Protein redistribution in TCR-directed NF-kB activation

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DESCRIPTION (provided by applicant): Antigen-stimulated T cell division and acquisition of effector functions are dependent upon signal transduction directed by the T cell receptor (TCR), and the consequent activation of a number of transcription factors that affect changes in gene expression. Transcription factors of the NF-?B family are of central importance in this process. The TCR-regulated NF-?B activation pathway will be investigated through a combination of molecular, biochemical, and highly advanced imaging techniques. The goal is to establish mechanistic relationships between 1) protein-protein interactions, 2) subcellular protein organization and 3) biochemical modification of cytoplasmic signal transducers in the TCR-directed NF-?B pathway. This goal will be accomplished via the execution of three specific aims: Aim1. To determine the mechanistic significance of localized protein-protein associations of signaling intermediates in TCR-directed activation of NF-?B. We will test the hypothesis that TCR-directed NF-?B signaling involves the orchestrated assembly and disassembly of defined protein complexes into discrete subcellular domains, with specific protein complexes playing a critical mechanistic role. Aim 2. To investigate, in individual cells, the mechanistic relationship between antigen signal strength, assembly of signal transduction complexes, and successful activation of the TCR-directed NF-?B signal transduction pathway. Experiments will be directed towards the evaluation of the hypothesis that antigen signals through the TCR are converted into "binary" outcomes of either no activation or full activation of effector functions. Furthermore, these functional outcomes are dependent on signal transduction decision events made at the single-cell level. Aim 3. To define the roles of phosphorylation and the MALT1 interaction domain in the TCR- mediated degradation and activation of BcHO. We will perform experiments to test the hypothesis that activation and proteolytic destruction of BcHO are mechanistically coupled, and that both processes are regulated by BcHO interaction with MALT1 and by PKC-dependent phosphorylation of BcHO. These studies will help establish the basic research foundation that could lead to the development of novel immuno-modulatory drugs, which would function via highly specific inhibition of antigen-receptor mediated activation of the NF-?B signaling cascade.