Transcriptional and translational analyses of the UL2 gene of equine herpesvirus 1: a homolog of UL55 of herpes simplex virus type 1 that is maintained in the genome of defective interfering particles

• Published in: Journal of Virology Vol. 67; no. 4; pp. 2255 - 2265
• Main Authors: Harty, R.N. (Mount Sinai School of Medicine, New York, NY), Holden, V.R, O'Callaghan, D.J
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for Microbiology 01.04.1993
• Subjects: Animals; Antigens, Viral - genetics; ARN MENSAJERO; ARN MESSAGER; Base Sequence; Biological and medical sciences; Cells, Cultured; Cloning, Molecular; Defective Viruses - genetics; EXPRESION GENICA; EXPRESSION DES GENES; Fundamental and applied biological sciences. Psychology; GENE; Gene Expression Regulation, Viral; GENES; Genes, Viral; Genetics; GENOMAS; GENOME; Herpesvirus 1, Equid - genetics; HERPESVIRUS EQUIN; HERPESVIRUS EQUINO; In Vitro Techniques; Mice; Microbiology; Molecular Sequence Data; Oligodeoxyribonucleotides - chemistry; Protein Biosynthesis; PROTEINAS; PROTEINE; Recombinant Fusion Proteins - immunology; RNA, Messenger - genetics; Transcription, Genetic; Viral Interference; Viral Proteins - genetics; Viral Structural Proteins - genetics; Virology; VIRUS DE LOS ANIMALES; VIRUS DES ANIMAUX; Virus; Proteins; Equine herpesvirus 1; Gene; Herpesvirus hominis 1; Herpesviridae; Alphaherpesvirinae; Interference; Homology; Gene expression; Defectivity
• ISSN: 0022-538X; 1098-5514
• DOI: 10.1128/JVI.67.4.2255-2265.1993

Defective interfering particles (DIPs) of equine herpesvirus 1 (EHV-1; Kentucky A strain) mediate persistent infection. DNA sequences at the L terminus, which contain the UL2 gene (homolog of UL55 of herpes simplex virus type 1 and open reading frame 3 of varicella-zoster virus) of standard EHV-1, have been shown to be highly conserved in all clones of the EHV-1 DIP genome. The UL2 mRNA was characterized by S1 nuclease analyses, which mapped the 5' and 3' termini of the 0.9-kb early UL2 mRNA to approximately 26 and 16 nucleotides downstream of a TTTAAA box and polyadenylation signal, respectively. The UL2 open reading frame, present within both the EHV-1 standard and DIP genomes, was inserted into the transcription expression vector pGEM-3Z to yield constructs pGEML2 and pDIL2, respectively. After in vitro transcription and translation, both constructs yielded a comigrating 23-kDa protein, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Polyclonal antiserum was raised against the UL2 protein by injecting rabbits with a TrpE/UL2 fusion protein expressed from plasmid pATH23L2 in Escherichia coli. The UL2-specific antiserum reacted in Western immunoblot and immunoprecipitation analyses with a 23-kDa polypeptide synthesized in cells infected with standard EHV-1 or DIP-enriched virus. These data also indicated that the UL2 polypeptide was more abundant in DIP-infected cells than in standard EHV-1-infected cells. Results from time course and pulse-chase analyses suggested that the UL2 polypeptide has a rapid turnover rate in DIP-infected cells