This is a competitive renewal of a previously funded application from an established investigator. T cell development and activation are dependent on multiple independent receptor signaling pathways. Since tyrosine kinases are known to play a critical role in receptor proximal signaling events, Dr. Berg previously set out to identify novel tyrosine kinases expressed in T cells. These efforts led to the cloning of Itk, a member of the Tec family of tyrosine kinases expressed only in T cells and NK cells. Btk, a close relative of Itk expressed in B cells and mast cells, has been shown to be the gene defective in human and mouse immunodeficiencies (X-linked agammaglobulinemia and XID, respectively). To examine the role of Itk in T cell signaling, Itk-deficient mice were generated and characterized. T cells from these mice are defective in TCR-induced calcium mobilization and cytokine production. A profound defect in positive selection in the thymus was also found. Using T cells from the Itk-deficient mice and retroviral gene transfer techniques, Dr. Berg proposes to define the protein domains and phosphorylation sites of Itk that are necessary for its role in TCR-induced calcium mobilization and cytokine production. She will characterize the biochemical defects in the Itk-deficient T cells, focusing on the transcription factors involved in IL-2 gene induction and the role of Itk in PLCgamma1 activation, and on CD28 signaling. Finally, she will address the role of Itk in T cell development by crossing the Itk-deficient mice to three different TCR transgenic lines, and then assess positive and negative selection in these mice. Together, these studies will provide important information on the role of Itk in both T cell development and T cell activation. As tyrosine kinases have been implicated in numerous human immunodeficiency diseases and cancers, and are involved in decisions leading to the proliferation versus differentiation in many cell lineages, these experiments will provide information relevant to an understanding of oncogenesis, autoimmunity, as well as genetic immunodeficiences.
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