Research Article|Articles in Press

Transcriptome-wide assessment of N6-methyladenosine modification identifies different gene expression and infection-associated pathways in Treponema pallidum-infected macrophage

Published:February 18, 2023DOI:


      • A global N6-methyladenosine (m6A) RNA modification pattern in Tp-infected macrophages was systematically dissected.
      • Aberrant genes with m6A-modification from these macrophages were enriched in infection-related pathways.
      • RNA-seq revealed function confusion of Tp-infected macrophage, which, together with MeRIP, displayed particular patterns.



      Treponema pallidum (Tp) is a widespread and destructive pathogen that leads to syphilis. As the acknowledged executor of host immunity, macrophage plays vital roles in combating the invasion and migration of Tp. However, the mechanisms of these processes are largely unknown, especially the critical driver genes and associated modifications.


      We aimed to systematically dissect the global N6-methyladenosine (m6A) RNA modification patterns in Tp-infected macrophages.


      The RNA of Tp-infected/non-infected macrophage was extracted, followed by mRNA sequencing and methylated RNA immunoprecipitation (MeRIP) sequencing. Bioinformatics analysis was executed by m6A peaks and motifs identification, Gene ontology and signaling pathways analysis of differentially expressed genes, and comprehensive comparison. The m6A levels were measured by RNA Methylation Assay, and m6A modified genes were determined by qPCR.


      Totally, 2623 unique and 3509 common m6A peaks were proved along with related transcripts in Tp-infected macrophages. The common m6A-related genes were enriched in the signals of oxidative stress, cell differentiation, and angiogenesis, while unique genes in those of metabolism, inflammation, and infection. And differentially expressed transcripts revealed various biological processes and pathways associated with catabolic and infection. They also experienced comprehensive analysis due to hyper-/hypo-methylation. And the m6A level of macrophage was elevated, along with qPCR validation of specific genes.


      With a particular m6A transcriptome-wide map, our study provides unprecedented insights into the RNA modification of macrophage stimulated by Tp in vitro, which partially differs from other infections and may provide clues to explore the immune process for syphilis.

