Characterization of an abundant, unique 1.7-kilobase bovine herpesvirus 4 (BHV-4) late RNA and mapping of a BHV-4 IE2 transactivator-binding site in its promoter-regulatory region

Abstract

We characterized an abundant late 1.7-kb cytoplasmic polyadenylated RNA (L1.7 RNA) transcribed from the bovine herpesvirus 4 (BHV-4) HindIII W fragment, in a region of the genome not conserved with Epstein-Barr virus and herpesvirus saimiri. L1.7 RNA contains only extremely short (<100 nucleotides) open reading frames followed by two repeat arrays. The first repeat array contains 11 copies of a 23-bp unit, TGGCACTA GTAGCATTTAACCCC. The second and third copies are each interrupted by 15-to 17-bp sequences that are identical to each other for the first 15 bp. In addition, the second and third copies of the repeat unit each contain two copies of nucleotides 5 to 9 (ACTAG) of the repeat unit, one at each end of the interruption. The second repeat array contains 12 copies of a 25-bp sequence, GCTGTGTATTATTGAGTATTTTTTA. The promoter-regulatory region of L1.7 was activated by the BHV-4 immediate-early gene 2 product (IE2), a homolog of the Epstein-Barr virus R transactivator, in cotransfection assays. We mapped an IE2 recognition site within a 167-bp fragment approximately 10 bp 5 to the start of L1.7 RNA transcription, using cotransfection assays and gel retardation assays. Using gel retardation assays, we mapped an IE2-binding site within this fragment to a 31-bp region from 56 to 86 bp 5 to the start of L1.7 RNA transcription. This IE2-binding site was able to transfer IE2 responsiveness to a heterologous promoter. However, IE2 responsiveness was affected by both position and orientation. Alignment of the L1.7 IE2-binding site sequence with sequences of two other BHV-4 IE2-binding sites resulted in a provisional IE2-binding site consensus sequence different from the Epstein-Barr virus R transactivator-binding site.

Two herpesviral noncoding PAN RNAs are functionally homologous but do not associate with common chromatin loci

“…Bioinformatic studies revealed ENEs, thus indicating the presence of PAN RNAs, in four other γ-herpesviruses: retroperitoneal fibromatosis-associated herpesvirus Macaca nemestrina (RFHVMn), Rhesus macaque rhadinovirus (RRV), and equine herpesvirus 2 and 5 (EHV-2 and EHV-5) [ 8 ]. A fifth PAN RNA homolog lacking an ENE was identified as a highly abundant noncoding RNA expressed from a syntenic genomic locus of bovine herpesvirus 4 (BHV-4) [ 8 , 17 ].…”

Establishment of a cell line persistently infected with bovine herpesvirus-4 by use of a recombinant virus

“…U, uninfected. The probe was a 32 P-labelled cloned cDNA of BHV-4 L1.7 RNA (Bermudez-Cruz et al ., 1997 ). Circular (C) and linear (L) viral genomes are indicated. ).…”

“…Total RNA prepared 24 h p.i. was assayed by Northern blot analysis for expression of representative early (BHV-4 homologue of herpes simplex virus 1 major DNA binding protein; Chang & van Santen, 1992 ) and late (L1.7; Bermudez-Cruz et al ., 1997 ) RNAs. Early RNA was detected in RD-4 cells at approximately one-fifth the level detected in BT cells (not shown).…”