MAbs, (2) the ability of ICP130 to complex with DNA was not sequence dependent, (3) ICP130 associated with both single- and double-stranded oligomers, and (4) similar supershifted patterns were produced using affinity-purified ICP130 and anti-ICP130 MAbs. During productive infection, ICP130 initially localized rapidly and exclusively to the infected cell's nucleus in a generalized, fine granular pattern. Over the course of infection, this pattern typically progressed to include several large, intensely reactive intranuclear granules, and by 6 hr p.i. some cytoplasmic reactivity also was visible. In < 5% of the cells, a dense, fibrillar network surrounding the nucleus was observed instead. The progressive changes in nuclear localization depended upon the onset of viral DNA replication, and once the late pattern was established, ongoing DNA synthesis was required to maintain it. The results indicate that ICP130 is the previously reported EHV-1 counterpart of the HSV major DNA-binding protein and is similar, but not identical, in many aspects.
authors.affiliation: Department of Oral Biology, Medical College of Georgia, Augusta 30912-1126.
authors.forename: J B
authors.initials: JB
authors.lastname: Lewis
authors.forename: Y G
authors.initials: YG
authors.lastname: Thompson
authors.forename: A C
authors.initials: AC
authors.lastname: Jenkins
authors.forename: G B
authors.initials: GB
authors.lastname: Caughman
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D000956:Antigens, Viral / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D006224:Cricetinae; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D004861:Herpesvirus 1, Equid / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D018259:Herpesvirus 1, Human / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D011817:Rabbits; D013347:Subcellular Fractions / Q000378:metabolism; D014709:Vero Cells
chemical_list: D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D004279:DNA, Viral; D004268:DNA-Binding Proteins