The overall goal of this project is to understand the role of the endoplasmic reticulum (ER) in regulating T cell receptor (TCR) expression in developing lymphocytes. The TCR complex expressed by mature T cells is comprised of polymorphic subunits that recognize antigen (TCRa and b), together with invariant proteins that transduce signals (CD3gde and z). TCR expression in thymocytes differs from that of mature T cells in two important ways: 1. Immature thymocytes express alternative forms of the TCR (calnexin-CD3 complexes, which consist of the molecular chaperone calnexin associated with CD3ge subunits; and pre-TCR complexes, which are thought to consist of the newly described pre-Ta subunit associated with TCRb and CD3 subunits) and these forms can transduce signals that regulate rearrangement of the gene loci encoding the antigen-binding TCR subunits. 2. Immature thymocytes express only 1/10 as many complete TCR complexes (abgdezz) on the cell surface despite making equivalent amounts of each TCR subunit. Failure to maintain TCR expression at these low levels can adversely affect the outcome of selection events responsible for generating mature functional T-cells, and the immune system's ability to destroy invading pathogens and cancer cells. It is hypothesized that TCR expression in thymocytes is controlled post-translationally by changes in the processing of TCR subunits in the ER. Clearly, the ER plays a central role in TCR expression because the folding and assembly of nascent TCR subunits is not passive, rather it requires the assistance of resident ER proteins called chaperones. Thus, it is likely that the extent to which TCR subunits are assembled and released to the cell surface is controlled by those resident ER proteins. The central hypothesis will be tested by investigating the developmental control of three distinct forms of the TCR (calnexin-CD3 complexes, pre-TCR complexes, and complete TCRs).
The aims are to, in immature thymocytes: 1. Examine the molecular mechanism underlying escape of calnexin-CD3 complexes from the ER. 2. Clarify pre-TCR structure and examine the requirements for surface expression. 3. Examine the mechanistic basis for impaired assembly to complete TCRs. Analysis of TCR assembly in developing thymocytes should provide insight into how changes in the function of the ER can have profound effects on the type and quantity of TCR expressed and as a result has profound effects on immune cell development.
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