Influenza A Virus-Induced Expression of a GalNAc Transferase, GALNT3, via MicroRNAs Is Required for Enhanced Viral Replication

Abstract

Influenza A virus (IAV) affects the upper and lower respiratory tracts and rapidly induces the expression of mucins, which are common O-glycosylated proteins, on the epithelial surfaces of the respiratory tract. Although mucin production is associated with the inhibition of virus transmission as well as characteristic clinical symptoms, little is known regarding how mucins are produced on the surfaces of respiratory epithelial cells and how they affect IAV replication. In this study, we found that two microRNAs (miRNAs), miR-17-3p and miR-221, which target GalNAc transferase 3 (GALNT3) mRNA, are rapidly downregulated in human alveolar basal epithelial cells during the early stage of IAV infection. We demonstrated that the expression of GALNT3 mRNA is upregulated in an IAV replication-dependent fashion and leads to mucin production in bronchial epithelial cells. A lectin microarray analysis revealed that the stable expression of GALNT3 by human alveolar basal epithelial cells induces mucin-type O-glycosylation modifications similar to those present in IAV-infected cells, suggesting that GALNT3 promotes mucin-type O-linked glycosylation in IAV-infected cells. Notably, analyses using short interfering RNAs and miRNA mimics showed that GALNT3 knockdown significantly reduces IAV replication. Furthermore, IAV replication was markedly decreased in embryonic fibroblast cells obtained from galnt3-knockout mice. Interestingly, IAV-infected galnt3-knockout mice exhibited high mortality and severe pathological alterations in the lungs compared to those of wild-type mice. Our results demonstrate not only the molecular mechanism underlying rapid mucin production during IAV infection but also the contribution of O-linked glycosylation to the replication and propagation of IAV in lung cells. IMPORTANCEViral infections that affect the upper or lower respiratory tracts, such as IAV, rapidly induce mucin production on the epithelial surfaces of respiratory cells. However, the details of how mucin-type O-linked glycosylation is initiated by IAV infection and how mucin production affects viral replication have not yet been elucidated. In this study, we show that levels of two miRNAs that target the UDP-GalNAc transferase GALNT3 are markedly decreased during the early stage of IAV infection, resulting in the upregulation of GALNT3 mRNA. We also demonstrate that the expression of GALNT3 initiates mucin production and affects IAV replication in infected cells. This is the first report demonstrating the mechanism underlying the miRNA-mediated initiation of mucin-type O-glycosylation in IAV-infected cells and its role in viral replication. Our results have broad implications for understanding IAV replication and suggest a strategy for the development of novel anti-influenza approaches.

SNP-mediated disruption of CTCF binding at the IFITM3 promoter is associated with risk of severe influenza in humans

“…Currently, there is no literature on the role that B4GALNT4 may play in host response to viral infection, but it is known that mice deficient in B4GALNT3, a paralog of B4GALNT4, display significantly reduced influenza virus replication during the early stages of infection 38 . This example illustrates that the complex regulation of this locus may affect multiple antiviral host pathways, warranting additional study in response to multiple infections.…”

microRNAs in viral acute respiratory infections: immune regulation, biomarkers, therapy, and vaccines

“…The inhibition of expression of the M1 protein of the type A IV is regulated by let-7c in A549 cells [60,61]. Low expression of miR-17-3p and miR-221 was found in human alveolar basal epithelial cells during IV infection [62].…”

Global Mapping of O-Glycosylation of Varicella Zoster Virus, Human Cytomegalovirus, and Epstein-Barr Virus

“…Information on site-specific O-glycosylation of virus and viral glycoproteins produced in different cellular systems could prove to be important because we predict that O-glycosylation may not only vary with respect to structures, but more importantly also vary considerably with respect to sites of O-glycan attachment. This is because the repertoire of polypeptide GalNAc transferases that controls the O-glycosylation capacity is cell-specific and may also be influenced by the viral infection itself, as evidenced by the induction of GalNAc-T3 by influenza A virus (18,89). We have recently developed a quantitative differential O-glycoproteomic strategy to address non-redundant contributions of individual GalNAc transferase isoforms to the O-glycosylation capacity of a cell (90), and this could be applied to address changes in viral O-glycosylation between clinical isolates or samples propagated in different cell types.…”