Foxp3 is a transcription factor expressed by a subset of lymphocytes called regulatory T cells (Treg). Unlike conventional T cells that mediate immunity, Treg suppress inflammation and are required for immunologic tolerance. Foxp3 is sufficient to specify the Treg program, as ectopic expression of this factor in conventional T cells causes them to adopt regulatory function, including the capacity to prevent inflammatory bowel disease and organ transplant rejection. In order to understand how Foxp3 can re-program conventional T cells into Treg, we undertook a global analysis of mouse Foxp3 binding and chromatin remodeling. These studies incorporated genome-wide analysis of gene expression, Foxp3 binding, nucleosome positioning, and active vs. repressive histone modifications by chromatin immunoprecipitation (ChIP) paired with massively parallel deep sequencing using the ABI-SOLiD platform. In conventional CD4+ T cells, thousands of active genes exhibited a nucleosome positioned downstream of the transcriptional start site, but expression of Foxp3 resulted transcriptional repression associated with a more random nucleosome distribution, indicative of large scale chromatin remodeling by this factor. Indeed, our epigenomic analysis revealed a predominant epigenetic pattern in cells expressing Foxp3, with thousands of genes co-exhibiting a reduction in the acetylation of H2A and H2B (k5-12-15), an increase in acetylation at histone motifs targeted by the MYST/TIP60 co-activator (H4k5-8-12-16), and an increase in the Polycomb complex-mediated H3k27me3 mark. A subset of these genes also exhibited a decrease in the H3k4me3 mark associated with active transcription. These results suggest that Foxp3 can displace co-activators such as CBP/p300 and Trithorax methyltransferases from nucleosomes positioned at genes normally active in conventional T cells, and can recruit MYST/TIP60 and Polycomb complexes to change the epigenetic landscape at thousands of genes. These data indicate that Foxp3 acts as a ‘pioneer factor’ that can induce large-scale re-programming of the epigenome at nearly one quarter of the T cell genome.

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