Intracellular interferons in fish : a unique means to combat viral infection

Peer reviewedPublisher PD

Sculpting the Bacterial <i>O</i>-Glycoproteome: Functional Analyses of Orthologous Oligosaccharyltransferases with Diverse Targeting Specificities

Protein glycosylation systems are widely recognized in bacteria, including members of the genus Neisseria. In most bacterial species, the molecular mechanisms and evolutionary contexts underpinning target protein selection and the glycan repertoire remain poorly understood. Broad-spectrum O-linked protein glycosylation occurs in all human-associated species groups within the genus Neisseria, but knowledge of their individual glycoprotein repertoires is limited. Interestingly, PilE, the pilin subunit of the type IV pilus (Tfp) colonization factor, is glycosylated in Neisseria gonorrhoeae and Neisseria meningitidis but not in the deeply branching species N. elongata subsp. glycolytica. To examine this in more detail, we assessed PilE glycosylation status across the genus and found that PilEs of commensal clade species are not modified by the gonococcal PglO oligosaccharyltransferase. Experiments using PglO oligosaccharyltransferases from across the genus expressed in N. gonorrhoeae showed that although all were capable of broad-spectrum protein glycosylation, those from a deep-branching group of commensals were unable to support resident PilE glycosylation. Further glycoproteomic analyses of these strains using immunoblotting and mass spectrometry revealed other proteins differentially targeted by otherwise remarkably similar oligosaccharyltransferases. Finally, we generated pglO allelic chimeras that begin to localize PglO protein domains associated with unique substrate targeting activities. These findings reveal previously unappreciated differences within the protein glycosylation systems of highly related bacterial species. We propose that the natural diversity manifest in the neisserial protein substrates and oligosaccharyltransferases has significant potential to inform the structure-function relationships operating in these and related bacterial protein glycosylation systems. IMPORTANCE Although general protein glycosylation systems have been well recognized in prokaryotes, the processes governing their distribution, function, and evolution remain poorly understood. Here, we have begun to address these gaps in knowledge by comparative analyses of broad-spectrum O-linked protein glycosylation manifest in species within the genus Neisseria that strictly colonize humans. Using N. gonorrhoeae as a well-defined model organism in conjunction with comparative genomics, intraspecies gene complementation, and glycoprotein phenotyping, we discovered clear differences in both glycosylation susceptibilities and enzymatic targeting activities of otherwise largely conserved proteins. These findings reveal previously unappreciated differences within the protein glycosylation systems of highly related bacterial species. We propose that the natural diversity manifest within Neisseria species has significant potential to elucidate the structure-function relationships operating in these and related systems and to inform novel approaches to applied glycoengineering strategies

Antiviral Activity of Salmonid Gamma Interferon against Infectious Pancreatic Necrosis Virus and Salmonid Alphavirus and Its Dependency on Type I Interferon▿

We investigated the antiviral activity and gene induction properties of interferon gamma (IFN-γ) compared to type I IFN (IFNa1) in Atlantic salmon. IFN-γ protected salmon cells against infectious pancreatic necrosis virus (IPNV)-induced cytopathic effect (CPE), reduced virus titers, and inhibited the synthesis of the viral structural protein VP3. Moreover, IFN-γ showed potent antiviral activity against salmonid alphavirus 3 (SAV3) measured as a reduction in virus nsP1 transcripts. IFN-γ (a type II IFN) had less specific antiviral activity against IPNV than IFNa1, showing a half-maximal effective concentration of 1.6 ng/ml versus 31 pg/ml determined in the CPE reduction assay. Compared to IFNa1, IFN-γ was a more effective inducer of the antiviral protein GBP, several interferon regulatory transcription factors (IRFs), and the chemokine IP-10. The antiviral activity of IFN-γ may also in part be ascribed to upregulation of Mx, ISG15, and viperin. These are typical type I IFN-induced genes in mammals and were also more strongly induced by IFNa1 than by IFN-γ in salmon cells. Fish and mammalian IFN-γ thus show strikingly similar gene induction properties. Interestingly, the antiviral activity of IFN-γ against IPNV and SAV3 and its ability to induce Mx and ISG15 markedly decreased in the presence of neutralizing antiserum against IFNa1. In contrast, antiIFNa1 had no effect on the induction of IRF-1 and IP-10 by IFN-γ. This suggests that the antiviral activity of IFN-γ is partially dependent on IFNa induction. However, because antiIFNa1 could not abolish the IFN-γ-mediated induction of Mx and ISG15 completely, IFN-γ may possibly also induce such genes directly