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 find that there are two critical regions of the IL-7 receptor alpha chain: 1)Box 1 binds Jak1 and initiates two pathways. One leads through an unknown kinase to serine phosphorylation of Bad and protects from cell death. The second leads to tyrosine phosphorylation of Y449 of the receptor itself.2)Phospho-Y449 activates Stat5, the pI3K pathway, and an unknown pathway that induces the bcl-2 gene and prevents Bax from translocating to mitochondriaOur initial studies of the IL-7 receptor were conducted in cell lines. We have now developed a retroviral infection of stem cells to examine the function of IL-7 receptor muteins in the development and survival of T cells in mice. We conclude that the same two regions, Box1 and Y449 also are required for thymocyte development, homeostasis, and TCRg rearrangement. We also observed that IL-7 receptor when ectopically expressed in hematopoietic stem cells induces myeloid proliferation. The intracellular domains of different cytokine receptors were exchanged with the IL-7 receptor intracellular domain to determine whether it delivered unique signals. We observed that the IL-9 receptor intracellular domain replaced that of IL-7 receptor for development of ab T cells but not rearrangement of the TCRg locus or development of gd T cells. 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 will also utilize phage display to identify the proteins that bind to the Y449 residue of the receptor.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.
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