TLR4 and C5aR crosstalk in dendritic cells induces a core regulatory network of RSK2, PI3Kβ, SGK1, and FOXO transcription factors

J Leukoc Biol. 2017 Oct;102(4):1035-1054. doi: 10.1189/jlb.2MA0217-058R. Epub 2017 Jul 21.

Abstract

Crosstalk between complement component 5a receptors (C5aRs) and TLRs in dendritic cells (DCs) occurs upon pathogen invasion; however, studies on C5aR and TLR crosstalk mainly focused on the modulating effect of C5a on TLR-induced cytokine production. To elucidate the breadth of C5aR and TLR4 crosstalk, the effect of simultaneous treatment with C5a and LPS was investigated in human monocyte-derived DCs (moDCs) 2 h after stimulation using whole transcriptome sequencing analysis. Although the effect of C5a on hallmark genes defining TLR4-induced DC maturation was limited at this time point, RNA sequencing analysis revealed a great variety of novel C5a targets, of which many interfere with TLR4-mediated immune activation. Analysis of functional relationships among these genes uncovered induction of a central immune regulatory network upon C5aR and TLR4 crosstalk, involving the transcription factors forkhead box (FOX)O1 and FOXO3 and the signaling molecules serum- and glucocorticoid-inducible kinase (SGK1), ribosomal S6 kinase 2 (RSK2), and PI3Kβ. C5aR and TLR crosstalk, furthermore, yielded down-regulation of mainly proinflammatory network branches, including IL-12B, IL-2Rα (IL-2RA), and jagged 1 (JAG1) and cooperative induction of predominantly anti-inflammatory network branches, including sphingosine kinase 1 (SPHK1), β2 adrenergic receptor (ADRB2), gastric inhibitory polypeptide receptor (GIPR), and four-and-a-half Lin11, Isl-1, and Mec-3 domains protein 2 (FHL2). Together, these data point toward induction of generalized immune regulation of DC function. Motif enrichment analysis indicate a prominent role for basic leucine zipper (bZIP) and IFN regulatory factor 4 (IRF4) transcription factors upon C5aR and TLR4 crosstalk. Additionally, differences were observed in the modulating capacity of C5a on DCs in the absence or presence of a pathogen (TLR stimulus). Our findings shed new light on the depth and complexity of C5aR and TLR4 crosstalk and provide new foci of research for future studies.

Keywords: C5a; IRF4; LPS; complement.

MeSH terms

  • Dendritic Cells / immunology*
  • Forkhead Box Protein O1 / immunology*
  • Forkhead Box Protein O3 / immunology*
  • Humans
  • Immediate-Early Proteins / immunology*
  • Phosphatidylinositol 3-Kinases / immunology*
  • Protein Serine-Threonine Kinases / immunology*
  • Receptor, Anaphylatoxin C5a / immunology*
  • Ribosomal Protein S6 Kinases, 90-kDa / immunology*
  • Signal Transduction / immunology*
  • Toll-Like Receptor 4 / immunology*

Substances

  • C5AR1 protein, human
  • FOXO1 protein, human
  • FOXO3 protein, human
  • Forkhead Box Protein O1
  • Forkhead Box Protein O3
  • Immediate-Early Proteins
  • Receptor, Anaphylatoxin C5a
  • TLR4 protein, human
  • Toll-Like Receptor 4
  • Protein Serine-Threonine Kinases
  • Ribosomal Protein S6 Kinases, 90-kDa
  • ribosomal protein S6 kinase, 90kDa, polypeptide 3
  • serum-glucocorticoid regulated kinase