Characterization of a human cytomegalovirus 1.6-kilobase late mRNA and identification of its putative protein product.
In a previous study (J. Martinez, R. S. Lahijani, and S. C. St. Jeor, J. Virol. 63:233-241, 1989), we identified a late, unspliced 1.6-kb mRNA that maps to the HindIII R fragment of human cytomegalovirus (HCMV) AD169. In the present study, the direction of transcription of this mRNA was determined by Northern (RNA) analysis with strand-specific probes. Primer extension was used to precisely map the 5' end of the mRNA. An open reading frame (ORF) designated ORF 2-1, located 176 nucleotides downstream from the cap site of the 1.6-kb mRNA, was identified. A synthetic peptide was made representing a hydrophilic region in the amino terminus of ORF 2-1. Immunoprecipitation and Western immunoblot analysis of infected HEL cell lysates, using affinity-purified antibody to the peptide (anti-P2-1), detected a viral protein with an apparent molecular mass of 58 kDa late in infection. Further support for the presence of this protein in infected-cell lysates was obtained by an enzyme-linked immunosorbent assay. Expression of viral antigens in intact infected HEL cells was assessed by immunofluorescence. General cytoplasmic staining was observed at 62 h postinfection, in contrast to a localized staining observed in the nuclear and perinuclear region at 96 h postinfection.
Lahijani
R S
RS
Department of Microbiology, University of Nevada, Reno 89557-0046.
Otteson
E W
EW
Adlish
J D
JD
St Jeor
S C
SC
D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D002460:Cell Line; D003587:Cytomegalovirus / Q000235:genetics*; D004279:DNA, Viral / Q000235:genetics; D005814:Genes, Viral; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D011487:Protein Conformation; D012333:RNA, Messenger / Q000235:genetics* / Q000302:isolation & purification; D015183:Restriction Mapping; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000235:genetics*
D004279:DNA, Viral; D012333:RNA, Messenger; D014764:Viral Proteins