Dataset Information

Disease Activity-Associated Alteration of mRNA m5 C Methylation in CD4+ T Cells of Systemic Lupus Erythematosus.

ABSTRACT: Epigenetic processes including RNA methylation, post-translational modifications, and non-coding RNA expression have been associated with the heritable risks of systemic lupus erythematosus (SLE). In this study, we aimed to explore the dysregulated expression of 5-methylcytosine (m5C) in CD4+ T cells from patients with SLE and the potential function of affected mRNAs in SLE pathogenesis. mRNA methylation profiles were ascertained through chromatography-coupled triple quadrupole mass spectrometry in CD4+ T cells from two pools of patients with SLE exhibiting stable activity, two pools with moderate-to-major activity, and two pools of healthy controls (HCs). Simultaneously, mRNA methylation profiles and expression profiling were performed using RNA-Bis-Seq and RNA-Seq, respectively. Integrated mRNA methylation and mRNA expression bioinformatics analysis was comprehensively performed. mRNA methyltransferase NSUN2 expression was validated in CD4+ T cells from 27 patients with SLE and 28 HCs using real-time polymerase chain reaction and western blot analyses. Hypomethylated-mRNA profiles of NSUN2-knockdown HeLa cells and of CD4+ T cells of patients with SLE were jointly analyzed using bioinformatics. Eleven methylation modifications (including elevated Am, 3'OMeA, m1A, and m6A and decreased ?, m3C, m1G, m5U, and t6A levels) were detected in CD4+ T cells of patients with SLE. Additionally, decreased m5C levels, albeit increased number of m5C-containing mRNAs, were observed in CD4+ T cells of patients with SLE compared with that in CD4+ T cells of HCs. m5C site distribution in mRNA transcripts was highly conserved and enriched in mRNA translation initiation sites. In particular, hypermethylated m5C or/and significantly up-regulated genes in SLE were significantly involved in immune-related and inflammatory pathways, including immune system, cytokine signaling pathway, and interferon signaling. Compared to that in HCs, NSUN2 expression was significantly lower in SLE CD4+ T cells. Notably, hypomethylated m5C genes in SLE and in NSUN2-knockdown HeLa cells revealed linkage between eukaryotic translation elongation and termination, and mRNA metabolism. Our study identified novel aberrant m5C mRNAs relevant to critical immune pathways in CD4+ T cells from patients with SLE. These data provide valuable perspectives for future studies of the multifunctionality and post-transcriptional significance of mRNA m5C modification in SLE.

SUBMITTER: Guo G

PROVIDER: S-EPMC7291606 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Disease Activity-Associated Alteration of mRNA m5 C Methylation in CD4+ T Cells of Systemic Lupus Erythematosus.

Guo Gangqiang G Wang Huijing H Shi Xinyu X Ye Lele L Yan Kejing K Chen Zhiyuan Z Zhang Huidi H Jin Zibing Z Xue Xiangyang X

Frontiers in cell and developmental biology 20200605

Epigenetic processes including RNA methylation, post-translational modifications, and non-coding RNA expression have been associated with the heritable risks of systemic lupus erythematosus (SLE). In this study, we aimed to explore the dysregulated expression of 5-methylcytosine (m5C) in CD4+ T cells from patients with SLE and the potential function of affected mRNAs in SLE pathogenesis. mRNA methylation profiles were ascertained through chromatography-coupled triple quadru ...[more]

