T cell receptor and cytokine signal integration in CD8+ T cells is mediated by the protein Themis.

T cell homeostasis and functional responsiveness require signals from self-peptide-major histocompatibility complex (self-pMHC) and cytokines, but the mechanisms controlling this signal integration are unknown. Using a conditional deletion of the T cell lineage-specific protein Themis, we show that Themis is required for the maintenance of peripheral CD8+ T cells and for proliferative CD8+ T cell responses to low-affinity pMHC aided by cytokines. Themis-deficient peripheral T cells show a phenotype indicative of reduced tonic signaling from self-pMHC, strongly suggesting that Themis is a positive regulator of T cell receptor signal strength in response to low-affinity self-pMHC in peripheral T cells. Signals from low-affinity pMHC and cytokines synergistically induce phosphorylation of the kinase Akt, metabolic changes and c-Myc transcription factor induction in CD8+ T cells only in the presence of Themis. This function of Themis is mediated through Shp1 phosphatase, as peripheral Themis and Shp1 double deletion rescues the peripheral CD8+ T cell maintenance.

Themis-associated phosphatase activity controls signaling in T cell development.

Thymocyte-expressed molecule involved in selection (Themis) has been shown to be important for T cell selection by setting the threshold for positive versus negative selection. Themis interacts with the protein tyrosine phosphatase (PTP) Src-homology domain containing phosphatase-1 (Shp1), a negative regulator of the T cell receptor (TCR) signaling cascade. However, how Themis regulates Shp1 is still not clear. Here, using a very sensitive phosphatase assay on ex vivo thymocytes, we have found that Themis enhances Shp1 phosphatase activity by increasing its phosphorylation. This positive regulation of Shp1 activity by Themis is found in thymocytes, but not in peripheral T cells. Shp1 activity is modulated by different affinity peptide MHC ligand binding in thymocytes. Themis is also associated with phosphatase activity, due to its constitutive interaction with Shp1. In the absence of Shp1 in thymocytes, Themis interacts with Shp2, which leads to almost normal thymic development in Shp1 conditional knockout (cKO) mice. Double deletion of both Themis and Shp1 leads to a thymic phenotype similar to that of Themis KO. These findings demonstrate unequivocally that Themis positively regulates Shp1 phosphatase activity in TCR-mediated signaling in developing thymocytes.

Autoimmune responses and inflammation in type 2 diabetes.

Obesity-induced insulin resistance is one of the largest noncommunicable disease epidemics that we are facing at the moment. Changes in lifestyle and greater availability of low nutritional value, high caloric food has led to the highest rates of obesity in history. Obesity impacts the immune system and obesity-associated inflammation contributes to metabolic diseases, such as type 2 diabetes. Both the adaptive and the innate immune system play a role in the regulation of glycemic control, and there is a need to understand how metabolic imbalances drive disease pathogenesis. This review discusses the cell types, mediators, and pathways that contribute to immunologic-metabolic crosstalk and explores how the immune system might be targeted as a strategy to treat metabolic disease.

A Dual Inhibitor of Cdc7/Cdk9 Potently Suppresses T Cell Activation.

T cell activation is mediated by signaling pathways originating from the T cell receptor (TCR). Propagation of signals downstream of the TCR involves a cascade of numerous kinases, some of which have yet to be identified. Through a screening strategy that we have previously introduced, PHA-767491, an inhibitor of the kinases Cdc7 and Cdk9, was identified to impede TCR signaling. PHA-767491 suppressed several T cell activation phenomena, including the expression of activation markers, proliferation, and effector functions. We also observed a defect in TCR signaling pathways upon PHA-767491 treatment. Inhibition of Cdc7/Cdk9 impairs T cell responses, which could potentially be detrimental for the immune response to tumors, and also compromises the ability to resist infections. The Cdc7/Cdk9 inhibitor is a strong candidate as a cancer therapeutic, but its effect on the immune system poses a problem for clinical applications.

Identification of Mediators of T-cell Receptor Signaling via the Screening of Chemical Inhibitor Libraries.

The T-cell receptor (TCR) signaling pathway comprises a multitude of mediators that transmit signals upon the activation of the TCR. Different strategies have been proposed and implemented for the identification of new mediators of TCR signaling, which would improve the understanding of T-cell processes, including activation and thymic selection. We describe a screening assay that enables the identification of molecules that influence TCR signaling based on the activation of developing thymocytes. Strong TCR signals cause developing thymocytes to activate apoptotic machinery in a process known as negative selection. Through the application of kinase inhibitors, those with targets that affect TCR signaling are able to override the process of negative selection. The method detailed in this paper can be used to identify inhibitors of canonical kinases with established roles in the TCR signaling pathways and also inhibitors of new kinases yet to be established in the TCR signaling pathways. The screening strategy here can be applied to screens of higher throughput for the identification of novel druggable targets in TCR signaling.

Development of a screening strategy for new modulators of T cell receptor signaling and T cell activation.

Activation of the T cell receptor (TCR) leads to the generation of a network of signaling events critical to the developmental decision making and activation of T cells. Various experimental approaches continue to identify new signaling molecules, adaptor proteins, and other regulators of TCR signaling. We propose a screening strategy for the identification of small molecules affecting TCR signaling based on the uncoupling of TCR stimulation from cellular responses in developing thymocytes. We demonstrate that this strategy successfully identifies inhibitors of kinases already shown to act downstream of TCR engagement, as well as new inhibitors. The proposed strategy is easily scalable for high throughput screening and will contribute to the identification of new druggable targets in T cell activation.