title: 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.
authors.affiliation: School of Veterinary Medicine and Science, University of Nottingham, Sutton Bonington LE12 5RD, United Kingdom. Electronic address: nevi.parameswaran@googlemail.com.
authors.forename: Nevi
authors.initials: N
authors.lastname: Parameswaran
authors.affiliation: Immunology-Vaccinology, Faculty of Veterinary Medicine, University of Liege, Liege, Belgium. Electronic address: bgdewals@ulg.ac.be.
authors.forename: Benjamin G
authors.initials: BG
authors.lastname: Dewals
authors.affiliation: Advanced Data Analysis Centre, University of Nottingham, Sutton Bonington LE12 5RD, United Kingdom. Electronic address: tom.giles@nottingham.ac.uk.
authors.forename: Tom C
authors.initials: TC
authors.lastname: Giles
authors.affiliation: Department of Biology, University of Virginia, Charlottesville, USA. Electronic address: deppmann@virginia.edu.
authors.forename: Christopher
authors.initials: C
authors.lastname: Deppmann
authors.affiliation: Centre for Genetics and Genomics, University of Nottingham, Nottingham, United Kingdom. Electronic address: martin.blythe@nottingham.ac.uk.
authors.forename: Martin
authors.initials: M
authors.lastname: Blythe
authors.affiliation: Immunology-Vaccinology, Faculty of Veterinary Medicine, University of Liege, Liege, Belgium. Electronic address: a.vdplasschen@ulg.ac.be.
authors.forename: Alain
authors.initials: A
authors.lastname: Vanderplasschen
authors.affiliation: School of Veterinary Medicine and Science, University of Nottingham, Sutton Bonington LE12 5RD, United Kingdom; Advanced Data Analysis Centre, University of Nottingham, Sutton Bonington LE12 5RD, United Kingdom. Electronic address: richard.emes@nottingham.ac.uk.
authors.forename: Richard D
authors.initials: RD
authors.lastname: Emes
authors.affiliation: School of Veterinary Medicine and Science, University of Nottingham, Sutton Bonington LE12 5RD, United Kingdom. Electronic address: david.haig@nottingham.ac.uk.
authors.forename: David
authors.initials: D
authors.lastname: Haig
mesh_terms: D000818:Animals; D005260:Female; D017353:Gene Deletion; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral; D006564:Herpesviridae / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology*; D054884:Host-Pathogen Interactions; D008304:Malignant Catarrh / Q000473:pathology / Q000821:virology*; D011817:Rabbits; D060888:Real-Time Polymerase Chain Reaction; D012376:Rodent Diseases / Q000473:pathology / Q000821:virology*; D017422:Sequence Analysis, DNA; D013601:T-Lymphocytes / Q000821:virology*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014774:Virulence; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism; D014779:Virus Replication
chemical_list: D014157:Transcription Factors; D037521:Virulence Factors
keywords: Cytotoxicity; Herpesvirus; Malignant catarrhal fever; Transcription regulation; bZIP protein
references.citation: Res Vet Sci. 1984 Mar;36(2):205-11
references.pmid: 6609389
references.citation: J Cell Physiol. 2005 May;203(2):305-18
references.pmid: 15389624
references.citation: Res Vet Sci. 1983 Jan;34(1):109-13
references.pmid: 6601284
references.citation: Vet Res. 2012 Jun 11;43:51
references.pmid: 22686373
references.citation: Res Vet Sci. 1986 Jul;41(1):76-81
references.pmid: 3764105
references.citation: Nat Rev Immunol. 2006 Aug;6(8):595-601
references.pmid: 16868550
references.citation: Vet Immunol Immunopathol. 1991 Aug;29(1-2):151-61
