RelA Signaling in <em>Scgb1a1</em>+ Progenitors Mediates Lower Airway Epithelial Atypia in RSV-Induced Post Viral Lung Disease

Respiratory syncytial virus (RSV), a member of genus Orthopneumovirus, is an etiological agent in infant lower respiratory tract infections (LRTIs) producing substantial global morbidity. Here, secretoglobin (Scgb1a1)-derived progenitors play a primary role in triggering innate, inflammatory, and cell state transitions in response to RSV LRTIs. Whether RSV activation of innate signaling in this epithelial sentinel population leads to chronic airway disease is unknown. To understand the role of innate signaling in Scgb1a1-derived progenitors, a model of RSV-post viral disease (PVLD) was developed and studied in the presence or absence of RelA conditional knockout (CKO). In wild type mice, we observed persistence of atypical, differentiation intermediate, alveolar type 2 (aAT2) cells characterized by tumor protein 63 (Trp63), Aquaporin-3 (Aqp3) and Itgb4 expression. Mechanistically, we found the formation of aAT2 population was prevented by RelA CKO. Lineage tracing using Scgb1a1 CreERTM X mTmG mice demonstrated that the Scgb1a1+ populations were precursors to the aAT2 population. Single-cell RNA sequencing (scRNA-seq) showed that RSV-PVLD coordinately upregulates Nuclear Receptor Subfamily 1 Group D (Nr1d1/2), Clock and Basic Helix-Loop-Helix ARNT Like 1 (Bmal) both in the aAT2 cell and its Pdgfra+-mesenchymal niche in a RelA-dependent manner. Systematic analysis of intercellular epithelial-mesenchymal communication in the scRNA-Seq data showed that the clock-dysregulated epithelial-mesenchymal niche produces aberrant Angptl4 expression. We conclude that RSV-PVLD is mediated, at least in part, by RelA signaling in Scgb1a1-derived epithelial progenitors dysregulating an aAT2-PDGFRa epithelial-mesenchymal niche associated with persistent RSV replication, ANGPTL4 signaling and inflammatory clock expression.