Aberrant expression of USF2 in refractory rheumatoid arthritis and its regulation of proinflammatory cytokines in Th17 cells

IL-17–producing Th17 cells are implicated in the pathogenesis of rheumatoid arthritis (RA) and TNF-α, a proinflammatory cytokine in the rheumatoid joint, facilitates Th17 differentiation. Anti-TNF therapy ameliorates disease in many patients with rheumatoid arthritis (RA). However, a significant proportion of patients do not respond to this therapy. The impact of anti-TNF therapy on Th17 responses in RA is not well understood. We conducted high-throughput gene expression analysis of Th17-enriched CCR6+CXCR3−CD45RA−CD4+T (CCR6+T) cells isolated from anti-TNF–treated RA patients classified as responders or nonresponders to therapy. CCR6+T cells from responders and nonresponders had distinct gene expression profiles. Proinflammatory signaling was elevated in the CCR6+T cells of nonresponders, and pathogenic Th17 signature genes were up-regulated in these cells. Gene set enrichment analysis on these signature genes identified transcription factor USF2 as their upstream regulator, which was also increased in nonresponders. Importantly, short hairpin RNA targetingUSF2in pathogenic Th17 cells led to reduced expression of proinflammatory cytokines IL-17A, IFN-γ, IL-22, and granulocyte-macrophage colony-stimulating factor (GM-CSF) as well as transcription factor T-bet. Together, our results revealed inadequate suppression of Th17 responses by anti-TNF in nonresponders, and direct targeting of the USF2-signaling pathway may be a potential therapeutic approach in the anti-TNF refractory RA.

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Loss of the Pioneer Factor FOXA1 Results in Genome-wide Epigenetic Reprogramming and activation of Interferon-Response Genes including CD274/PD-L1

10.1101/2020.09.11.294041 ◽

2020 ◽

Author(s):

Wenhuo Hu ◽

Hironobu Yamashita ◽

Jenna Craig ◽

Vonn Walter ◽

Joshua I. Warrick ◽

...

Keyword(s):

Gene Expression ◽

Bladder Cancer ◽

Transcription Factor ◽

Epigenetic Modification ◽

Regulatory Elements ◽

Epigenetic Reprogramming ◽

Gene Set Enrichment Analysis ◽

Interferon Response ◽

Integrated Analysis ◽

The Impact

AbstractForkhead Box A1 (FOXA1) is a pioneer transcription factor critical in epigenetic regulation of chromatin and cell fate determination. Reduced FOXA1 expression is an independent predictor of poor overall survival in bladder cancer patients. However, the impact of FOXA1 loss on chromatin epigenetics in bladder cancer is unknown. Therefore, we determined the impact of FOXA1 knock out (KO) on epigenetic modification of chromatin and associated gene expression. We identified 8,230 differentially expressed genes following FOXA1 KO. Surprisingly, Gene Set Enrichment Analysis (GSEA) identified IFNɑ/ɣ gene expression signatures as enriched following FOXA1 KO. FOXA1 KO induced both increased and decreased numbers of histone 3 lysine 27 acetylation (H3K27ac) sites throughout the genome. As expected, the majority of differences in H3K27ac across genomic areas in FOXA1 KO cells is mapped to intergenic and intronic regions where enhancers reside. In addition, a subset of differential H3K27ac levels were also mapped to proximal promoters and within gene bodies. Integrated analysis of RNA/ChIP-seq data shows changes in gene expression that are mirrored by differences in H3K27ac. Motif analysis of DNA sequence enriched for H3K27ac identified significant increases in transcription factor binding motifs including the interferon sensitive response element (ISRE) and interferon response factors such as IRF1. Moreover, we identified increased H3K27ac of regulatory elements as being associated with several upregulated interferon sensitive genes (ISGs) in FOXA1 KO cells, including CD274/PD-L1. Western blotting and Q-RT-PCR confirmed upregulation of CD274/PD-L1 following FOXA1 KO. Analysis of TCGA data confirmed an inverse relationship between FOXA1 and CD274 in bladder cancer, as well as in other cancers. In summary, we provide evidence of widespread epigenetic reprogramming after FOXA1 KO in bladder cancer cells. Additionally, we provide evidence that FOXA1 KO-induced epigenetic changes contribute to activation of a global interferon-dominant expression signature, including the immune checkpoint target CD274/PD-L1 in a cancer cell-intrinsic manner.

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Molecular remission at T cell level in patients with rheumatoid arthritis

Scientific Reports ◽

10.1038/s41598-021-96300-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Jun Inamo ◽

Katsuya Suzuki ◽

Masaru Takeshita ◽

Yasushi Kondo ◽

Yuumi Okuzono ◽

...

