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

Gangqiang Guo; Huijing Wang; Xinyu Shi; Lele Ye; Kejing Yan; Zhiyuan Chen; Huidi Zhang; Zibing Jin; Xiangyang Xue

doi:10.3389/fcell.2020.00430

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

Front Cell Dev Biol. 2020 Jun 5:8:430. doi: 10.3389/fcell.2020.00430. eCollection 2020.

Authors

Gangqiang Guo^{1

2}, Huijing Wang³, Xinyu Shi², Lele Ye^{2

4}, Kejing Yan², Zhiyuan Chen², Huidi Zhang⁵, Zibing Jin⁶, Xiangyang Xue²

Affiliations

¹ School of Life Sciences and Technology, Tongji University, Shanghai, China.
² Department of Microbiology and Immunology, Institute of Molecular Virology and Immunology, Institute of Tropical Medicine, School of Basic Medical Sciences, Wenzhou Medical University, Wenzhou, China.
³ Kidney Disease Center, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, China.
⁴ Department of Gynecologic Oncology, Wenzhou Central Hospital, Wenzhou, China.
⁵ Department of Nephrology, The First Affiliated Hospital, Wenzhou Medical University, Wenzhou, China.
⁶ Laboratory for Stem Cell and Retinal Regeneration, Institute of Stem Cell Research, Division of Ophthalmic Genetics, The Eye Hospital, Wenzhou Medical University, Wenzhou, China.

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 Ψ, 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.

Keywords: 5-methylcytosine (m5C); CD4+ T cell; NSUN2; epigenetics; systemic lupus erythematosus.