Single-cell profiling uncovers regulatory programs of pathogenic Th2 cells in allergic asthma
Lung pathogenic T helper type 2 (pTh2) cells are important drivers of allergic asthma, but fundamental questions remain regarding their regulation and heterogeneity. The differentiation and effector functions of immune cells are tightly regulated by epigenetic processes. Histone deacetylase 1 (HDAC1) is an important epigenetic regulator of T cells, however, its role in pTh2 cells is yet to be determined. Here we investigate immune regulation in allergic asthma by single-cell RNA sequencing (scRNA-seq) in mice challenged with house dust mite, in the presence and absence of HDAC1 function. Our analyses reveal two distinct subsets of lung pTh2 cells: pathogenic effector Th2 (peTh2) and pathogenic Th2 tissue-resident memory (Th2 Trm) cells. Both pTh2 cell subsets are highly proinflammatory and exhibit distinct transcriptional and phenotypic signatures as compared with other lung Th subsets. Based on our scRNA-seq analysis, we identify conditions to generate pTh2 cells in vitro and confirm that these in vitro generated pTh2 cells have a similar transcriptional profile as lung peTh2 cells. Using our new in vitro model, we demonstrate that the p38 mitogen-activated protein kinase pathway is critical for interleukin-5 (IL-5) and IL-13 expression in pTh2 cells. Our data further underline the importance of HDAC1 in limiting the pathogenicity of lung and in vitro pTh2 cells and in the formation of lung Th2 Trm cells. In summary, we have generated novel insights into pTh2 cell biology and established a new in vitro model for investigating pTh2 cells that will be useful for discovering molecular mechanisms involved in pTh2-mediated allergic asthma. Overall design: To understand how pathogenic Th2 differentiation is established at a chromatin level, and how HDAC1 regulates this process, we isolated naive CD4+ T cells from HDAC1fl/fl and HDAC1fl/fl Cd4-Cre mice, and cultured these cells either under "conventional" Th2 conditions (anti-CD3/anti-CD28 with IL-4) or "pathogenic" Th2 conditons (anti-CD3/anti-CD28 with IL-4, anti-GITR agonist, TSLP).To have a time resolved chromatin accessibility profiling. the cells were harvested after 24 and 48 hours of culture and processed for assay for transposase-accessible chromatin using sequencing (ATAC-seq). For each condition, three experimental replicates were performed.