Keyword(s):

Rheumatoid Arthritis ◽

T Cells ◽

T Cell ◽

Disease Control ◽

Expression Profiling ◽

Molecular Mechanisms ◽

Principal Component ◽

Cell Level ◽

Molecular Remission ◽

Signature Genes

AbstractWhile numerous disease-modifying anti-rheumatic drugs (DMARDs) have brought about a dramatic paradigm shift in the management of rheumatoid arthritis (RA), unmet needs remain, such as the small proportion of patients who achieve drug-free status. The aim of this study was to explore key molecules for remission at the T cell level, which are known to be deeply involved in RA pathogenesis, and investigate the disease course of patients who achieved molecular remission (MR). We enrolled a total of 46 patients with RA and 10 healthy controls (HCs). We performed gene expression profiling and selected remission signature genes in CD4+ T cells and CD8+ T cells from patients with RA using machine learning methods. In addition, we investigated the benefits of achieving MR on disease control. We identified 9 and 23 genes that were associated with clinical remission in CD4+ and CD8+ T cells, respectively. Principal component analysis (PCA) demonstrated that their expression profiling was similar to those in HCs. For the remission signature genes in CD4+ T cells, the PCA result was reproduced using a validation cohort, indicating the robustness of these genes. A trend toward better disease control was observed during 12 months of follow-up in patients treated with tocilizumab in deep MR compared with those in non-deep MR, although the difference was not significant. The current study will promote our understanding of the molecular mechanisms necessary to achieve deep remission during the management of RA.

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OP0021 IDENTIFICATION OF DIFFERENTIALLY EXPRESSED GENES IN EARLY RHEUMATOID ARTHRITIS PATIENTS RESPONDING TO TOCILIZUMAB

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2021-eular.3668 ◽

2021 ◽

Vol 80 (Suppl 1) ◽

pp. 12.2-12

Author(s):

I. Muller ◽

M. Verhoeven ◽

H. Gosselt ◽

M. Lin ◽

T. De Jong ◽

...

Keyword(s):

Rheumatoid Arthritis ◽

Gene Expression ◽

T Cells ◽

Gene Ontology ◽

Regulatory T Cells ◽

Early Rheumatoid Arthritis ◽

Whole Blood ◽

Differentially Expressed ◽

Rna Seq ◽

Double Blind

Background:Tocilizumab (TCZ) is a monoclonal antibody that binds to the interleukin 6 receptor (IL-6R), inhibiting IL-6R signal transduction to downstream inflammatory mediators. TCZ has shown to be effective as monotherapy in early rheumatoid arthritis (RA) patients (1). However, approximately one third of patients inadequately respond to therapy and the biological mechanisms underlying lack of efficacy for TCZ remain elusive (1). Here we report gene expression differences, in both whole blood and peripheral blood mononuclear cells (PBMC) RNA samples between early RA patients, categorized by clinical TCZ response (reaching DAS28 < 3.2 at 6 months). These findings could lead to identification of predictive biomarkers for TCZ response and improve RA treatment strategies.Objectives:To identify potential baseline gene expression markers for TCZ response in early RA patients using an RNA-sequencing approach.Methods:Two cohorts of RA patients were included and blood was collected at baseline, before initiating TCZ treatment (8 mg/kg every 4 weeks, intravenously). DAS28-ESR scores were calculated at baseline and clinical response to TCZ was defined as DAS28 < 3.2 at 6 months of treatment. In the first cohort (n=21 patients, previously treated with DMARDs), RNA-sequencing (RNA-seq) was performed on baseline whole blood PAXgene RNA (Illumina TruSeq mRNA Stranded) and differential gene expression (DGE) profiles were measured between responders (n=14) and non-responders (n=7). For external replication, in a second cohort (n=95 therapy-naïve patients receiving TCZ monotherapy), RNA-seq was conducted on baseline PBMC RNA (SMARTer Stranded Total RNA-Seq Kit, Takara Bio) from the 2-year, multicenter, double-blind, placebo-controlled, randomized U-Act-Early trial (ClinicalTrials.gov identifier: NCT01034137) and DGE was analyzed between 84 responders and 11 non-responders.Results:Whole blood DGE analysis showed two significantly higher expressed genes in TCZ non-responders (False Discovery Rate, FDR < 0.05): urotensin 2 (UTS2) and caveolin-1 (CAV1). Subsequent analysis of U-Act-Early PBMC DGE showed nine differentially expressed genes (FDR < 0.05) of which expression in clinical TCZ non-responders was significantly higher for eight genes (MTCOP12, ZNF774, UTS2, SLC4A1, FECH, IFIT1B, AHSP, and SPTB) and significantly lower for one gene (TND2P28M). Both analyses were corrected for baseline DAS28-ESR, age and gender. Expression of UTS2, with a proposed function in regulatory T-cells (2), was significantly higher in TCZ non-responders in both cohorts. Furthermore, gene ontology enrichment analysis revealed no distinct gene ontology or IL-6 related pathway(s) that were significantly different between TCZ-responders and non-responders.Conclusion:Several genes are differentially expressed at baseline between responders and non-responders to TCZ therapy at 6 months. Most notably, UTS2 expression is significantly higher in TCZ non-responders in both whole blood as well as PBMC cohorts. UTS2 could be a promising target for further analyses as a potential predictive biomarker for TCZ response in RA patients in combination with clinical parameters (3).References:[1]Bijlsma JWJ, Welsing PMJ, Woodworth TG, et al. Early rheumatoid arthritis treated with tocilizumab, methotrexate, or their combination (U-Act-Early): a multicentre, randomised, double-blind, double-dummy, strategy trial. Lancet. 2016;388(10042):343-55.[2]Bhairavabhotla R, Kim YC, Glass DD, et al. Transcriptome profiling of human FoxP3+ regulatory T cells. Human Immunology. 2016;77(2):201-13.[3]Gosselt HR, Verhoeven MMA, Bulatovic-Calasan M, et al. Complex machine-learning algorithms and multivariable logistic regression on par in the prediction of insufficient clinical response to methotrexate in rheumatoid arthritis. Journal of Personalized Medicine. 2021;11(1).Disclosure of Interests:None declared

