MERS-CoV and H5N1 influenza virus antagonize antigen presentation by altering the epigenetic landscape

Menachery, Vineet D.; Schäfer, Alexandra; Burnum-Johnson, Kristin E.; Mitchell, Hugh D.; Eisfeld, Amie J.; Walters, Kevin B.; Nicora, Carrie D.; Purvine, Samuel O.; Casey, Cameron P.; Monroe, Matthew E.; Weitz, Karl K.; Stratton, Kelly G.; Webb-Robertson, Bobbie-Jo M.; Gralinski, Lisa E.; Metz, Thomas O.; Smith, Richard D.; Waters, Katrina M.; Sims, Amy C.; Kawaoka, Yoshihiro; Baric, Ralph S.

However, it is possible that DNA methylation is due to stress induced by viral infection. Both MERS-CoV and H5N1-VN1203 produced significant cytopathic effect by 24 h postinfection (hpi), contrasting both SARS-CoV and H1N1-09. However, while a global increase in methylation was observed following MERS-CoV and H5N1-VN1203 infection, it was far from uniform across the entire genome. More in-depth analysis of methylation data found that, following MERS-CoV infection, the (Table S3) found lower meCpG/Mbp rates, suggesting specificity in the magnitude of DNA methylation at the MHC locus and potentially other genomic locations. Similarly, H5N1-VN1203 methylation within the MHC locus corresponded to 34.29 meCpG/Mbp vs. 7.7 meCpG/Mbp over the remainder of the chr. 6. Together, these results suggest that DNA methylation is at least one of the primary mechanisms exploited by MERS-CoV to antagonize antigen-presentation gene expression in infected cells, while H5N1-VN1203 may use a combination of DNA methylation and histone-modification alterations to achieve a similar affect. Both Viral and Host Processes Contribute to DNA Methylation. To examine how these respiratory viruses target antigen presentation via epigenetic modulation, we utilized MERS-CoV and H5N1-VN1203 mutants that lacked key viral proteins. Previous work by our group implicated influenza NS1 in histone modification and ISG down-regulation compared with a similarly replication attenuated PB2 mutant (6) . In this study, we extended this analysis to antagonism of antigen-presentation molecules. Following infection of Calu3 cells with wild-type (WT), C-terminal truncation of NS1 (NS1trunc124), or PB2 encoding an amino acid substitution (PB2-K627E) mutant viruses, we examined changes in viral replication as well as HLA-A and -C peptide abundance ( Fig. 6 A and B ). While the overall peptide level was decreased relative to mock, truncation of H5N1-VN1203 NS1 resulted in augmented protein expression of both MHC class I molecules relative to the WT virus. Importantly, the reduction of MHC class I downregulation was not a product of attenuated viral replication, as the PB2 mutant had similar replication kinetics and titer, but maintained HLA-A and -C antagonism (Fig. S3A) . Together, the results implicate a role for NS1 in modulation of antigen presentation following H5N1-VN1203 infection.