The USP7-STAT3-granzyme-Par-1 axis regulates allergic inflammation by promoting differentiation of IL-5-producing Th2 cells
Uncontrolled type 2 immunity by type 2 helper T (Th2) cells causes intractable allergic diseases; however, whether the interaction of CD4+ T cells shapes the pathophysiology of allergic diseases remains unclear. We identified a subset of Th2 cells that produced the serine proteases granzyme A and B early in differentiation. Granzymes cleave protease-activated receptor (Par)-1 and induce phosphorylation of p38 mitogen-activated protein kinase (MAPK), resulting in the enhanced production of IL-5 and IL-13 in both mouse and human Th2 cells. Ubiquitin-specific protease 7 (USP7) regulates IL-4-induced phosphorylation of STAT3, resulting in granzyme production during Th2 cell differentiation. Genetic deletion of Usp7 or Gzma and pharmacological blockade of granzyme B ameliorated allergic airway inflammation. Furthermore, PAR-1+ and granzyme+ Th2 cells were colocalized in nasal polyps from patients with eosinophilic chronic rhinosinusitis. Thus, the USP7-STAT3-granzymes-Par-1 pathway is a potential therapeutic target for intractable allergic diseases. Overall design: In this study, genomes were recovered from Th2 cells and sequenced; Th2 cell differentiation was performed in wild-type, Cre-ERT2 Usp7+/+, Cre-ERT2 Usp7fl/fl, Stat3fl/fl, and CD4-Cre Stat3fl/fl mice derived from Splenic naive CD4 T cells (CD44loCD62Lhi) were purified using a magnetic cell sorter (autoMACS; Miltenyi Biotec, Bergisch Gladbach, NRW, Germany). cultures of Th cells were performed as previously described. for Th2 cells, Naive CD4 T cells were cultured with IL-2 (25 U/mL), IL-4 (100 U/mL), anti-IFN-? mAb, and CD28 mAb (1 µg/mL) in the presence of TCRß mAb. For library preparation, libraries were prepared using Chromium Single Cell 3' Reagent Kits v3.1 according to the manufacturer's protocol (10X Genomics, Pleasanton, CA, USA). The prepared scRNA-seq libraries were sequenced using NovaSeq 6000 (Illumina) for 127 cycles (paired-end reads).