Introduction: Bach2 is a transcriptional repressor that plays a critical role in regulating Th2-type immune response. However, the underlying molecular mechanisms remain unclear.
Methods: Bach2 flox/flox mice and Batf flox/flox mice were crossed with CD4-Cre TG mice to generate T cell-specific gene manipulation mice (Bach2 KO: Bach2 flox/flox × CD4-Cre TG, Batf KO: Batf flox/flox × CD4-Cre TG). Bach2 interaction protein(s) were identified by using an AlphaScreen with cell-free technology for protein synthesis.
Results: We screened Bach2 interaction protein(s), and Batf family transcription factors (Batf and Batf3) were identified by possible candidates. Binding of Bach2-Batf complex to the AP-1 consensus oligonucleotide was detected by an oligonucleotide precipitation assay. A ChIP-sequencing revealed that the major enriched motifs for Bach2-binding contain Batf and Irf4 binding motifs. The bindings of Batf and Irf4 to the Bach2 binding regions were increased in Bach2 KO effector CD4 T cells suggesting that Bach2-Batf complex inhibit the recruitment of the Batf-Irf4 complex. Furthermore, we found that Bach2 associates with the DNase I hypersensitive region (RHS6) at the control region of the Th2 cytokine gene locus. Increased bindings of Batf-Irf4 complex to the Bach2 binding regions and a rise of active enhancer mark of histone H3 (K27 acetylation level) were validated in Bach2 KO effector CD4 T cells. Moreover, the deletion of Batf gene improved the Th2-type lung inflammation in T cell-specific Bach2 KO mice.
Conclusion: Bach2-Batf interactions are required to prevent an excessive Th2 response and subsequent development of Th2-type lung inflammation.