references.pmid: 1683050
references.citation: PLoS One. 2009 Jul 15;4(7):e6265
references.pmid: 19603070
references.citation: Nucleic Acids Res. 2005 Feb 24;33(4):e36
references.pmid: 15731329
references.citation: J Virol. 2011 Jul;85(14):6941-54
references.pmid: 21593175
references.citation: Proc Natl Acad Sci U S A. 2004 Aug 10;101(32):11815-20
references.pmid: 15289599
references.citation: Immunology. 2001 Jan;102(1):44-52
references.pmid: 11168636
references.citation: Adv Exp Med Biol. 2009;647:64-93
references.pmid: 19760067
references.citation: Res Vet Sci. 1989 Jul;47(1):90-6
references.pmid: 2772411
references.citation: PLoS One. 2008 Feb 20;3(2):e1627
references.pmid: 18286184
references.citation: J Clin Microbiol. 1994 Jul;32(7):1674-9
references.pmid: 7523438
references.citation: J Comp Pathol. 2008 Feb-Apr;138(2-3):131-44
references.pmid: 18331739
references.citation: Vet Pathol. 1980 Jan;17(1):73-83
references.pmid: 7352365
references.citation: Res Vet Sci. 1975 Sep;19(2):159-66
references.pmid: 1166120
references.citation: Vaccine. 2008 Aug 18;26(35):4461-8
references.pmid: 18601965
references.citation: J Virol. 2000 Feb;74(4):1593-601
references.pmid: 10644328
references.citation: Curr Opin Virol. 2012 Aug;2(4):453-8
references.pmid: 22858118
references.citation: Vet Pathol. 1980 Jan;17(1):58-72
references.pmid: 7352364
references.citation: Virology. 1993 Sep;196(1):232-40
references.pmid: 8356795
references.citation: Vet J. 2009 Mar;179(3):324-35
references.pmid: 18760944
references.citation: Virus Res. 2012 Oct;169(1):246-54
references.pmid: 22925730
references.citation: Mol Biol Evol. 2006 Aug;23(8):1480-92
references.pmid: 16731568
references.citation: J Gen Virol. 2006 Mar;87(Pt 3):509-517
references.pmid: 16476972
references.citation: Vet Immunol Immunopathol. 1998 Nov 6;66(1):67-81
references.pmid: 9847021
references.citation: Cancer Res. 2004 Apr 15;64(8):2774-81
references.pmid: 15087393
references.citation: J Virol. 2013 Mar;87(6):3597-604
references.pmid: 23302876
references.citation: Proc Natl Acad Sci U S A. 2013 May 21;110(21):E1933-42
references.pmid: 23630278
references.citation: Vet Microbiol. 2006 Aug 25;116(1-3):29-36
references.pmid: 16621344
references.citation: BMC Bioinformatics. 2004 Aug 19;5:113
references.pmid: 15318951
references.citation: J Gen Virol. 2007 Jan;88(Pt 1):28-39
references.pmid: 17170433
references.citation: J Gen Virol. 2003 Aug;84(Pt 8):1941-1949
references.pmid: 12867624
references.citation: Nature. 2013 Jun 13;498(7453):236-40
references.pmid: 23685454
references.citation: J Gen Virol. 2007 Dec;88(Pt 12):3224-3233
references.pmid: 18024890
references.citation: Nucleic Acids Res. 2012 Jan;40(Database issue):D84-90
references.pmid: 22086963
references.citation: J Gen Virol. 1996 May;77 ( Pt 5):953-61
references.pmid: 8609492
references.citation: Vet Microbiol. 2010 Sep 28;145(1-2):47-53
references.pmid: 20381274
references.citation: J Comp Pathol. 1988 Jan;98(1):99-109
references.pmid: 3346394
references.citation: Front Microbiol. 2012 Jul 09;3:247
references.pmid: 22787458
references.citation: J Virol Methods. 2005 Nov;129(2):186-90
references.pmid: 15998545
references.citation: Vet Immunol Immunopathol. 2010 Aug 15;136(3-4):284-91
references.pmid: 20413164
references.citation: Oncogene. 2003 Aug 11;22(33):5108-21
references.pmid: 12910248
references.citation: J Comp Pathol. 2007 Feb-Apr;136(2-3):156-66
references.pmid: 17374379