Characterising the role of murine gammaherpesvirus-68 host shutoff protein in mediating cellular mRNA degradation

Sheridan, Victoria Characterising the role of murine gammaherpesvirus-68 host shutoff protein in mediating cellular mRNA degradation. Doctor of Philosophy thesis, University of Liverpool.

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Abstract

During lytic infection with a number of pathogenic viruses, host cells have been found to exhibit loss of mRNA culminating in loss of cellular proteins, a phenomenon termed host shutoff. In γ-herpesviruses, the precise mechanism by which host shutoff takes place is yet to be fully determined. Initially, host shutoff in Kaposi’s sarcoma-associated herpesvirus (KSHV) was associated with the viral DNA exonuclease SOX (shutoff and exonuclease), which was followed by the identification of host shutoff in Epstein Barr virus (EBV) linked to the SOX homologue BGLF5. Murine γ-herpesvirus-68 (MHV-68) is a γ-herpesvirus that infects murid rodents including laboratory mice, with extensive viral genome homology to KSHV and EBV. MHV-68 was also found to induce host shutoff via the SOX homologue ORF37. The similarities between MHV-68 and the human γ-herpesviruses make the mouse virus an ideal model for studying host shutoff and consequences of host shutoff in disease pathogenesis. In this study, MHV-68 was used to investigate the role of ORF37 in MHV-68 infection in vitro and in vivo and to examine possible mechanisms of mRNA degradation mediated by ORF37. A stop codon mutant of MHV-68 (ORF37Stop) and its corresponding revertant (ORF37StopRev) were engineered. ORF37Stop virus exhibited highly restrictive phenotype in immunocompetent cells. In contrast, lack of ORF37 was tolerated in cells lacking a functional intereferon- α/β receptor (IFNαβRKO). These in vitro observations were replicated, to some extent, in vivo with the mutant virus establishing lytic infection in the lungs of IFNαβRKO mice. Interestingly, host shutoff was found to proceed even in the absence of ORF37, but only where functional type I IFN-mediated responses were lacking. A poly(A) tail assay was used to investigate the possibility of mRNA degradation at the 3’ mRNA. In contrast to a previously published study, the results suggest that cellular poly(A) tails do not appear to be elongated as a result of MHV-68 lytic infection. Data generated during this project have identified a novel function by which ORF37-mediated host shutoff causes mRNA degradation and that a primary role of this viral protein is to overcome the type I interferon-receptor-mediated responses. The observation that host shutoff can proceed in the absence of ORF37 in cells unable to respond to type I IFNs highlight two important possibilities: other viral proteins may mediate host shutoff; and type I IFN effector mechanisms are a key player in host shutoff. These novel observations together with lack of a mechanism by which ORF37 mediates mRNA degradation warrant further studies in this field.

Item Type:	Thesis (Doctor of Philosophy)
Additional Information:	Date: 2012-09 (completed)
Uncontrolled Keywords:	MHV-68, DNA exonuclease, host shutoff, gammaherpesvirus
Subjects:	?? QH426 ??
Divisions:	Faculty of Health and Life Sciences
Depositing User:	Symplectic Admin
Date Deposited:	10 Sep 2013 08:52
Last Modified:	16 Dec 2022 04:38
DOI:	10.17638/00009273
Supervisors:	Ebrahimi, Bahram Stewart, James
URI:	https://livrepository.liverpool.ac.uk/id/eprint/9273