The aim of the project was to investigate the role of LCPTP (Leucocyte phosphotyrosine phosphatase) in T cell signalling and activation. The investigation was divided into three parts. In the first part, biochemical studies were used to identify possible LCPTP substrates/interacting proteins. Antibodies specific for LCPTP were used to characterise potential interactions of endogenous LCPTP protein in cells. Recombinant LCPTP proteins, including wildtype and a substrate-trap protein were also generated. These were used to show that LCPTP interacts with members of the MAP kinase family, specifically ERK1 and ERK2, suggesting that these proteins are major LCPTP substrates. In the second part, the effect of overexpressing LCPTP on T cell function was investigated. The Jurkat T cell line was transfected to produce stable cell lines expressing wildtype and mutant forms of LCPTP. LCPTP transfected cells were stimulated via the TCR and CD28 and monitored for cytokine production, surface marker expression and cell proliferation. It was shown that LCPTP inhibited IL-2 production and expression of the surface IL-2 receptor, CD25. LCPTP also protected cells from TCR induced cell death and promoted homotypic aggregation. Following on from the observations made in the Jurkat T cell line, the final part of the project was aimed at looking at the effects of LCPTP in primary T cells. LCPTP adenovirus constructs were made and used to infect both na?ve T cells as well as antigen differentiated Th1 and Th2 populations. Overexpressed LCPTP substrate-trap protein lead to increased levels of cellular phosphoERK. Increased levels of LCPTP resulted in changes in cytokine production in TCR stimulated cells relative to control cells. Cells overexpressing LCPTP had a higher basal level of proliferation. The results obtained support a role for LCPTP in regulating the early stages of T cell activation by changing MAP kinase activity.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:408406 |
| Date | January 2004 |
| Creators | Brodeur, Isabelle |
| Publisher | University College London (University of London) |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://discovery.ucl.ac.uk/1446578/ |
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