Arch Virol (1998) 143: 2391–2412
Characterization of a bovine herpesvirus 4(BHV-4) 1.1-kb RNA
and its transactivation by BHV-4 immediate-early 2 gene product
, L. Zhang
, and V. L. van Santen
Department of Pathobiology, Auburn University, Auburn, Alabama, U.S.A.
Accepted June 5, 1998
Summary. We determined the structure of a 1.1-kb cytoplasmic polyadenylated
RNA transcribed from a region of the bovine herpesvirus 4 (BHV-4) genome
not conserved among gammaherpesviruses by sequencing its cDNA, by S1
nuclease analysis, and by primer extension analysis. We found that the RNA
consists of a short, approximately 193-nucleotide (nt), 5
exon spliced to a 799-nt
exon and contains two short (53 and 57 codons) overlapping open reading
frames (ORFs). The 53-codon ORF was previously designated BORFE1.
Neither ORF exhibits detectable amino acid sequence homology with ORFs
of other gammaherpesviruses. The 1.1-kb RNA promoter-regulatory region was
speciﬁcally transactivated by the BHV-4 IE2 gene product, a homolog of the
Epstein-Barr virus R transactivator, in cotransfection assays. In gel retardation
experiments, IE2 protein formed a complex with DNA in a 129-bp fragment
between −23 and −151 relative to the transcription start site of the 1.1-kb RNA,
and less efﬁciently with a 57-bp subfragment between −78 and −22. A sequence
similar to sequences of IE2-binding fragments of other BHV-4 IE2 responsive
promoters was found partly in the 57-bp subfragment, extending into the portion
of the 129-bp fragment not found in the 57-bp fragment. The 129-bp fragment,
but not the 57-bp fragment, was sufﬁcient for transactivation of a promoterless
chloramphenicol acetyl transferase (CAT) reporter gene by IE2. However, the
129-bp fragment did not function efﬁciently as an IE2-responsive enhancer when
inserted approximately 140 base pairs (bp) 5
to the transcription start site of
a CAT reporter gene driven by an enhancerless simian virus 40 early promoter.
Based on this and other observations, we propose that IE2 functions as a promoter
factor rather than an enhancer factor.
Present addresses: Department of Genetics and Molecular Biology, Centro de Investi-
gacion y de Estudios Avanzados del IPN, Zacatenco, Mexico D.F.
Veterans Affairs Medical Center 151, Decatur, GA, U.S.A.