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        • Hook 3rd, E.W.
        Lancet. 2017; 389: 1550-1557
        • Peeling R.W.
        • Mabey D.
        • Kamb M.L.
        • Chen X.S.
        • Radolf J.D.
        • Benzaken A.S.
        Nat. Rev. Dis. Prim. 2017; 317073
        • Peeling R.W.
        • Hook 3rd, E.W.
        The pathogenesis of syphilis: the great mimicker, revisited.
        J. Pathol. 2006; 208: 224-232
        • Radolf J.D.
        • Deka R.K.
        • Anand A.
        • Šmajs D.
        • Norgard M.V.
        • Yang X.F.
        Treponema pallidum, the syphilis spirochete: making a living as a stealth pathogen.
        Nat. Rev. Microbiol. 2016; 14 (744-59)
        • Li J.H.
        • Guo H.
        • Gao X.H.
        • Chen H.D.
        Multiple skin ulcers from malignant syphilis.
        Lancet. 2015; 386: 1564
        • Forrestel A.K.
        • Kovarik C.L.
        • Katz K.A.
        Sexually acquired syphilis: Historical aspects, microbiology, epidemiology, and clinical manifestations.
        J. Am. Acad. Dermatol. 2020; 82: 1-14
        • Cruz A.R.
        • Ramirez L.G.
        • Zuluaga A.V.
        • et al.
        Immune evasion and recognition of the syphilis spirochete in blood and skin of secondary syphilis patients: two immunologically distinct compartments.
        PLOS Negl. Trop. Dis. 2012; 6e1717
        • Carlson J.A.
        • Dabiri G.
        • Cribier B.
        • Sell S.
        The immunopathobiology of syphilis: the manifestations and course of syphilis are determined by the level of delayed-type hypersensitivity.
        Am. J. Dermatopathol. 2011; 33: 433-460
        • Baker-Zander S.A.
        • Lukehart S.A.
        Macrophage-mediated killing of opsonized Treponema pallidum.
        J. Infect. Dis. 1992; 165: 69-74
        • Huang T.
        • Yang J.
        • Zhang J.
        • et al.
        MicroRNA-101-3p downregulates TLR2 expression, leading to reduction in cytokine production by treponema pallidum-stimulated macrophages.
        J. Investig. Dermatol. 2020; 140 (e1): 1566-1575
        • Xu S.L.
        • Lin Y.
        • Liu W.
        • et al.
        The P2×7 receptor mediates NLRP3-dependent IL-1β secretion and promotes phagocytosis in the macrophage response to Treponema pallidum.
        Int. Immunopharmacol. 2020; 82106344
        • Xu S.L.
        • Lin Y.
        • Zhu X.Z.
        • et al.
        Autophagy promotes phagocytosis and clearance of Treponema pallidum via the NLRP3 inflammasome in macrophages.
        J. Eur. Acad. Dermatol. Venereol. 2020; 34: 2111-2119
        • Zhang Q.
        • Cao X.
        Epigenetic regulation of the innate immune response to infection.
        Nat. Rev. Immunol. 2019; 19 (417-32)
        • Geula S.
        • Moshitch-Moshkovitz S.
        • Dominissini D.
        • et al.
        Stem cells. m6A mRNA methylation facilitates resolution of naïve pluripotency toward differentiation.
        Science. 2015; 347: 1002-1006
        • Deng X.
        • Su R.
        • Weng H.
        • Huang H.
        • Li Z.
        • Chen J.
        RNA N(6)-methyladenosine modification in cancers: current status and perspectives.
        Cell Res. 2018; 28 (507-17)
        • Frye M.
        • Harada B.T.
        • Behm M.
        • He C.
        RNA modifications modulate gene expression during development.
        Science. 2018; 361: 1346-1349
        • Liu J.
        • Dou X.
        • Chen C.
        • et al.
        N (6)-methyladenosine of chromosome-associated regulatory RNA regulates chromatin state and transcription.
        Science. 2020; 367: 580-586
        • Du J.
        • Liao W.
        • Liu W.
        • et al.
        N(6)-adenosine methylation of Socs1 mRNA is required to sustain the negative feedback control of macrophage activation.
        Dev. Cell. 2020; 55: 737-753
        • Wang X.
        • Ji Y.
        • Feng P.
        • et al.
        The m6A reader IGF2BP2 regulates macrophage phenotypic activation and inflammatory diseases by stabilizing TSC1 and PPARγ.
        Adv. Sci. 2021; 82100209
        • Li N.
        • Hui H.
        • Bray B.
        • et al.
        METTL3 regulates viral m6A RNA modification and host cell innate immune responses during SARS-CoV-2 infection.
        Cell Rep. 2021; 35109091
        • Liu Y.
        • You Y.
        • Lu Z.
        • et al.
        N (6)-methyladenosine RNA modification-mediated cellular metabolism rewiring inhibits viral replication.
        Science. 2019; 365: 1171-1176
        • Wang X.
        • Li Y.
        • Chen W.
        • et al.
        Transcriptome-wide reprogramming of N(6)-methyladenosine modification by the mouse microbiome.
        Cell Res. 2019; 29 (167-70)