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Modeling transcriptional regulation using gene regulatory networks based on multi-omics data sources

BMC Bioinformatics ◽

10.1186/s12859-021-04126-3 ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Neel Patel ◽

William S. Bush

Keyword(s):

Gene Expression ◽

Transcription Factor ◽

Transcriptional Regulation ◽

Gene Regulatory Networks ◽

Regulatory Networks ◽

Prediction Models ◽

Long Distance ◽

Chromatin Looping ◽

Gene Regulatory ◽

The Impact

Abstract Background Transcriptional regulation is complex, requiring multiple cis (local) and trans acting mechanisms working in concert to drive gene expression, with disruption of these processes linked to multiple diseases. Previous computational attempts to understand the influence of regulatory mechanisms on gene expression have used prediction models containing input features derived from cis regulatory factors. However, local chromatin looping and trans-acting mechanisms are known to also influence transcriptional regulation, and their inclusion may improve model accuracy and interpretation. In this study, we create a general model of transcription factor influence on gene expression by incorporating both cis and trans gene regulatory features. Results We describe a computational framework to model gene expression for GM12878 and K562 cell lines. This framework weights the impact of transcription factor-based regulatory data using multi-omics gene regulatory networks to account for both cis and trans acting mechanisms, and measures of the local chromatin context. These prediction models perform significantly better compared to models containing cis-regulatory features alone. Models that additionally integrate long distance chromatin interactions (or chromatin looping) between distal transcription factor binding regions and gene promoters also show improved accuracy. As a demonstration of their utility, effect estimates from these models were used to weight cis-regulatory rare variants for sequence kernel association test analyses of gene expression. Conclusions Our models generate refined effect estimates for the influence of individual transcription factors on gene expression, allowing characterization of their roles across the genome. This work also provides a framework for integrating multiple data types into a single model of transcriptional regulation.

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Analysis of chromatin organization and gene expression in T cells identifies functional genes for rheumatoid arthritis

10.1101/827923 ◽

2019 ◽

Author(s):

Jing Yang ◽

Amanda McGovern ◽

Paul Martin ◽

Kate Duffus ◽

Xiangyu Ge ◽

...

Keyword(s):

Rheumatoid Arthritis ◽

Gene Expression ◽

T Cells ◽

Complex Disease ◽

Target Genes ◽

Disease Risk ◽

Association Studies ◽

Dna Interaction ◽

Genome Wide Association Studies ◽

Causal Genes

AbstractGenome-wide association studies have identified genetic variation contributing to complex disease risk. However, assigning causal genes and mechanisms has been more challenging because disease-associated variants are often found in distal regulatory regions with cell-type specific behaviours. Here, we collect ATAC-seq, Hi-C, Capture Hi-C and nuclear RNA-seq data in stimulated CD4+ T-cells over 24 hours, to identify functional enhancers regulating gene expression. We characterise changes in DNA interaction and activity dynamics that correlate with changes gene expression, and find that the strongest correlations are observed within 200 kb of promoters. Using rheumatoid arthritis as an example of T-cell mediated disease, we demonstrate interactions of expression quantitative trait loci with target genes, and confirm assigned genes or show complex interactions for 20% of disease associated loci, including FOXO1, which we confirm using CRISPR/Cas9.

