IL-7 is a cytokine produced by non-lymphoid cells in the thymus and lymphoid organs. Signals from the IL-7 receptor are essential for normal thymocyte development and survival of T cells after leaving the thymus. Clinically, defects in genes for the components of IL-7 receptor are a common cause of human severe combined immunodeficiency disease (SCID). IL-7 is a promising therapeutic in treatment of AIDS, cancer and in immunization, whereas blocking the IL-7 pathway may be desirable in autoimmune diseases and lymphoid cancers. IL-7 protects lymphocytes from apoptosis and our lab has been investigating the intracellular mechanisms. We have shown that IL-7 withdrawal activates p38MAPK, which in turn activates NHE1 inducing intracellular alkalinization which activates the death protein Bax. We showed that Bax is a critical death mediator because deletion of Bax rescues T cell development in mice deficient in the IL-7 receptor. This rescue is incomplete and we sought additional death mediators among the Bcl-2 family. This led to identification of two other death proteins, Bax and Bim, which we show undergo posttranslational modification following IL-7 withdrawal.Withdrawal of IL-7 induces G1 arrest. We have evidence that this arrest occurs through p38MAPK phosphorylating Cdc25a which is required for S phase progression. We also have evidence that P38MAPK leads to increased stability of p27Kip1 which inhibits cdk2. This pathway to p27Kip1 stability appears to be a new mechanism of cell cycle regulation that does not involve the previously identified regulators Scp2 and Cks1. This new pathway may be critical in regulating lymphocyte, and lymphoma proliferation.We previously characterized the intracellular domain of IL-7 receptor and the regions required to induce survival and differentiation. In the past year we showed that, despite the unique requirement for IL-7 receptor, many of its functions can be duplicated by the intracellular domains of related receptors. One function was shown to be unique to the IL-7 receptor intracellular domain, that is the opening of chromatin of the TCR gamma locus, permitting rearrangement of this gene.To identify the missing components in the IL-7 signaling pathway we have developed proteomics methods consisting of 2D gels followed by mass spectroscopy - we have identified over 30 proteins and are evaluating their relevance. We previously focused on the effects of IL-7 on thymic development and are beginning to examine intracellular pathways by which IL-7 regulates homeostasis of peripheral T cells. Two signals are required for survival of peripheral T cells, one from the IL-7 receptor and the other is a low affinity self recognition by the antigen receptor. We are analyzing the intracellular interactions of these two pathways. IL-7 is critical for lymphocyte survival, but little is known about the regulation of its production. We are examining the types of cells that produce it, whether it is recognized in solution or is displayed on surfaces, and if the latter, characterize the binding mechanism. There is very little IL-7 protein made and it has not been possible to visualize it using standard immunohistochemical technique. We have use new recombineering methods (in collaboration with S.Warming and N.Copeland) to knock a non-secreted GFP reporter into the first exon of IL-7 and create transgenic mice. These mice allowed us to visualize IL-7 producing cells for the first time and show that the thymic epithelial cell has extensive dendritic branches containing IL-7 and contacting lymphocytes.
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