Avian Foxp3 acts as a switch for IL-10 by directly binding the T3G motif: revealing a novel epigenetic modulation...

Foxp3 is the master transcription factor for regulatory T (Treg) cell differentiation and function, essential for immune homeostasis and tolerance. While its role is well-characterized in mammals, its functional mechanisms in avian species remain poorly understood.

We conducted comparative genomic and functional analyses of chicken Foxp3 (chFoxp3). DNA-binding specificity was determined using motif analysis and genome-wide scanning. An H9N2 avian influenza challenge model was used to assess Foxp3 expression dynamics and CD4⁺CD25⁺ T cell frequency in vivo. The transcriptional regulation of IL-10 by chFoxp3 was investigated through promoter binding and transactivation assays, with key domains mapped via mutagenesis.

Avian Foxp3 has evolved independently from its mammalian ortholog. Unlike human Foxp3, which binds the FKHM motif, chFoxp3 specifically recognizes a T3G microsatellite motif. Genome-wide scanning identified potential targets enriched in the FoxO signaling pathway, including IL-10. In an H9N2 infection model, Foxp3 expression and CD4⁺CD25⁺ T cell frequency exhibited biphasic dynamics. Mechanistically, chFoxp3, but not human FoxP3, directly binds the T3G motif in the IL-10 promoter to activate transcription. This transactivation function critically depends on the HNT linker within its Fork-head domain.

Our study reveals a novel mode of transcriptional regulation by chFoxp3 via T3G motif recognition, underscoring its functional diversification during evolution. These findings provide a foundation for understanding immune regulation in birds under both physiological and pathological conditions, such as avian influenza infection.