Protein phosphorylation is a key mechanism for transmitting growth signals into cells. In T lymphocytes, both the T cell antigen receptor (TCR) and the receptor for the growth factor interleukin-2 (IL-2) use multiple well known protein kinase cascades for signaling to the nucleus. Much less is known about the protein phosphatases that dephosphorylate and inactivate these kinases and their substrates. The PI has recently found that the hematopoietic tyrosine-specific phosphatase HePTP can negatively regulate TCR-induced activation of the IL-2 gene and proposes that the mechanism involves dephosphorylation of MAP kinases through a specific targeting mechanism. This grant focuses on HePTP, which is known to be altered in leukemias and other hematological disorders, and attempts to elucidate in detail how it acts on MAP kinases, which also are known to be altered in acute leukemias and to regulate a wide array of aspects of T cell development, proliferation and death.
Specific Aim I (Functional interaction between HePTP and MAP kinases in T cells) will address the PI's finding that HePTP forms complexes with the MAP kinases Erk1 Erk2, and p38alpha (but not with Jnk). He will test whether other MAP kinases bind, whether the complexes contain additional molecules, and how signals from the TCR or IL-2 receptor modify the association. He hypothesizes HePTP/Erk complexes dissociate in a phosphorylation-dependent manner allowing the required nuclear translocation of the MAP kinases.
Specific Aim 2 (Structural basis for HePTP-MAP kinase association) will take the analysis of Erk/p38 binding to HePTP to the atomic level and will culminate in the co-crystallization and resolution of the 3-dimensional structure of the complex.
This Aim will also refine the analysis of binding and its regulation by phosphorylation to the mechanistic level. Finally, Specific Aim 3 (Involvement of HePTP in T cell physiology) steps back to a much broader cell biology / T cell physiology level and examines the role and importance of HePTP in gene regulation in T cells, concentrating on cytokines, cell death-inducing surface proteins, and protooncogenes. Finally, the PI will use the DNA array technology to determine which genes HePTP expression will up- or down-regulate in a tetracycline-controlled inducible system. This study will thoroughly illuminate the biology of HePTP. The new information will be useful for the design of therapies to treat a variety of hematological diseases such as myelodysplastic syndrome, T cell leukemias and lymphomas.
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