Abstract
The paramyxoviridae, respiratory syncytial virus (RSV), and murine respirovirus are enveloped, negative-sense RNA viruses that are the etiological agents of vertebrate lower respiratory tract infections (LRTIs). We observed that RSV infection in human small airway epithelial cells induced accumulation of glycosylated proteins within the endoplasmic reticulum (ER), increased glutamine-fructose-6-phosphate transaminases (GFPT1/2) and accumulation of uridine diphosphate (UDP)-N-acetylglucosamine, indicating activation of the hexosamine biosynthetic pathway (HBP). RSV infection induces rapid formation of spliced X-box binding protein 1 (XBP1s) and processing of activating transcription factor 6 (ATF6). Using pathway selective inhibitors and shRNA silencing, we find that the inositol-requiring enzyme (IRE1α)-XBP1 arm of the unfolded protein response (UPR) is required not only for activation of the HBP, but also for expression of mesenchymal transition (EMT) through the Snail family transcriptional repressor 1 (SNAI1), extracellular matrix (ECM)-remodeling proteins fibronectin (FN1), and matrix metalloproteinase 9 (MMP9). Probing RSV-induced open chromatin domains by ChIP, we find XBP1 binds and recruits RNA polymerase II to the IL6, SNAI1, and MMP9 promoters and the intragenic superenhancer of glutamine-fructose-6-phosphate transaminase 2 (GFPT2). The UPR is sustained through RSV by an autoregulatory loop where XBP1 enhances Pol II binding to its own promoter. Similarly, we investigated the effects of murine respirovirus infection on its natural host (mouse). Murine respirovirus induces mucosal growth factor response, EMT, and the indicators of ECM remodeling in an IRE1α-dependent manner, which persists after viral clearance. These data suggest that IRE1α-XBP1s arm of the UPR pathway is responsible for paramyxovirus-induced metabolic adaptation and mucosal remodeling via EMT and ECM secretion.
Highlights
The paramyxoviridae are species-adapted, enveloped negative-sense RNA viruses responsible for respiratory tract infections in vertebrates
To investigate the effect of respiratory syncytial virus (RSV) on uridine diphosphate (UDP)-GlcNAc metabolism, we focused our experiments on telomerase-immortalized hSAECs. hSAECs are derived from primary human small airway epithelial cells, maintain representative genomic and proteomic signatures [20], replicate RSV [37], produce pathogenic mucin- and T-helper-2 lymphocyte-activating cytokines that mediate disease [20], and most importantly, exhibit inducible cell state changes [22, 38, 39] without exhibiting cellular senescence
To confirm whether RSV infection activates the hexosamine biosynthetic pathway (HBP), we examined changes in the cellular N-glycosylated protein distribution. hSAECs were infected with sucrose-purified RSV [multiplicity of infection (MOI) =1] for 24 h, fixed, and stained for N-acetylglucosamino-peptides using fluorescein-conjugated wheat germ agglutinin (FITC-WGA)
Summary
The paramyxoviridae are species-adapted, enveloped negative-sense RNA viruses responsible for respiratory tract infections in vertebrates. Through an interconnected network of cell-surface and cytoplasmic pattern recognition receptors, RSV replication triggers a robust innate antiviral response consisting of secretion of cytokine [10, 11], interferon [12], and damage-associated patterns [13]. These factors act in paracrine manner to initiate leukocytic inflammation [13,14,15], mucous production [16], and reduce viral replication [17]
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