An amphiregulin reporter mouse enables transcriptional and clonal expansion analysis of reparative lung Treg cells [scRNA-Seq]

Treg cells play critical roles in tissue repair processes, certain of which are mediated by production of the growth factor amphiregulin (Areg). We developed a knock-in mouse model for detection of Areg production in live Treg cells, inserting a Thy1a (Thy1.1) construct into the endogenous Areg locus, such that for every transcript of Areg that is translated, a Thy1.1 transcript is also translated and trafficked separately to the cell surface. We used this mouse model to sort for Thy1.1- or Thy1.1+ (Areg-producing or -non-producing) lung Treg cells in models of lung damage (influenza A virus infection and bleomycin acute lung injury), and used them for assessment of gene expression and clonal expansion of these populations. AregThy1.1/Thy1.1 mice were administered saline (control), influenza A virus (IAV) (275 TCID50) or bleomycin (1 mg/kg). For experiment 1, timepoints used were 5 days post-instillation (dpi) for saline (control), 5 dpi for IAV, and 12 dpi for bleomycin. For experiment 2, timepoints used were 21 dpi for saline (control) and 21 dpi for bleomycin. Lung Treg cells were sorted for Thy1.1- (Areg-non-producing) and Thy1.1+ (Areg-producing) populations, then sorted populations were prepared for single cell RNA-seq using 10x Genomics protocols. GEX: gene expression; TCR: T cell receptor; HTO: hash tag oligos.