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

Respiratory syncytial virus (RSV), a member of the genus <i>Orthopneumovirus</i>, is an etiological agent in infant lower respiratory tract infections (LRTIs) producing substantial global morbidity. Here, secretoglobin (<i>Scgb1a1</i>)-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 <i>Scgb1a1</i>-derived progenitors, a model of RSV post-viral disease (PVLD) was developed and studied in the presence or absence of RelA conditional knockout (CKO). Single-cell RNA sequencing (scRNA-seq) studies showed that RSV-PVLD induced a transition of atypical, differentiation-intermediate, alveolar type 2 (aAT2) cells characterized by tumor protein 63 (TRP63), aquaporin 3 (AQP3), and Itgβ4 expression, as well as changes in PDGFRβ mesenchyme. A single-cell trajectory analysis and lineage-tracing experiments using <i>Scgb1a1</i> CreER^TM X mTmG mice demonstrated that the <i>Scgb1a1+</i> populations were precursors to the aAT2 population. Mechanistically, we found that the formation of the aAT2 population was prevented by RelA CKO. A differential gene expression analysis revealed that RSV-PVLD coordinately upregulates nuclear receptor subfamily 1 group D (<i>Nr1d1/2</i>), clock and basic helix-loop-helix ARNT-like 1 (<i>Bmal</i>) genes both in the aAT2 cell and in its <i>Pdgfrα</i>+ mesenchymal niche in a RelA-dependent manner. A systematic analysis of intercellular epithelial-mesenchymal communication in the scRNA-seq data showed that the clock-dysregulated epithelial-mesenchymal niche produces aberrant <i>ANGPTL4</i> expression. ANGPTL4 upregulation was confirmed by the measurement of both its mRNA and protein. Moreover, ANGPTL4 is biologically active in the BALF of RSV-PVLD mice, inhibiting lipoprotein lipase activity. We conclude that RSV-PVLD is mediated, at least in part, by RelA signaling in <i>Scgb1a1</i>-derived epithelial progenitors, dysregulating ANGPTL4 signaling in an epithelial-mesenchymal niche, resulting in persistence of atypical alveolar epithelial cells with dysregulated of clock gene expression.