This proposal will investigate the direct regulation CD4+ T lymphocyte function through the TLR2 and TLR4 signaling pathways. We have previously shown that TLR2 activation in T cells can drive the generation and effector function of Th17 cells. Our preliminary data indicates that TLR4 signaling additionally may function in a similar manner within T lymphocytes through the promotion of proliferation and survival. Furthermore, direct activation of the TLR2 or the TLR4 pathway in CD4+ T cells promotes the development of experimental autoimmune encephalomyelitis (EAE), the mouse model of multiple sclerosis. Importantly, CD4+ T cells were found to promote the pathogenesis of EAE through the activation of TLR2 by endogenously generated signals that are released as a byproduct of inflammation. Whether the same is true for endogenous TLR4 signaling has yet to be determined. Thus, we will first perform EAE using a model of WT and TLR4-deficient CD4+ T cell transfer in the absence of exogenous TLR4 stimuli to determine the potential importance of this pathway in driving autoimmune inflammation. Furthermore, we aim to ascertain the identity of such endogenous TLR signals capable of activating the TLR2 and TLR4 pathways in Th17 cells. Two candidate molecules have been identified, HMGB1 and biglycan, both of which stimulate TLR2 and TLR4 and are found in inflamed CNS tissue. We will also investigate is targeting these endogenous TLR ligands, rather than TLRs themselves, is a viable strategy for EAE/MS therapy. These studies will include directly neutralizing endogenous ligands through antibody treatment. Our in vitro observations have clearly demonstrated that TLR2 signaling promotes proliferation and the direct polarization of naive CD4+ T cells into the Th17 lineage. TLR4 signaling, on the other hand, had no such effect on lineage commitment but instead enhanced proliferation and survival of Th17 cells. These results collectively suggest that under sub optimal conditions, signaling through either TLR could enhance Th17 cytokine production and the persistence of this inflammatory subset. Thus, we will analyze the epigenetic, signaling, and transcriptional mechanisms TLRs utilize to regulate proliferation, survival, and the production of IL-17 in CD4+ T lymphocytes. Our laboratory has found that specific histone modifications enriched at the IL-17 locus allow permissive access of the transcriptional machinery during Th17 differentiation. Furthermore, TLR2 activation on Th17 cells enhanced the expression of Th17-related transcription factors such as ROR?t. Thus, we will determine if histone H3 acetylation and methylation events are amplified through TLR2 signaling during Th17 differentiation. Additionally, we aim to globally investigate the downstream targets of TLR2 and TLR4 signaling in CD4+ T cells through microarray analysis. Identification of novel signaling and transcriptional pathways governing T cell survival and cytokine production as a result of TLR signaling will provide new lines of experimentation that will allow the candidate to establish an independent laboratory, gain separation from the research interests of the current mentor, and aid in the application for R01 funding.
This proposal addresses the direct role of toll-like receptor signaling in T cells during the development of inflammation. Our previous research has shown that toll-like receptor activation in T lymphocytes can promote the development of a mouse model of multiple sclerosis. This proposal will directly examine the mechanisms for these findings with the overall goal being the development of new therapeutic strategies in the treatment of autoimmune disease such as multiple sclerosis. PROJECT NARRATIVE: This proposal addresses the direct role of toll-like receptor signaling in T cells during the development of inflammation. Our previous research has shown that toll-like receptor activation in T lymphocytes can promote the development of a mouse model of multiple sclerosis. This proposal will directly examine the mechanisms for these findings with the overall goal being the development of new therapeutic strategies in the treatment of autoimmune disease such as multiple sclerosis.