Activation and repression of transcription of protein coding genes is key to all facets of biology. Over the past decade or so, the field has learned a lot about how writing and reading of the epigenetic code, alterations in chromosome conformation and distal enhancers contribute to this regulation. In this regard, we have examined how these elements contribute to activation and repression of the gene that encodes IFN-?, a critical cytokine for function of both innate and adaptive arms of the immune system. More recently the field has learned that much of the genome is transcribed in some cell lineage at some stage in development. Two newly discovered classes of RNA regulatory elements include long non-coding RNAs (lncRNA) and RNAs transcribed from distal enhancers (eRNAs). We identified a lncRNA adjacent to IFNG that activates IFNG transcription. We also identified eRNAs synthesized from IFNG distal enhancers in a Th1/Th2/Th17 lineage and developmental specific context. Here, we propose an integrated set of experiments to understand underlying principles by which lncRNAs contribute to both regulation of genes, IFNG, IL4/L13, IL17a/f, that encode signature cytokines that exemplify Th1/Th2/Th17 lineage commitment and genes encoding the 'master regulators' of this lineage commitment. We believe that examination of these classes of lncRNA, regulators of 'signature gene expression' and of 'master regulators', will identify underlying principles that can be integrated into a greater whole than studies targeting a single effector Th lineage. Our proposal will link functions of lncRNAs, eRNAs, and distal enhancer elements in a murine model system that permits analysis of the evolution of the genomic IFNG locus as T cells differentiate from the naive to effector to memory stage of development. An additional aspect of our studies will focus on genetic polymorphisms that increase risk to develop complex diseases. Genome-wide association studies in human populations have identified numerous single nucleotide polymorphisms (SNPs) associated with risk of developing complex diseases. The vast majority of these disease risk alleles are found in areas of the genome that do not code for proteins. Examples are the four SNPs within the TMEVPG1-IFNG-IL26-IL22 genomic region, three of which are associated with inflammatory bowel diseases. Our studies will focus on the function of these disease risk alleles. We propose that we have a unique understanding of this genomic region and the necessary expertise to undertake these studies. The hypothesis we will test is that presence of these SNPs and the associated co-inherited haplotype blocks alter distal enhancer function to change expression of adjacent protein-coding genes to impact disease. We propose an integrated series of experiments to explore this hypothesis. Not only will these studies increase our understanding of how genetic risk for IBD is conferred, diseases that affects 0.5-1% of the human population, but they will also serve as a general roadmap to address how disease risk alleles located in protein non-coding regions of the genome alter expression of key proteins contributing to disease pathogenesis.

Public Health Relevance

Two newly discovered general classes of regulatory elements include the long non-coding RNAs (lncRNA) and RNAs transcribed from distal enhancers (eRNAs). The central hypothesis of this proposal is that integration of contributions of lncRNAs and eRNAs with that which we already understand about regulation of protein-coding gene expression will help produce a more unifying understanding of activation and silencing of protein-coding genes. A corollary of this reasoning is that we can use this understanding to define how genetic polymorphisms that increase risk to develop complex diseases and are localized in the non-coding portion of the genome actually regulate gene expression and contribute to disease risk.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
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Cellular and Molecular Immunology - A Study Section (CMIA)
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Kelly, Halonna R
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Vanderbilt University Medical Center
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Gibbons, Hunter R; Shaginurova, Guzel; Kim, Laura C et al. (2018) Divergent lncRNA GATA3-AS1 Regulates GATA3 Transcription in T-Helper 2 Cells. Front Immunol 9:2512
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