Gene transcription dynamics associated with cell lineage differentiation can be modeled on three sequential processes: First, a rapid and transient onset of activities of transcription factors (TFs) responsible for turning on genes associated with differentiated states; second, suppression of genes associated with the immediate precursor state or alternate cell fate choice; and third, long lasting induction of gene associated with differentiated states. The first draft of a compendium of transcriptomes of maturing innate ??TCR+ thymocyte subsets produced in conjunction with the Immunological Genome Project (ImmGen) matches the model and predicts that other innate lymphoid cells (ILCs) with shared function, but acting at unique anatomical locations, are controlled by regulatory networks with thematic commonality. To uncover the gene regulatory networks that specify critical innate immune function we propose to test the predicted regulatory modules controlling innate IL-17 production from ??T cells (T??17). IL-17 family of cytokines has emerged as the central effectors of inflammatory responses contributing to autoimmune disorders in humans. While the rules for IL-17 production by adaptive T cells are extensively studied how IL-17 production from specialized ILC subsets is regulated is unknown. T??17 cells are the primary source of IL-17 during bacterial infection and its capacity for IL-17 production is programmed in the thymus. The gene regulatory network controlling the programming was unknown. The architectural blueprint responsible for T??17 differentiation, once discovered, can be used as a guide to understand all effector ILC development. A comprehensive understanding of innate T effector lineage differentiation can be accomplished by three interconnected approaches: First, identification of the initial wave of TFs that define differentiated T??17 state and the progenitors in which the initial programming is evident; second, a systematic mapping of TF regulator occupancy of target T??17 genes; and third, perturbation of the network (gene KO mice and pathogen challenge) from the apex of the gene regulatory network followed by impact analyses to determine functional interconnectivity of gene modules within the network. Functional characterization of the primary gene nodes in the regulatory network specifying T??17 cell fate has so far revealed five essential transcription factors (TFs): SOX13, SOX4, ROR?t, TCF1 and LEF1, of which only one, ROR?t, was previously identified. Mice deficient in Sox13 or Sox4 have impaired T??17 differentiation. Mice lacking TCF1 generate T cells with hyper-production of IL-17 while LEF1 expression is biphasic, excluded from IL-17 innate effectors. The five TFs therefore constitute the core regulators of innate IL-17 production and they are embedded in other ILC effector programs, supporting the prediction that a common gene network blueprint generates ILC effectors and understanding their interconnected function will be central to potential targeted immunotherapies of inflammatory disorders.
IL-17 family of cytokines (IL-17A, F and IL-22) and factors that induce it (IL-1 and IL-23) have been shown to be critical drivers of autoimmunity as well as protective immunity, particularly at mucosal sites. Initial priming of immune responses to pathogenic challenges is performed by innate lymphoid cells (ILCs) with programmed capacity to rapidly secrete IL-17 and related cytokines. IL-17 from innate ?? T cells (T??17) mediates pulmonary fibrosis and neutrophilia and represents the primary source of early inflammatory response in the skin and lung against Mycobacterium tuberculosis, Klebsiella pneumoniae, Listeria monocytogenes and Escherichia coli. Genes that control IL-17 production in ILCs are predicted to be distinct from those involved in adaptive, conventional T cells that are responsible for the delayed, second wave of IL-17 production. The clonal antigen receptor signaling is the prototypic switch of adaptive effector T cell differentiation, but for most ILCs n the gut that lack classical antigen receptors, and even those equipped with invariant receptors, other regulatory molecular switches must be operational. In conjunction with the Immunological Genome Project we mapped the gene regulatory network predicted to control innate T??17 cell differentiation. The network is composed of factors that respond to evolutionarily conserved morphogen signaling and variations of the network are embedded across ILC subsets. In this grant proposal we will systematically map the hierarchy of functionally interconnected gene modules in the regulatory network that programs innate IL-17 production. This map of unparalleled scope and detail will guide ongoing searches for novel targets of immunotherapy to treat human diseases resulting from deregulated IL-17 production.
