A bifunctional FoxP3+ regulatory T cell subset converts to pro-inflammatory helper T cells

Researchers have described several functional subsets of CD4⁺ FoxP3⁺ regulatory T cells (T_REGS). These subsets differ in plasticity — their ability to convert into other T_H subtypes. For example, T_REGS that express RORγt promote pro-tumor immune function in colorectal cancer patients.

In Immunity, Sharma et al. identified another T_REG subset: FoxP3⁺ T_REGS that lose expression of Eos. These cells convert into a pro-inflammatory helper subtype. This subtype drives naïve CD8⁺ T cells to become potent effector cells.

Eos is a transcription factor in the Ikaros family. It acts as an obligate co-repressor: it pairs with FoxP3 to block FoxP3-repressed genes. Researchers had seen that T_REGS in inflammatory environments could turn pro-inflammatory without losing FoxP3. To find out why, Sharma et al. examined Eos expression in FoxP3⁺ T_REGS under inflammatory conditions.

How T_REGS Convert to an Inflammatory Phenotype

The researchers tracked acquired expression of IL-2, IL-17, and CD40L in the draining lymph nodes of a vaccination site. FoxP3⁺ T_REGS at distant lymph nodes did not gain this function. In the converted, inflammatory FoxP3⁺ T_REGS, Eos expression was quickly lost.

IL-6 was required for this loss of Eos. T_REGS in mice lacking IL-6 kept their Eos expression under the same conditions. But IL-6 alone was not enough. The conversion also needed:

• Interaction with MHC class II on activated dendritic cells
• Loss of Eos expression itself — T_REGS engineered to overexpress Eos did not convert

Not All T_REGS Convert Equally

Not all FoxP3⁺ T_REGS were equally likely to lose Eos and turn pro-inflammatory. Researchers tested thymic FoxP3⁺ T_REGS for Eos stability under cycloheximide treatment and found two distinct groups:

• CD38⁺CD69⁺CD103^– T_REGS were "Eos-labile" — they lost Eos within one hour of treatment.
• CD38^–CD69^–CD103⁺ T_REGS kept their Eos expression.

Other T_REG markers, including CD25 and CTLA-4, were expressed equally in both groups. So these standard markers cannot tell the two populations apart. When researchers sorted the T_REGS into CD38⁺CD103^– and CD38^–CD103⁺ subsets and transferred them into mice, only the CD38⁺CD103^– T_REGS lost Eos and gained CD40L and IL-2 expression after vaccination.

The Eos-labile T_REGS still showed normal suppressive function in several models. This included protection from colitis in a Rag-deficient CD45RB^HI effector-cell-driven autoimmune colitis model, and suppression of anti-CD3-driven T cell proliferation in vitro.

Development and Function of the Eos-Labile Subset

Researchers found the Eos-labile subset in the thymus as part of the natural T_REG repertoire. This led them to study the signals needed for its development. IL-6 was required again — this subset did not arise in IL-6^-/- mice. Epigenetic analysis of DNA methylation across these FoxP3⁺ T_REG subsets found distinct methylation patterns. Still, the subsets stayed much more closely related to each other than to FoxP3^– CD4⁺ T cells. Future studies need to pin down these epigenetic differences and which signals IL-6 controls.

The researchers also studied how the Eos-labile subset contributes to CD8⁺ priming in the vaccination model. Depleting T_REGS caused:

• Loss of CD8⁺ T cell proliferation and granzyme B expression
• Loss of CD86 upregulation on dendritic cells

Adding back just the Eos-labile subset — or IL-2 plus CD40-agonist antibodies — rescued these defects. The Eos-labile subset did not help reactivate memory CD4⁺ T cells, though. This suggests these cells have a specific role in priming naïve T cells. Despite having regulatory activity, they are critical for priming CD8⁺ T cell responses by supplying IL-2 and CD40L signals.

IDO Blocks the Conversion

Indoleamine 2,3-dioxygenase (IDO) blocked Eos downregulation and blocked the gain of IL-2, IL-17, and CD40L expression. In a mouse tumor vaccination model, blocking IDO was important for inducing FoxP3⁺ inflammatory T_REGS and generating anti-tumor effector CD8⁺ T cell responses.

IDO appears to block Eos downregulation partly by opposing the IL-6-STAT3 pathway. It does this through kynurenine-pathway metabolites, which activate the aryl hydrocarbon receptor (AhR). Different AhR ligands are known to regulate T_H17 cell induction differently than T_REG induction (Quintana et al.), and kynurenine is a T_REG-inducing AhR ligand (Mezrich et al.). The contrasting effects of IL-6 and IDO will matter for priming immune responses.

Overall, this study identified the mechanisms that turn on and block this newly defined Eos-labile T_REG subset. This subset keeps FoxP3 expression and typical T_REG suppressive activity, yet plays a key role in priming effector T cell immune responses. Future studies need to address how these cells balance regulatory and priming roles, and how this subset relates to the many other T_REG subsets already described.

An inherently bifunctional subset of foxp3(+) T helper cells is controlled by the transcription factor eos.  Sharma MD, Huang L, Choi JH, Lee EJ, Wilson JM, Lemos H, Pan F, Blazar BR, Pardoll DM, Mellor AL, Shi H, Munn DH. Immunity. 2013 May 23;38(5):998-1012. doi: 10.1016/j.immuni.2013.01.013. Epub 2013 May 16.

Eos, goddess of treg cell reprogramming. Rieder SA, Shevach EM. Immunity. 2013 May 23;38(5):849-50. doi: 10.1016/j.immuni.2013.05.001.

Control of T(reg) and T(H)17 cell differentiation by the aryl hydrocarbon receptor.  Quintana FJ, Basso AS, Iglesias AH, Korn T, Farez MF, Bettelli E, Caccamo M, Oukka M, Weiner HL. Nature. 2008 May 1;453(7191):65-71. doi: 10.1038/nature06880. Epub 2008 Mar 23.

An interaction between kynurenine and the aryl hydrocarbon receptor can generate regulatory T cells.  Mezrich JD, Fechner JH, Zhang X, Johnson BP, Burlingham WJ, Bradfield CA. J Immunol. 2010 Sep 15;185(6):3190-8. doi: 10.4049/jimmunol.0903670. Epub 2010 Aug 18.

Sanguine supplies research-grade human CD4+ T cells for studies like this.