Premature expression of the latency-related RNA encoded by bovine herpesvirus type 1 correlates with higher levels of beta interferon RNA expression in productively infected cells

Abstract

Bovine herpesvirus type 1 (BHV-1) is an important pathogen that can initiate bovine respiratory disease complex. Like other members of the subfamily Alphaherpesvirinae, BHV-1 establishes latency in sensory neurons. The latency-related (LR) gene expresses a family of alternatively spliced transcripts in infected sensory neurons that have the potential to encode several LR proteins. An LR mutant virus that contains three stop codons near the 59 terminus of the first open reading frame in the LR gene does not express two LR proteins or reactivate from latency. In addition, the LR mutant virus induces higher levels of apoptosis in trigeminal ganglionic neurons and grows less efficiently in certain tissues of infected calves. In spite of the reduced pathogenesis, the LR mutant virus, wild-type BHV-1 and the LR rescued virus exhibit identical growth properties in cultured bovine cells. In this study, we demonstrated that during early phases of productive infection the LR mutant virus expressed higher levels of LR-RNA relative to the LR rescued virus or wt BHV-1. Bovine kidney cells infected with the LR mutant virus also induced higher levels of beta interferon RNA and interferon response genes. These results suggest that inappropriate expression of LR-RNA, in the absence of LR protein expression, may influence the latency-reactivation cycle and pathogenic potential of BHV-1.

Dexamethasone Treatment of Calves Latently Infected with Bovine Herpesvirus 1 Leads to Activation of the bICP0 Early Promoter, in Part by the Cellular Transcription Factor C/EBP-Alpha

“…Previous studies demonstrated that the LR rescued virus but not the LR mutant virus reactivated from latency after DEX treatment (14,(31)(32)(33). To test whether IE, E, or L genes were expressed in TG following DEX treatment, RT-PCR was performed using primers that specifically amplify viral genes.…”

“…TG were stored at Ϫ80°C for nucleic acid extraction or processed for histological procedures. Part of the TG used for this study was also used for previous studies (14,(31)(32)(33).…”

“…BHV-1-free crossbred calves (ϳ200 kg) were used for this study. Calves were inoculated with 10 7 PFU of the indicated virus into each nostril and eye, for a total of 4 ϫ 10 7 PFU/animal, as described previously (14,15,25,(31)(32)(33). Calves were housed under strict isolation and given antibiotics before and after BHV-1 infection to prevent secondary bacterial infection.…”

“…For the construction of the LR mutant virus, 25 bp of the LR gene sequence from the Cooper strain were replaced with an oligonucleotide that contains a unique EcoRI restriction site and three stop codons to prevent protein expression (15). The LR mutant virus was previously characterized both in vitro and in vivo (14,15,25,(31)(32)(33). Stock cultures of the respective viruses were prepared in bovine cells (MDBK).…”

“…Calves infected with the LR mutant virus exhibit diminished clinical symptoms and reduced shedding of infectious virus in the eye, tonsil, or TG (14,15,33). The LR mutant virus prematurely expresses LR RNA relative to rescued or wt BHV-1, which correlates with an enhanced interferon response in cells infected with the LR mutant virus (31).…”