title: Granzyme B inhibits vaccinia virus production through proteolytic cleavage of eukaryotic initiation factor 4 gamma 3.
abstract: Cytotoxic T lymphocytes (CTLs) are the major killer of virus-infected cells. Granzyme B (GrB) from CTLs induces apoptosis in target cells by cleavage and activation of substrates like caspase-3 and Bid. However, while undergoing apoptosis, cells are still capable of producing infectious viruses unless a mechanism exists to specifically inhibit viral production. Using proteomic approaches, we identified a novel GrB target that plays a major role in protein synthesis: eukaryotic initiation factor 4 gamma 3 (eIF4G3). We hypothesized a novel role for GrB in translation of viral proteins by targeting eIF4G3, and showed that GrB cleaves eIF4G3 specifically at the IESD(1408)S sequence. Both GrB and human CTL treatment resulted in degradation of eIF4G3 and reduced rates of translation. When Jurkat cells infected with vaccinia virus were treated with GrB, there was a halt in viral protein synthesis and a decrease in production of infectious new virions. The GrB-induced inhibition of viral translation was independent of the activation of caspases, as inhibition of protein synthesis still occurred with addition of the pan-caspase inhibitor zVAD-fmk. This demonstrated for the first time that GrB prevents the production of infectious vaccinia virus by targeting the host translational machinery.
authors.affiliation: Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada.
authors.forename: Marcelo
authors.initials: M
authors.lastname: Marcet-Palacios
authors.forename: Brenda Lee
authors.initials: BL
authors.lastname: Duggan
authors.forename: Irene
authors.initials: I
authors.lastname: Shostak
authors.forename: Michele
authors.initials: M
authors.lastname: Barry
authors.forename: Tracy
authors.initials: T
authors.lastname: Geskes
authors.forename: John A
authors.initials: JA
authors.lastname: Wilkins
authors.forename: Akiko
authors.initials: A
authors.lastname: Yanagiya
authors.forename: Nahum
authors.initials: N
authors.lastname: Sonenberg
authors.forename: R Chris
authors.initials: RC
authors.lastname: Bleackley
mesh_terms: D017209:Apoptosis / Q000502:physiology; D053148:Caspase 3 / Q000378:metabolism; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D039603:Eukaryotic Initiation Factor-4G / Q000378:metabolism*; D053804:Granzymes / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D010448:Peptide Initiation Factors / Q000378:metabolism*; D059748:Proteolysis; D040901:Proteomics; D013602:T-Lymphocytes, Cytotoxic / Q000378:metabolism*; D014616:Vaccinia virus / Q000378:metabolism* / Q000502:physiology; D014779:Virus Replication / Q000502:physiology
chemical_list: D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D039603:Eukaryotic Initiation Factor-4G; D010448:Peptide Initiation Factors; C077382:eIF-4gamma; C509213:GZMB protein, human; D053804:Granzymes; D053148:Caspase 3; D020169:Caspases