The A2 gene of alcelaphine herpesvirus-1 is a transcriptional regulator affecting cytotoxicity in virus-infected T cells but is not required for malignant catarrhal fever induction in rabbits

Abstract

Alcelaphine herpesvirus-1 (AlHV-1) causes malignant catarrhal fever (MCF). The A2 gene of AlHV-1 is a member of the bZIP transcription factor family. We wished to determine whether A2 is a virulence gene or not and whether it is involved in pathogenesis by interference with host transcription pathways. An A2 gene knockout (A2ΔAlHV-1) virus, revertant (A2revAlHV-1) virus, and wild-type virus (wtAlHV-1) were used to infect three groups of rabbits. A2ΔAlHV-1-infected rabbits succumbed to MCF, albeit with a delayed onset compared to the control groups, so A2 is not a critical virulence factor. Differential gene transcription analysis by RNAseq and qRT-PCR validation of a selection of these was performed in infected large granular lymphocyte (LGL) T cells obtained in culture from the MCF-affected animals. A2 was involved in the transcriptional regulation of immunological, cell cycle and apoptosis pathways. In particular, there was a bias towards γδ T cell receptor (TCR) expression and downregulation of αβ TCR. TCR signalling, apoptosis, cell cycle, IFN-γ and NFAT pathways were affected. Of particular interest was partial inhibition of the cytotoxicity-associated pathways involving perforin and the granzymes A and B in the A2ΔAlHV-1-infected LGLs compared to controls. In functional assays, A2ΔAlHV-1-infected LGLs were significantly less cytotoxic than wtAlHV-1- and A2revAlHV-1-infected LGLs using rabbit corneal epithelial cells (SIRC) as targets. This implies that A2 is involved in a pathway enhancing the expression of LGL cytotoxicity. This is important as virus-infected T cell cytotoxicity in vivo has been suggested as a potential mechanism of disease induction in MCF.

Genomic duplication and translocation of reactivation transactivator and bZIP-homolog genes is a conserved event in alcelaphine herpesvirus 1

“…The entire genome of strain C500 has been cloned from low-passage, virulent virus as a bacterial artificial chromosome (BAC), in which infectivity and pathogenicity have been shown to be preserved24. The BAC clone is therefore an invaluable tool for studying the biology and pathogenesis of MCF78252627. The overall genome of strain C500 has been shown to be present in the BAC clone24, but the full sequence, and therefore the possible existence of undetected genetic changes, has not been determined.…”

Ov2 is a modulator of OvHV-2 RTA mediated gene expression

“…Ov2 shares significant homology, around the b-ZIP domain with A2, its positional homolog in AlHV-1 (Parameswaran et al 2014 ). The observation made by Parameswaran et al that apoptotic pathways are upregulated in rabbit LGL cells derived from rabbits infected with an A2 deletant virus, compared to controls is consistent with a model in which A9 expression is regulated through A2 (Parameswaran et al 2014 ). The availability of a bacterial artificial chromosome of AlHV-1, may prove a valuable tool in delineating to what extent the functions of Ov2/A2, and those of Ov9/A9, are conserved (Dewals et al 2006 ).…”

“…The genome of AlHV-1, a related Macavirus , contains a positional homologue of Ov2, termed A2, that also contains a bZIP domain (Ensser et al 1997 ). In a rabbit model of MCF, up regulation of cellular apoptosis pathways was observed following infection with a virus carrying a deletion of the A2 gene (Parameswaran et al 2014 ).…”

Wildebeest-Derived Malignant Catarrhal Fever: A Bovine Peripheral T Cell Lymphoma Caused by Cross-Species Transmission of Alcelaphine Gammaherpesvirus 1

“…AlHV-1 encodes the basic leucine zipper (bZIP) family of proteins that likely regulate host and/or virus gene transcription, leading to changes that could result in lymphoproliferation [ 119 ]. Interestingly, AlHV-1 A2 is a bZIP family protein [ 103 ]. Deletion of A2 from the AlHV-1 genome does not alter viral replication nor the ability of AlHV-1 to induce WD-MCF in a rabbit model, suggesting that it is not essential on its own in driving WD-MCF pathogenesis [ 103 ].…”