Dataset Information

Identification of a Distal Locus Enhancer Element That Controls Cell Type-Specific TNF and LTA Gene Expression in Human T Cells.

ABSTRACT: The human TNF/LT locus genes TNF, LTA, and LTB are expressed in a cell type-specific manner. In this study, we show that a highly conserved NFAT binding site within the distal noncoding element hHS-8 coordinately controls TNF and LTA gene expression in human T cells. Upon activation of primary human CD4+ T cells, hHS-8 and the TNF and LTA promoters display increased H3K27 acetylation and nuclease sensitivity and coordinate induction of TNF, LTA, and hHS-8 enhancer RNA transcription occurs. Functional analyses using CRISPR/dead(d)Cas9 targeting of the hHS-8-NFAT site in the human T cell line CEM demonstrate significant reduction of TNF and LTA mRNA synthesis and of RNA polymerase II recruitment to their promoters. These studies elucidate how a distal element regulates the inducible cell type-specific gene expression program of the human TNF/LT locus and provide an approach for modulation of TNF and LTA transcription in human disease using CRISPR/dCas9.

SUBMITTER: Jasenosky LD

PROVIDER: S-EPMC7576113 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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Identification of a Distal Locus Enhancer Element That Controls Cell Type-Specific TNF and LTA Gene Expression in Human T Cells.

Jasenosky Luke D LD Nambu Aya A Tsytsykova Alla V AV Ranjbar Shahin S Haridas Viraga V Kruidenier Laurens L Tough David F DF Goldfeld Anne E AE

Journal of immunology (Baltimore, Md. : 1950) 20200925 9

The human TNF/LT locus genes TNF, LTA, and LTB are expressed in a cell type-specific manner. In this study, we show that a highly conserved NFAT binding site within the distal noncoding element hHS-8 coordinately controls TNF and LTA gene expression in human T cells. Upon activation of primary human CD4+ T cells, hHS-8 and the TNF and LTA promoters display increased H3K27 acetylation and nuclease sensitivity and coordinate induct ...[more]

PMID: 32978279

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Project description:BackgroundGenome-wide association studies (GWAS) have identified more than 100 genetic loci associated with type 2 diabetes (T2D). However, the underlying biological mechanisms for many of these associations remain unknown. GWAS signals close to the glucokinase regulatory protein gene (GCKR) have been reported for lipid and glucose metabolism traits and the risk of T2D. We investigated the regulatory function of an intronic locus at GCKR represented by the lead single nucleotide polymorphism (SNP) rs780094.MethodsWe used ENCODE project histone modification and transcription factor binding data to determine the regulatory features of a GCKR intronic locus formed by the high linkage disequilibrium rs780094(C/T), rs780095(G/A), and rs780096(G/C) SNPs. Characterization of the transcriptional activity of this region was assessed by luciferase reporter assays in HepG2 cells and mouse primary hepatocytes. ChIP-qPCR was used to determine the levels of haplotype specific transcription factor binding and histone marks. A CRISPR-dCas9 transcriptional activator system and qPCR were used to activate the locus and measure GCKR expression, respectively. Differential haplotype expression was measured from human liver biopsies.ResultsThe ENCODE data suggest the existence of a liver-specific intragenic enhancer at the locus represented by s780094. We observed that FOXA2 increased the transcriptional activity of this region in a haplotype specific way (CGG > TAC; rs780094, rs780095, and rs780096). In addition, the CGG haplotype showed higher binding to FOXA2 and higher levels of the H3K27Ac histone mark. The epigenetic activation of this locus increased the expression of endogenous GCKR in HepG2 cells, confirming that GCKR is the direct target gene of the enhancer. Finally, we confirmed that the CGG haplotype exhibits higher levels of transcription in human liver.ConclusionsOur results demonstrate the existence of a liver-specific FOXA2-regulated transcriptional enhancer at an intronic T2D locus represented by rs780094, rs780095, and rs780096 SNPs that increases GCKR expression. Differential haplotype regulation suggests the existence of cis regulatory effects that may contribute to the associated traits at this locus.

Dataset Information

Identification of a Distal Locus Enhancer Element That Controls Cell Type-Specific TNF and LTA Gene Expression in Human T Cells.

Publications

Identification of a Distal Locus Enhancer Element That Controls Cell Type-Specific <i>TNF</i> and <i>LTA</i> Gene Expression in Human T Cells.

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