        • Lukehart S.A.
        • Marra C.M.
        Isolation and laboratory maintenance of Treponema pallidum.
        Curr. Protoc. Microbiol. 2007; (Chapter 12: Unit 12A.1)
        • Chen S.
        • Zhou Y.
        • Chen Y.
        • Gu J.
        fastp: an ultra-fast all-in-one FASTQ preprocessor.
        Bioinformatics. 2018; 34 (i884-i90)
        • Li H.
        • Durbin R.
        Fast and accurate short read alignment with Burrows-Wheeler transform.
        Bioinformatics. 2009; 25: 1754-1760
        • Meng J.
        • Lu Z.
        • Liu H.
        • et al.
        A protocol for RNA methylation differential analysis with MeRIP-Seq data and exomePeak R/Bioconductor package.
        Methods. 2014; 69: 274-281
        • Heinz S.
        • Benner C.
        • Spann N.
        • et al.
        Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities.
        Mol. Cell. 2010; 38: 576-589
        • Thorvaldsdóttir H.
        • Robinson J.T.
        • Mesirov J.P.
        Integrative genomics viewer (IGV): high-performance genomics data visualization and exploration.
        Brief. Bioinform. 2013; 14: 178-192
        • Young M.D.
        • Wakefield M.J.
        • Smyth G.K.
        • Oshlack A.
        Gene ontology analysis for RNA-seq: accounting for selection bias.
        Genome Biol. 2010; 11: R14
        • Xie C.
        • Mao X.
        • Huang J.
        • et al.
        KOBAS 2.0: a web server for annotation and identification of enriched pathways and diseases.
        Nucleic Acids Res. 2011; 39: W316-W322
        • Kanehisa M.
        • Goto S.
        KEGG: kyoto encyclopedia of genes and genomes.
        Nucleic Acids Res. 2000; 28: 27-30
        • Hawley K.L.
        • Cruz A.R.
        • Benjamin S.J.
        • et al.
        IFNγ enhances CD64-potentiated phagocytosis of treponema pallidum opsonized with human syphilitic serum by human macrophages.
        Front. Immunol. 2017; 8: 1227
        • Weng H.
        • Huang H.
        • Wu H.
        • et al.
        METTL14 inhibits hematopoietic stem/progenitor differentiation and promotes leukemogenesis via mRNA m(6)A modification.
        Cell Stem Cell. 2018; 22 (e9): 191-205
        • Lichinchi G.
        • Zhao B.S.
        • Wu Y.
        • et al.
        Dynamics of human and viral RNA methylation during zika virus infection.
        Cell Host Microbe. 2016; 20 (666-73)
        • Tang L.
        • Wei X.
        • Li T.
        • et al.
        Emerging perspectives of RNA N (6)-methyladenosine (m(6)A) modification on immunity and autoimmune diseases.
        Front. Immunol. 2021; 12630358
        • Li Z.
        • Teng M.
        • Jiang Y.
        • et al.
        YTHDF1 negatively regulates treponema pallidum-induced inflammation in THP-1 macrophages by promoting SOCS3 translation in an m6A-dependent manner.
        Front. Immunol. 2022; 13857727
        • Lichinchi G.
        • Gao S.
        • Saletore Y.
        • et al.
        Dynamics of the human and viral m(6)A RNA methylomes during HIV-1 infection of T cells.
        Nat. Microbiol. 2016; 1: 16011
        • Kim G.W.
        • Imam H.
        • Khan M.
        • et al.
        HBV-induced increased N6 methyladenosine modification of PTEN RNA affects innate immunity and contributes to HCC.
        Hepatology. 2021; 73 (533-47)
        • West A.P.
        • Brodsky I.E.
        • Rahner C.
        • et al.
        TLR signalling augments macrophage bactericidal activity through mitochondrial ROS.
        Nature. 2011; 472: 476-480
        • Martínez I.
        • Oliveros J.C.
        • Cuesta I.
        • et al.
        Apoptosis, toll-like, RIG-I-like and NOD-like receptors are pathways jointly induced by diverse respiratory bacterial and viral pathogens.
        Front. Microbiol. 2017; 8: 276
        • Krueger S.
        • Kuester D.
        • Bernhardt A.
        • Wex T.
        • Roessner A.
        Regulation of cathepsin X overexpression in H. pylori-infected gastric epithelial cells and macrophages.
        J. Pathol. 2009; 217: 581-588
        • Lewis M.S.
        • Danelishvili L.
        • Rose S.J.
        • Bermudez L.E.
        MAV_4644 interaction with the host cathepsin Z protects mycobacterium avium subsp. hominissuis from rapid macrophage killing.
        Microorganisms. 2019; 7: 5
        • Adams L.A.
        • Möller M.
        • Nebel A.
        • et al.
        Polymorphisms in MC3R promoter and CTSZ 3′UTR are associated with tuberculosis susceptibility.
        Eur. J. Hum. Genet. 2011; 19: 676-681
        • Kuo D.
        • Ding J.
        • Cohn I.S.
        • et al.
        HBEGF(+) macrophages in rheumatoid arthritis induce fibroblast invasiveness.
        Sci. Transl. Med. 2019; 11: 491