Post-translational modification of the myxoma-virus anti-inflammatory serpin SERP-1 by a virally encoded sialyltransferase

Abstract

SERP-1 is a secreted serpin (serine-proteinase inhibitor) encoded by myxoma virus, a poxvirus pathogen of rabbits. SERP-1 is required for myxoma-virus virulence, and the purified protein has been shown to possess independent anti-inflammatory activity in animal models of restenosis and antigen-induced arthritis. As an inhibitor of serine proteinases, SERP-1 acts against tissue-type plasminogen activator, urokinase-type plasminogen activator, plasmin, thrombin and Factor Xa. In the present study, examination of SERP-1 glycosylation-site mutants showed that the N-linked glycosylation of Asn(172) was essential for SERP-1 secretion, whereas mutation of Asn(99) decreased secretion efficiency, indicating that N-linked glycosylation plays an essential role in the processing and trafficking of SERP-1. Furthermore, comparison of SERP-1 from wild-type myxoma virus and a virus containing a targeted disruption of the MST3N sialyltransferase locus demonstrated that SERP-1 is specifically modified by this myxoma-virus-encoded sialyltransferase, and is thus the first reported viral protein shown to by modified by a virally encoded glycosyltransferase. Sialylation of SERP-1 by the MST3N gene product creates a uniquely charged species of secreted SERP-1 that is distinct from SERP-1 produced from other eukaryotic expression systems, though this has no apparent effect upon the kinetics of in vitro proteinase inhibition. Rather, the role of viral sialylation of SERP-1 likely relates to masking antigenicity or targeting SERP-1 to specific sites of action in vivo.

Myxoma Virus Protein M029 Is a Dual Function Immunomodulator that Inhibits PKR and Also Conscripts RHA/DHX9 to Promote Expanded Host Tropism and Viral Replication

“…HRP-conjugated goat anti-rabbit and anti-mouse IgG antibodies were purchased from Jackson ImmunoResearch Laboratories. Generation of rabbit polyclonal and mouse monoclonal antibodies against MYXV proteins M-T7 and Serp1 was described before [61], [62].…”

M062 Is a Host Range Factor Essential for Myxoma Virus Pathogenesis and Functions as an Antagonist of Host SAMD9 in Human Cells

“…For Western blot analysis, antibodies targeting the human SAMD9 protein (Sigma-Aldrich), c-Myc (9E10; Santa Cruz Biotechnology), anti-V5 (Invitrogen), anti-␤-actin (Ambion), goat anti-mouse IgG conjugated with peroxidase (Thermo Scientific), and goat anti-rabbit IgG conjugated with peroxidase (Santa Cruz Biotechnology) were purchased commercially for this study. Antibodies against MYXV SERP-1 and M063 were previously described (4,19). Briefly, after the addition of Laemmli loading buffer and heat treatment, 25 to 50 g of total protein or samples from pull-down or co-IP were separated by SDS-PAGE before transferring to HyBond-P hydrophobic polyvinylidene difluoride (PVDF) membrane (GE Healthcare) using the XCell II Blot Module (Invitrogen).…”

Bitter-sweet symphony: glycan–lectin interactions in virus biology

“…Considering the restricted glycosylation enzyme and precursor availabilities, it is conceivable that overexpression of viral glycoproteins in an infected target cell can result in an increased glycan heterogeneity of both viral and cellular glycoconjugates. Moreover, viruses may also actively modify the host and viral glycome by modulating the expression of host cell glycoenzymes (Hiraiwa et al, 1997;Cebulla et al, 2000;Hiraiwa et al, 2003) or via expression of virally encoded glycoenzymes in infected cells (Jackson et al, 1999;Willer et al, 1999;Nash et al, 2000;Sujino et al, 2000;Vanderplasschen et al, 2000;Markine-Goriaynoff et al, 2003, 2004a.…”