DOCK8 regulates fitness and function of regulatory T cells through modulation of IL-2 signaling
Akhilesh K. Singh, Ahmet Eken, David Hagin, Khushbu Komal, Gauri Bhise, Azima Shaji, Tanvi Arkatkar, Shaun W. Jackson, Estelle Bettelli, Troy R. Torgerson, Mohamed Oukka
Akhilesh K. Singh, Ahmet Eken, David Hagin, Khushbu Komal, Gauri Bhise, Azima Shaji, Tanvi Arkatkar, Shaun W. Jackson, Estelle Bettelli, Troy R. Torgerson, Mohamed Oukka
View: Text | PDF
Research Article Immunology

DOCK8 regulates fitness and function of regulatory T cells through modulation of IL-2 signaling

Abstract

Foxp3+ Tregs possess potent immunosuppressive activity, which is critical for maintaining immune homeostasis and self-tolerance. Defects in Treg development or function result in inadvertent immune activation and autoimmunity. Despite recent advances in Treg biology, we still do not completely understand the molecular and cellular mechanisms governing the development and suppressive function of these cells. Here, we have demonstrated an essential role of the dedicator of cytokinesis 8 (DOCK8), guanine nucleotide exchange factors required for cytoskeleton rearrangement, cell migration, and immune cell survival in controlling Treg fitness and their function. Treg-specific DOCK8 deletion led to spontaneous multiorgan inflammation in mice due to uncontrolled T cell activation and production of proinflammatory cytokines. In addition, we show that DOCK8-deficient Tregs are defective in competitive fitness and in vivo suppressive function. Furthermore, DOCK8 controls IL-2 signaling, crucial for maintenance and competitive fitness of Tregs, via a STAT5-dependent manner. Our study provides potentially novel insights into the essential function of DOCK8 in Tregs and immune regulation, and it explains the autoimmune manifestations associated with DOCK8 deficiency.

Authors

Akhilesh K. Singh, Ahmet Eken, David Hagin, Khushbu Komal, Gauri Bhise, Azima Shaji, Tanvi Arkatkar, Shaun W. Jackson, Estelle Bettelli, Troy R. Torgerson, Mohamed Oukka

×

Figure 5

Tregs require DOCK8 for optimal STAT5 activation and competitive fitness.

Options: View larger image (or click on image) Download as PowerPoint
Tregs require DOCK8 for optimal STAT5 activation and competitive fitness...
(A–C) Combined cells from spleen and LN were stimulated with indicated concentration of IL-2 for 15 minutes, and then cells were analyzed for pSTAT5 by flow cytometry. (A) FACS plot showing the pSTAT5, (B) histogram showing the mean fluorescent intensity (MFI) of pSTAT5, and (C) dose response analysis of pSTAT5 in response to IL-2 stimulation in CD45+CD4+YFP+ Tregs isolated from control or Foxp3CreDOCK8fl/fl mice. Results described in this figure are representative of 3 independent experiments with minimum of 2–3 mice per group. (D–H) DOCK8-deficient Tregs are defective in competitive fitness in the presence of WT Tregs. (D) Representative FACS plots showing the frequency of Tregs (CD45+CD4+Foxp3+) in thymus, spleen, and LN of Foxp3Cre/wt DOCK8wt/wt and Foxp3Cre/WT DOCK8fl/fl female mice. (E and G) The frequency (E) and absolute number (G) of Foxp3+YFP– Tregs (E, left panel) and Foxp3+YFP+ Tregs (E, right panel) in indicated organs. (F and H) Plot showing the MFI of Foxp3 (F) and CD25 (H) in thymus, spleen, and LN of Foxp3Cre/wt DOCK8wt/wt and Foxp3Cre/WT DOCK8fl/fl female mice. Data represents at least 3 independent experiments with 4–6 mice per group. (I and J) Combined cells from spleen and LN of female Foxp3Cre/WT DOCK8fl/fl mice were stimulated with 10 unit/ml of IL-2 for 15 minutes, and then cells were analyzed for pSTAT5 by flow cytometry. (I) FACS plot and (J) histogram showing the frequency of pSTAT5+ cells in response to IL-2 stimulation in CD45+CD4+CD25+YFP– (WT) and CD45+CD4+CD25+YFP+ (KO) Tregs. Results described in these figures are representative of 3 independent experiments with 3 mice per group. (K) Representative FACS plot showing the frequency of Tregs in the spleen of control and Foxp3CreDOCK8fl/fl mice after PBS or IL-2 complexes injections. (L) Histogram showing the proliferation of sorted Tregs from control and Foxp3CreDOCK8fl/fl mice after 12 days of culture in the presence of IL-2– and CD3/D28–coated beads. The data shown are the mean ± SD. Statistics were performed with Prism software by using t test. *P < 0.05, **P < 0.01,***P < 0.001.