FoxP3 and the control of CD4+CD25+ regulatory T cell development and function.
The forkhead-family transcription factor FoxP3 has been implicated in the development and function of CD4+CD25+ regulatory T cells (Tregs). In the mouse, FoxP3 expression is both necessary and sufficient for generating Tregs, while FoxP3 expression has been shown to correlate with Treg function in humans. The laboratory is taking several approaches to better understand the role of this protein.
Structure/function analysis of FoxP3. We have shown that FoxP3 functions as a transcriptional repressor, targeting composite NF-AT/AP-1 sites in cytokine gene promoters. We have taken advantage of mutations in the FoxP3 gene found in human patients with IPEX (Immune dysfunction/Polyendocrinopathy/Enteropathy/X-linked) syndrome to study the function of FoxP3. Using these mutations, as well as deletions, we have mapped the region responsible for NF-AT inhibition to the amino terminus of FoxP3. We are currently examining the mechanism of FoxP3-mediated NF-AT inhibition, and the role of this amino terminal domain of FoxP3 in this process.
Regulation of Treg/Th17 differentiation by FoxP3. We have found that Foxp3 interacts with members of the retinoic receptor-like orphan receptor ROR) family of nuclear steroid receptors. This family includes RORgt, which has been shown to be critical for the differentiation of Th17 cells, a CD4 T cell subset implicated in several autoimmune diseases. We have shown that Foxp3 and RORgt physically interact, and that Foxp3 inhibits the ability of RORgt to activate the transcription of gene critical for Th17 differentiation. Our current work is designed to understand the physiological significance of this interaction in both normal immune homeostasis and in settings of autoimmune disease and infection.
Regulatory T cells and autoimmune diabetes. We have established a model of Type 1 diabetes where disease proceeds in spite of the presence of islet antigen-specific regulatory T cells in the infiltrated islets. We are now using this model to identify and characterize the factors present in the inflamed pancreas that abrogate the ability of antigen-specific Tregs to control the ongoing autoimmune response in these models.
TSLP and allergic inflammation
We are also studying a cytokine called thymic stromal lymphopoietin (TSLP). TSLP is an IL-7-like cytokine that is expressed by epithelial cells in the lung, skin, gut, and thymus. The TSLP receptor complex is a heterodimer comprised of the TSLPR and IL-7Ra. The receptor is expressed primarily on monocytes and myeloid-derived dendritic cells, as well as on B cells. Recent work has shown that TSLP treatment of human dendritic cells has several outcomes, including increased survival, upregulation of co-stimulatory molecules, and the production of the Th2-attracting chemokines CCL17 and CCL22. When T cells are primed on TSLP-treated DC, they produce inflammatory cytokines when restimulated (IL-4, -5, -13, and TNFa, but no IL-10). In support of its role in allergic inflammation, keratinocytes from patients with atopic dermatitis produce high levels of TSLP, while keratinocytes from normal individuals do not.
We have begun to model human allergic diseases using transgenic models that express TSLP at specific sites. Thus far we have models that express an inducible TSLP transgene in the skin, and models that express both constitutive and inducible TSLP transgene in the lung. In both cases the models develop the corresponding disease when the transgene is expressed-atopic dermatitis in the skin and asthma in the lung. The diseases developed by these models closely resemble their human counterparts. The approaches we are currently taking to further analyze these models are outlined below.
In our model, TSLP initiates the inflammatory cascade that leads to eventual disease. To identify and characterize the downstream mediators of TSLP-induced disease the TSLP transgenic models are being bred to a series of models carrying targeted mutations in genes upregulated in the target tissue.
We have established an acute model of TSLP-mediated airway inflammation using intranasal administration of TSLP (plus an antigen such as ovalbumin). Using this system we have begun to study the cellular interactions that are critical for TSLP function in the lung.
We have established collaborations to screen lung samples from patients with a variety of inflammatory lung diseases for TSLP expression.
We have shown that models that lack TSLP responses (both TSLP- and TSLPR-deficient models) lack the ability to mount effective Th2 responses. We have now found, using a contact hypersensitivity model, that TSLP is also involved in regulating Th1-type responses. We are currently investigating the underlying mechanisms at play in this system.
We have begun to examine the regulation of TSLP expression in epithelial cells. Interestingly, the cytokines that are at elevated levels in the TSLP transgenics, and also in humans with asthma or atopic dermatitis, induce TSLP expression. This suggests that TSLP not only initiates disease, but is also part of a feed-back loop to perpetuate disease. We have identified 3 regions upstream of the TSLP gene that appear to be involved in its regulation. We are currently characterizing these sequences.