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The Functionalities and Clinical Significance of Tumor-Infiltrating Immune Cells in Esophageal Squamous Cell Carcinoma

BioMed Research International ◽

10.1155/2021/8635381 ◽

2021 ◽

Vol 2021 ◽

pp. 1-9

Author(s):

Jishuai Zhang ◽

Haifeng Wang ◽

Haitao Wu ◽

Guangliang Qiang

Keyword(s):

Gene Expression ◽

Squamous Cell Carcinoma ◽

T Cells ◽

Esophageal Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Natural Killer ◽

Clinical Significance ◽

Squamous Cell ◽

Immune Cells ◽

Gene Set Enrichment Analysis

Tumor-infiltrating immune cells have been implicated in the tumorigenesis and progression of esophageal squamous cell carcinoma (ESCC). However, the functionalities and clinical significance of immune cells remain largely unveiled. In this study, the gene expression data from the Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) were extracted. The relative infiltrating levels were estimated by single-sample gene set enrichment analysis. Some cytotoxic immune cells were attenuated, and resting cytotoxic immune cells were accumulated in ESCC. Remarkably, we also observed that infiltrating levels of macrophage M2 and resting natural killer (NK) cells were increased in nonresponders of CRT, and T cells that had anticancer activities such as activated memory CD4 and T helper 2 (Th2) cells were significantly reduced in ESCC tissues of the nonresponders. Moreover, the high infiltrations of the resting natural killer (NK) and dendritic cell (DC) were observed to result in a shorter overall survival in ESCC. Consistently, high expression of immune checkpoint genes, CTLA4 and HAVCR2, was associated with poor prognosis. Furthermore, STAT5B, a key transcription factor, as well as its target genes, involved in the regulation of T cells, was significantly downregulated in ESCC, especially subgroup I, indicating that downregulation of STAT5B might be associated with reduced T cell-mediated anticancer activity. In conclusion, the present study significantly improved our understanding of the regulatory roles of immune cells in ESCC.

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The Fate of Th17 Cells is Shaped by Epigenetic Modifications and Remodeled by the Tumor Microenvironment

International Journal of Molecular Sciences ◽

10.3390/ijms21051673 ◽

2020 ◽

Vol 21 (5) ◽

pp. 1673 ◽

Cited By ~ 3

Author(s):

Elodie Renaude ◽

Marie Kroemer ◽

Romain Loyon ◽

Delphine Binda ◽

Christophe Borg ◽

...

Keyword(s):

Dna Methylation ◽

T Cells ◽

Transcription Factor ◽

Tumor Microenvironment ◽

Th17 Cell ◽

Th17 Cells ◽

Epigenetic Modifications ◽

High Plasticity ◽

Th17 Cell Differentiation ◽

Promising Treatment

Th17 cells represent a subset of CD4+ T cells characterized by the master transcription factor RORγt and the production of IL-17. Epigenetic modifications such as post-translational histone modifications and DNA methylation play a key role in Th17 cell differentiation and high plasticity. Th17 cells are highly recruited in many types of cancer and can be associated with good or bad prognosis. Here, we will review the remodeling of the epigenome induced by the tumor microenvironment, which may explain Th17 cell predominance. We will also discuss the promising treatment perspectives of molecules targeting epigenetic enzymes to remodel a Th17-enriched tumor microenvironment.

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Requirement for Transcription Factor NFAT in Interleukin-2 Expression

Molecular and Cellular Biology ◽

10.1128/mcb.19.3.2300 ◽

1999 ◽

Vol 19 (3) ◽

pp. 2300-2307 ◽

Cited By ~ 122

Author(s):

Chi-Wing Chow ◽

Mercedes Rincón ◽

Roger J. Davis

Keyword(s):

