title: Bovine herpesvirus 5 BICP0 complements the bovine herpesvirus 1 homolog.
abstract: Bovine herpesvirus 1 (BoHV-1) and BoHV-5 are closely related (82% amino acid identity) but differ strongly in neuropathogenesis. The immediate-early gene for BICP0 is less conserved (70% amino acid identity) and may contribute to a dissimilar phenotype. A peculiar difference is a guanosine hexamer in the BICP0-1 gene which aligns with only five guanosines in the BICP0-5 gene and therefore results in a frameshift in the latter open reading frame. Thus, the C-terminal amino acid sequence (residues 643-676 of BICP0-1 vs. 655-720 of BICP0-5) is completely different. We introduced the BICP0-5 frameshift into the BoHV-1 genome cloned as a bacterial artificial chromosome (BoHV-1 BAC) using the Red recombination system with galK selection and counterselection. Transfection of MDBK cells with the resulting BAC produced recombinant virus that replicated like wild type BoHV-1 in vitro. Attempts to exchange the entire BICP0-1 gene by the BoHV-5 homolog using the same approach failed repeatedly. Therefore, we cotransfected purified BICP0(-)/galK(+)-BoHV-1 BAC DNA with a recombination plasmid coding for BICP0-5 with or without a HA tag into MDBK cells. BoHV-1 recombinants expressing the respective proteins were characterized. In vitro, all recombinants grew to similar titers as the parental viruses, which demonstrates that BICP0-5 compensates for the growth defect of BICP0(-)/galK(+)-BoHV-1 and functionally complements BICP0-1 of BoHV-1. We conclude that BICP0 may be suitable to positively select BoHV-1 recombinants with deletions or insertions of additional genes of interest.
authors.affiliation: Faculty of Veterinary Medicine, Institute of Virology, University of Zurich, Winterthurerstrasse 266A, CH-8057 Zurich, Switzerland.
authors.forename: Florian
authors.initials: F
authors.lastname: Steiner
authors.forename: Anita
authors.initials: A
authors.lastname: Zumsteg
authors.forename: Bernd
authors.initials: B
authors.lastname: Vogt
authors.forename: Mathias
authors.initials: M
authors.lastname: Ackermann
authors.forename: Martin
authors.initials: M
authors.lastname: Schwyzer
mesh_terms: D000818:Animals; D002417:Cattle; D002460:Cell Line; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics; D016368:Frameshift Mutation; D005816:Genetic Complementation Test; D007242:Herpesvirus 1, Bovine / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D025881:Herpesvirus 5, Bovine / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D017874:Immediate-Early Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011995:Recombination, Genetic; D015534:Trans-Activators / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: C551101:BICP0 protein, Bovine herpesvirus 5; D017874:Immediate-Early Proteins; D015534:Trans-Activators; D014764:Viral Proteins; D044767:Ubiquitin-Protein Ligases; C086729:bICP0 protein, Bovine herpesvirus 1