PMID: 32582707

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Project description:The emerging epitranscriptome plays an essential role in autoimmune disease. As a novel mRNA modification, N4-acetylcytidine (ac4C) could promote mRNA stability and translational efficiency. However, whether epigenetic mechanisms of RNA ac4C modification are involved in systemic lupus erythematosus (SLE) remains unclear. Herein, we detected eleven modifications in CD4+ T cells of SLE patients using mass spectrometry (LC-MS/MS). Furthermore, using samples from four CD4+ T cell pools, we identified lower modification of ac4C mRNA in SLE patients as compared to that in healthy controls (HCs). Meanwhile, significantly lower mRNA acetyltransferase NAT10 expression was detected in lupus CD4+ T cells by RT-qPCR. We then illustrated the transcriptome-wide ac4C profile in CD4+ T cells of SLE patients by ac4C-RIP-Seq and found ac4C distribution in mRNA transcripts to be highly conserved and enriched in mRNA coding sequence regions. Using bioinformatics analysis, the 3879 and 4073 ac4C hyper-acetylated and hypoacetylated peaks found in SLE samples, respectively, were found to be significantly involved in SLE-related function enrichments, including multiple metabolic and transcription-related processes, ROS-induced cellular signaling, apoptosis signaling, and NF-κB signaling. Moreover, we demonstrated the ac4C-modified regulatory network of gene biological functions in lupus CD4+ T cells. Notably, we determined that the 26 upregulated genes with hyperacetylation played essential roles in autoimmune diseases and disease-related processes. Additionally, the unique ac4C-related transcripts, including USP18, GPX1, and RGL1, regulate mRNA catabolic processes and translational initiation. Our study identified novel dysregulated ac4C mRNAs associated with critical immune and inflammatory responses, that have translational potential in lupus CD4+ T cells. Hence, our findings reveal transcriptional significance and potential therapeutic targets of mRNA ac4C modifications in SLE pathogenesis.

Project description:Objective: The mechanism of CD4+ T-cell dysfunction in systemic lupus erythematosus (SLE) has not been fully understood. Increasing evidence show that long noncoding RNAs (lncRNAs) can regulate immune responses and take part in some autoimmune diseases, while little is known about the lncRNA expression and function in CD4+ T of SLE. Here, we aimed to detect the expression profile of lncRNAs in lupus CD4+ T cells and explore the mechanism that how lincRNA00892 in CD4+ T cells is involved in the pathogenesis of SLE. Methods: The expression profiles of lncRNAs and mRNAs in CD4+ T cells from SLE patients and healthy controls were detected by microarray. LincRNA00892 and CD40L were chosen for validation by quantitative real-time PCR (qRT-PCR). Coexpression network was conducted to predict the potential target genes of lincRNA00892. Then lincRNA00892 was overexpressed in normal CD4+ T cells via lentivirus transfection. The expression of lincRNA00892 was detected by qRT-PCR. The expression of CD40L was detected by qRT-PCR, western blotting, and flow cytometry, respectively. The expression of CD69 and CD23 was measured by flow cytometry. The secretion of IgG was determined by enzyme-linked immunosorbent assay (ELISA). The proteins targeted by lincRNA00892 were measured by RNA pulldown and subsequent mass spectrometry (MS). The interaction between heterogeneous nuclear ribonucleoprotein K (hnRNP K) and lincRNA00892 or CD40L was detected by RNA immunoprecipitation (RIP) assay. Results: A total of 1887 lncRNAs and 3375 mRNAs were found to be aberrantly expressed in CD4+ T cells of SLE patients compared to healthy controls. LincRNA00892 and CD40L were confirmed to be upregulated in CD4+ T cells of SLE patients by qRT-PCR. The lncRNA-mRNA coexpression network analysis indicated that CD40L was a potential target of lincRNA00892. Overexpression of lincRNA00892 enhanced CD40L protein levels while exerting little influence on CD40L mRNA levels in CD4+ T cells. In addition, lincRNA00892 could induce the activation of CD4+ T cells. Furthermore, lincRNA00892 led to the activation of B cells and subsequent secretion of IgG in a CD4+ T-cell-dependent manner. Finally, hnRNP K was found to be among the proteins pulled down by lincRNA00892, and hnRNP K could bind to lincRNA00892 or CD40L directly. Conclusion: Our results showed that the lncRNA expression profile was altered in CD4+ T cells of SLE. LincRNA00892 possibly contributed to the pathogenesis of SLE by targeting hnRNP K and subsequently upregulating CD40L expression to activate CD4+ T and B cells. These provided us a potential target for further mechanistic studies of SLE pathogenesis.

Dataset Information

Disease Activity-Associated Alteration of mRNA m5 C Methylation in CD4+ T Cells of Systemic Lupus Erythematosus.

Publications

Disease Activity-Associated Alteration of mRNA m<sup>5</sup> C Methylation in CD4<sup>+</sup> T Cells of Systemic Lupus Erythematosus.

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