The IRE1α–XBP1s Arm of the Unfolded Protein Response Activates N-Glycosylation to Remodel the Subepithelial Basement Membrane in Paramyxovirus Infection

Abstract

Respiratory syncytial virus (RSV) causes severe lower respiratory tract infections (LRTI) associated with decreased pulmonary function, asthma, and allergy. Recently, we demonstrated that RSV induces the hexosamine biosynthetic pathway via the unfolded protein response (UPR), which is a pathway controlling protein glycosylation and secretion of the extracellular matrix (ECM). Because the presence of matrix metalloproteinases and matricellular growth factors (TGF) is associated with severe LRTI, we studied the effect of RSV on ECM remodeling and found that RSV enhances the deposition of fibronectin-rich ECM by small airway epithelial cells in a manner highly dependent on the inositol requiring kinase (IRE1α)–XBP1 arm of the UPR. To understand this effect comprehensively, we applied pharmacoproteomics to understand the effect of the UPR on N-glycosylation and ECM secretion in RSV infection. We observe that RSV induces N-glycosylation and the secretion of proteins related to ECM organization, secretion, or proteins integral to plasma membranes, such as integrins, laminins, collagens, and ECM-modifying enzymes, in an IRE1α–XBP1 dependent manner. Using a murine paramyxovirus model that activates the UPR in vivo, we validate the IRE1α–XBP1-dependent secretion of ECM to alveolar space. This study extends understanding of the IRE1α–XBP1 pathway in regulating N-glycosylation coupled to structural remodeling of the epithelial basement membrane in RSV infection.

Genomic targets of the IRE1-XBP1s pathway in mediating metabolic adaptation in epithelial plasticity

“…Although EMP is orchestrated by interactions of the IKK signaling with multiple intracellular phosphorylation cascades, maintenance of the EMT pathway is dependent on cellular survival and adaptation to ER stress through protein N-glycosylation ( 13 , 37 ). EMP-induced N-glycosylation is dependent on the abundance of intracellular uridine 5′-diphosphate-N-acetyl-d-glucosamine (UDP-GlcNAc), which is a rate-limiting substrate for N-glycan synthesis and subsequently N-glycosylation of asparagines on secreted proteins, enabling proper protein folding, transport to Golgi and secretion, relieving proteotoxicity ( 13 , 37 ). The details of how IRE1α-XBP1s pathway controls UPR-induced metabolic adaptations are incompletely understood.…”

“…Activation of the UPR involves expression of immediate-early homeostatic genes that prevent proteotoxicity via protein N-glycosylation ( 13 , 37 ). Our previous ATAC-Seq studies have shown that the majority of RSV-inducible genes are silent, yet maintained in an open chromatin configuration associated with H3K27Ac marks ( 16 , 29 ).…”

“…IRE1α alternatively splices a 26 nt fragment from XBP1u mRNA, whose translation forms the bZIP-class XBP1s transcription factor that translocates into the nucleus to interact with chromatin-bound transcriptional regulators. In addition to activating the SNAI1 and ZEB1 mesenchymal transcription factors, IRE1-XBP1s signaling shifts glucose flux from glycolysis to uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) production ( 11 , 13 ). UDP-GlcNAc is a rate limiting substrate for protein N-glycosylation; its increased production during the UPR enables native folding and secretion of basement membrane modifying enzymes involved in collagen crosslinking, elastin degradation and assembling a fibronectin-enriched matrix, remodeling the basement membrane ( 13 ).…”

“…In addition to activating the SNAI1 and ZEB1 mesenchymal transcription factors, IRE1-XBP1s signaling shifts glucose flux from glycolysis to uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) production ( 11 , 13 ). UDP-GlcNAc is a rate limiting substrate for protein N-glycosylation; its increased production during the UPR enables native folding and secretion of basement membrane modifying enzymes involved in collagen crosslinking, elastin degradation and assembling a fibronectin-enriched matrix, remodeling the basement membrane ( 13 ). The mechanistic details how IRE1α-XBP1s pathway activates N glycosylation is not fully understood.…”

Metatranscriptome Reveals Specific Immune and Microbial Signatures of Respiratory Syncytial Virus Infection in Children

“…Interestingly, enrichment of extracellular matrix-related genes was specific to RSV infection compared with both the non-RSV infection and control groups. Studies have indicated that RSV infection could potentiate changes in extracellular matrix dynamics, which might be linked to asthma development ( 22 – 24 ). RSV-infected human lung fibroblasts could induce hyaluronan-enriched extracellular matrix production, promoting the adhesion of mast cells, which have long been recognized as a type of critical cell in the development of allergic airway inflammation and asthma.…”