The herpesvirus saimiri ORF50 gene, encoding a transcriptional activator homologous to the Epstein-Barr virus R protein, is transcribed from two distinct promoters of different temporal phases

Abstract

The mRNA species encoding the herpesvirus saimiri (HVS) homolog of the Epstein-Barr virus R transcriptional activator (termed ORF50) have been identified and used to determine transcriptional start sites within the gene. The first transcript is spliced and starts from a promoter within ORF49 containing a single intron; the second is produced from a promoter within the second exon and is in the same reading frame. The spliced transcript is detected at early times during productive virus replication in OMK cells, whereas the nonspliced transcript is detected later. The spliced transcript is fivefold-more potent in activating the delayed-early ORF6 promoter; the function of the nonspliced transcript is unclear. Thus, the role of this protein in activating herpesvirus saimiri from the latent state may differ significantly from that of the Epstein-Barr virus R protein.Herpesvirus saimiri (HVS) establishes asymptomatic infections involving a subset of T lymphocytes in its natural host, the squirrel monkey (Saimiri sciureus), but causes fatal T-cell lymphomas and lymphoproliferative diseases in other species of New World primates (6). HVS has been classified as a gamma-2 herpesvirus and shares significant homology with Epstein-Barr virus (EBV) (1,2,9,10,16,24). The genomes of HVS and EBV are generally colinear, in that homologous genes are found in approximately equivalent locations and in the same relative orientations in the two viruses (22). As in many other herpesviruses, gene expression during lytic infection occurs in three main temporal phases: immediate-early (IE), delayed-early (DE), and late (17). The major IE transcript in HVS is encoded by the HindIII-G IE gene (ORF14) (2,(25)(26)(27). Analysis of the sequence shows that it does not exhibit homology with any EBV-encoded proteins (25), but it does contain local homology with a putative superantigen (29); however, its function remains to be established. The 52-kDa product of the HVS IE gene (ORF57) is homologous to the EBV transactivator encoded by BMLF1 (also known as Mta, M-IE, or IE-2) and to the 27-kDa or IE 63-kDa protein encoded by UL54 of herpes simplex virus 1 and RF4 of varicellazoster virus (4,14,21,22,24).The EcoRI-D or HVS R protein transactivator (ORF50) is homologous to the BRLF1 gene product (also known as R or Rta) (23). BRLF1 is translated from a bicistronic mRNA which also encodes another IE gene product, BZLF1 (12, 20). These two EBV gene products function in activating expression from a number of viral (3,5,8,13,15,18,28) and perhaps cellular (7) promoters, thereby initiating the lytic cascade. However, HVS R RNA is expressed as an early transcript in HVS-infected cells (23, 25). The HVS R gene is proposed to be spliced with more than one exon, although the initiation codon has not been mapped. Despite the likelihood of a spliced transcript, there is evidence that a DNA sequence within the identified exon is able to produce a functionally active protein capable of activating a DE ORF6 gene promoter, a component of the major DNA binding prot...

The Activation Domain of Herpesvirus Saimiri R Protein Interacts with the TATA-Binding Protein

“…Reduced CAT activity was observed when the deletion constructs were used to transactivate pAW-CAT2. However, pBK50b was shown to transactivate the ORF 6 promoter to levels similar to those found previously (52). This suggested that the sequences contained within bp 72350 to 72402 of the published sequence, which were deleted in pBK50⌬1, are essential for transactivation by the ORF 50b gene product.…”

“…Gene expression in HVS is modulated by the two major transcriptional regulating genes encoded by open reading frame (ORF) 50 and ORF 57 (41,42,52,54,55). The ORF 57 gene product encodes a multifunctional protein capable of both transactivation and repression of viral gene expression.…”

“…The spliced transcript is fivefold more potent in activating the delayedearly ORF 6 promoter. However, the function of the nonspliced transcript is unclear (41,52). Further analysis of the ORF 50 gene products have demonstrated that they activate transcription directly following interactions with promoters containing a specific sequence motif.…”

The Immediate-Early Gene Product Encoded by Open Reading Frame 57 of Herpesvirus Saimiri Modulates Gene Expression at a Posttranscriptional Level

“…ORF 50 produces two transcripts. The first is spliced containing a single intron and is detected at early times during the productive cycle, whereas the second is expressed later and is produced from a promoter within the second exon (30). In this paper, we show that the ORF 57 gene product represses the transactivation capability of the ORF 50a gene product (which is produced from a spliced transcript), but activates that of ORF 50b (an unspliced transcript).…”

“…We were particularly interested in determining whether the ORF 57 gene product exerted any regulatory effect on another transcriptional activator encoded by HVS, ORF 50. This gene encodes two transcripts: the first is spliced and is detected at early times during the productive cycle, whereas the second is expressed later and is produced from a promoter within the second exon (30). Therefore, transfection studies were performed to assess the effect of the ORF 57 gene product on the transactivation capability of the ORF 50 gene products.…”