E2A/HEB MEDIATED TRANSCRIPTIONAL REGULATION IN T CELL DEVELOPMENT Abstract T lymphocyte development in the thymus gives rise to multiple lineages of T cells that are clearly separated from each other based on their distinct effector fates and associated immune functions. Invariant natural killer T (iNKT) cells represent a group of innate-like T cells produced in parallel with CD4 helper and CD8 killer lineage T cells that provide adaptive immunity. In contrast to adaptive T cells, which constitute the vast clonal diversity but remain mostly functionally naïve in circulation, iNKT cells have already acquired effector functions upon maturation in the thymus or arrival at the peripheral tissues. This unique feature endorses iNKT cells to play important roles in the early phase of immune defense, long before the elapsing window needed for clonal activation and expansion of antigen specific CD4 helper or CD8 killer T cells. Because iNKT cells use a semi- invariant T cell receptor (TCR) that can be stimulated with small lipid antigens to provide potent effector function, they have a great potential to be applied to cell-based immune therapy. However, due to lack of understanding of the regulatory mechanisms underpinning iNKT lineage development and homeostatic maintenance of the effector populations, therapeutic manipulation of iNKT cells in vivo or adoptive transfer of ex vivo expanded iNKT cells still remain in early phase of clinical trials. Our recent studies have revealed a critical role for E-protein transcription factors, encoded by the E2A and HEB genes, in driving iNKT lineage specification and expansion. Strikingly, we saw a dramatic enhancement of iNKT cell development and expansion upon deletion of the Id2 and Id3 genes, which encode inhibitors of E-proteins. The expansion of iNKT cells, induced by Id gene deletion, eventually drives their malignant transformation into iNKT lymphomas in young adult mice. These findings offer a new angle of approach to uncovering the developmental timing and regulatory pathways of iNKT lineage development and expansion. In this proposal we will leverage the mouse genetic tools established in our lab to define the separate roles of E proteins in iNKT lineage commitment and subsequent clonal expansion. iNKT cells are highly conserved between mice and humans in regards to their developmental origin, antigenic specificity, and functionality. The outcome of the proposed research will provide new strategies for controlled manipulation of iNKT cells that can be used in immune therapy, either alone or in combination with the adaptive T cells, without the risk of malignant transformation.