Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics
Differential cleavage of viral polypeptides by allotypic variants of granzyme B skews immunity to mouse cytomegalovirus
Section snippets
List of non-standard abbreviations
MCMV mouse cytomegalovirus GzmB granzyme B B6 inbred mouse strain C57BL/6 AAD benzyloxycarbonyl-Ala-Ala-Asp-S Benzyl IEPD Acetyl-Ile- Glu-Pro-Asp-paranitroanilide MOMP mitochondrial outer membrane permeabilization tBid truncated Bid MHC major histocompatibility complex AV annexin V PI propidium iodide TAILS terminal amine isotopic labelling of substrate MEF mouse embryonic fibroblast CTL cytotoxic T lymphocyte NK natural killer Asp-ase cleavage after a P1 aspartate residue.
Mice
Typical inbred B6 mice (which have the P allele, GzmBP/P) [15] were purchased from the Walter and Eliza Hall Institute (WEHI), Melbourne, Australia. B6.GzmAB−/− and B6.GzmBW/W mice were bred and maintained at the Peter MacCallum Cancer Centre. Male and female mice were used at 6–10 weeks of age. The study was conducted according to the Australian Code of Practice for the care and use of animals for scientific purposes and the Australian National Health and Medical Research guidelines and
Results
We previously made the surprising finding that CD8+ CTLs generated by B6 mice homozygous for the BP allele of GzmB efficiently kill target cells infected with MCMV whereas those of congenic mice homozygous for BW fail to do so. This lack of cytotoxicity in vitro translates into marked susceptibility of mice homozygous for GzmBW to acute infection with MCMV Δ157 in vivo, as 100% of these mice die from overwhelming viral hepatitis and liver failure within 7 days, whereas GzmBP-expressing mice
Discussion
Although they recognise and bind dangerous cells via very different mechanisms, CTL and NK cells share a single, contact-dependent pathway to inflict target cell death once a stable immune synapse has formed. Apart from erythrocytes, which do not express major histocompatibility (MHC) molecules, any somatic cell can be eliminated via ‘granule exocytosis’ – most commonly because they are harbouring a virus or are on a path to malignant transformation [1,2]. Through granule exocytosis, two quite
Conclusion
The current study has advanced our investigations into the mechanism through which heterogeneity in GzmB structure and subtle variations in its enzymatic function can lead to radical differences in outcome following MCMV infection in vivo. Using a variety of biochemical and proteomic approaches, we identified and characterised a number of endogenous and viral proteins which are differentially processed by the GzmBW and GzmBP alloforms. We found no definitive evidence that activation of
Credit author statement
Vivien Sutton – conceptualisation; investigation, supervision of research staff; writing of original manuscript draft; review and editing; Chris Andoniou - conceptualisation; investigation; Michael Leeming – data curation; formal analysis; manuscript review and editing; Colin House – investigation and methodology; data curation; manuscript review and editing; Sally Watt – investigation and methodology; Sandra Verschoor – investigation and methodology; Annette Ciccone – investigation and
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgements
The work described here was supported by Program and Project grants to JAT, IV, MD-E and CA from the National Health and Medical Research Council of Australia. JAT is supported through generous philanthropic support as a Rosie Lew Fellow of the Peter Mac Research Foundation.
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Cited by (2)
Beyond target cell death – Granzyme serine proteases in health and disease
2022, Molecular Aspects of MedicineCitation Excerpt :In contrast, granzyme deficient mice have severely impaired MCMV control, although unlike perforin deficient mice, granzyme deficient mice survive infection again highlighting differences in perforin versus granzyme function (van Dommelen et al., 2006). In the case of MCMV, direct cleavage of MCMV proteins by granzymes may be more important for viral control than granzyme-dependent cytotoxicity (Andoniou et al., 2014; Sutton et al., 2020). Ectromelia is one of the few reported viral models where granzyme deficiency can phenocopy perforin loss in terms of mortality during infection (Müllbacher et al., 1999a, 1999b).