Gene Expression ◽

T Cells ◽

Transcription Factor ◽

Nuclear Translocation ◽

Interleukin 2 ◽

Inhibitory Effect ◽

Dominant Negative ◽

Activated T Cells ◽

Nfat Activation ◽

Nfat Transcription Factor

ABSTRACT The nuclear factor of activated T cells (NFAT) transcription factor is implicated in expression of the cytokine interleukin-2 (IL-2). Binding sites for NFAT are located in the IL-2 promoter. Furthermore, pharmacological studies demonstrate that the drug cyclosporin A inhibits both NFAT activation and IL-2 expression. However, targeted disruption of the NFAT1 and NFAT2 genes in mice does not cause decreased IL-2 secretion. The role of NFAT in IL-2 gene expression is therefore unclear. Here we report the construction of a dominant-negative NFAT mutant (dnNFAT) that selectively inhibits NFAT-mediated gene expression. The inhibitory effect of dnNFAT is mediated by suppression of activation-induced nuclear translocation of NFAT. Expression of dnNFAT in cultured T cells caused inhibition of IL-2 promoter activity and decreased expression of IL-2 protein. Similarly, expression of dnNFAT in transgenic mice also caused decreased IL-2 gene expression. These data demonstrate that NFAT is a critical component of the signaling pathway that regulates IL-2 expression.

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Direct and widespread role for the nuclear receptor EcR in mediating the response to ecdysone in Drosophila

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1900343116 ◽

2019 ◽

Vol 116 (20) ◽

pp. 9893-9902 ◽

Cited By ~ 14

Author(s):

Christopher M. Uyehara ◽

Daniel J. McKay

Keyword(s):

Gene Expression ◽

Transcription Factor ◽

Nuclear Receptor ◽

Binding Sites ◽

Transcriptional Responses ◽

Enhancer Activity ◽

Developmental Transitions ◽

Experimental Systems ◽

The Impact

The ecdysone pathway was among the first experimental systems employed to study the impact of steroid hormones on the genome. In Drosophila and other insects, ecdysone coordinates developmental transitions, including wholesale transformation of the larva into the adult during metamorphosis. Like other hormones, ecdysone controls gene expression through a nuclear receptor, which functions as a ligand-dependent transcription factor. Although it is clear that ecdysone elicits distinct transcriptional responses within its different target tissues, the role of its receptor, EcR, in regulating target gene expression is incompletely understood. In particular, EcR initiates a cascade of transcription factor expression in response to ecdysone, making it unclear which ecdysone-responsive genes are direct EcR targets. Here, we use the larval-to-prepupal transition of developing wings to examine the role of EcR in gene regulation. Genome-wide DNA binding profiles reveal that EcR exhibits widespread binding across the genome, including at many canonical ecdysone response genes. However, the majority of its binding sites reside at genes with wing-specific functions. We also find that EcR binding is temporally dynamic, with thousands of binding sites changing over time. RNA-seq reveals that EcR acts as both a temporal gate to block precocious entry to the next developmental stage as well as a temporal trigger to promote the subsequent program. Finally, transgenic reporter analysis indicates that EcR regulates not only temporal changes in target enhancer activity but also spatial patterns. Together, these studies define EcR as a multipurpose, direct regulator of gene expression, greatly expanding its role in coordinating developmental transitions.

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Modulatory Effects ofAstragalusPolysaccharides on T-Cell Polarization in Mice with Polymicrobial Sepsis

Mediators of Inflammation ◽

10.1155/2015/826319 ◽

2015 ◽

Vol 2015 ◽

pp. 1-10 ◽

Cited By ~ 17

Author(s):

Yu-Chen Hou ◽

Jin-Ming Wu ◽

Ming-Yang Wang ◽

Ming-Hsun Wu ◽

Kuen-Yuan Chen ◽

...

Keyword(s):

T Cells ◽

Th17 Cells ◽

Cecal Ligation ◽

Cell Polarization ◽

Polymicrobial Sepsis ◽

Organ Injury ◽

Th2 Cells ◽

High Dose ◽

Th2 Response ◽

The Impact

Background.This study evaluated the impact of different doses ofAstragaluspolysaccharides (APS) on the functional status and phenotype of T cells during polymicrobial sepsis.Methods.On day 1 after cecal ligation and puncture, mice were treated with either saline, 100 (A100), 200 (A200), or 400 mg APS/kg body weight (BW) (A400) by an intraperitoneal injection daily for 4 days. All mice were sacrificed 5 days after the operation.Results.APS treatment reversed the sepsis-induced decrement in the T helper (Th) cell population, and the percentage of activated Th cells also increased in the spleen and Peyer’s patches. APS administration downregulated the percentages of circulating Th2 cells and regulatory T cells (Treg), and the percentage of Th17 cells in blood was upregulated in the A400 group. Weight loss and kidney injury were attenuated in the A100 and A200 groups but not in the A400 group at the end of the study.Conclusions.Treatments with 100 and 200 mg APS/kg BW reduced Treg populations and elicited a more-balanced Th1/Th2 response that consequently attenuated immunosuppression in polymicrobial sepsis. High-dose APS administration led to excessive responses of Th17 cells which may have adverse effects in sepsis-induced organ injury.

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