Treg-Cell-Derived IL-35-Coated Extracellular Vesicles Promote Infectious Tolerance

Cell Rep. 2020 Jan 28;30(4):1039-1051.e5. doi: 10.1016/j.celrep.2019.12.081.


Interleukin-35 (IL-35) is an immunosuppressive cytokine composed of Epstein-Barr-virus-induced protein 3 (Ebi3) and IL-12α chain (p35) subunits, yet the forms that IL-35 assume and its role in peripheral tolerance remain elusive. We induce CBA-specific, IL-35-producing T regulatory (Treg) cells in TregEbi3WT C57BL/6 reporter mice and identify IL-35 producers by expression of Ebi3TdTom gene reporter plus Ebi3 and p35 proteins. Curiously, both subunits of IL-35 are displayed on the surface of tolerogen-specific Foxp3+ and Foxp3neg (iTr35) T cells. Furthermore, IL-35 producers, although rare, secrete Ebi3 and p35 on extracellular vesicles (EVs) targeting a 25- to 100-fold higher number of T and B lymphocytes, causing them to acquire surface IL-35. This surface IL-35 is absent when EV production is inhibited or if Ebi3 is genetically deleted in Treg cells. The unique ability of EVs to coat bystander lymphocytes with IL-35, promoting exhaustion in, and secondary suppression by, non-Treg cells identifies a novel mechanism of infectious tolerance.

Keywords: CD81; Ebi3; IL-35; Treg; cytokines; extracellular vesicles; p35; tetraspanin; tolerance; exosome.

MeSH terms

  • Animals
  • B-Lymphocytes / immunology
  • B-Lymphocytes / metabolism
  • CD4-Positive T-Lymphocytes / immunology
  • CD4-Positive T-Lymphocytes / metabolism
  • CD8-Positive T-Lymphocytes / immunology
  • CD8-Positive T-Lymphocytes / metabolism
  • Coculture Techniques
  • Extracellular Vesicles / immunology
  • Extracellular Vesicles / metabolism*
  • Extracellular Vesicles / ultrastructure
  • Female
  • Forkhead Transcription Factors / metabolism
  • Gene Knockout Techniques
  • Heart Transplantation
  • Immune Tolerance*
  • Immunosuppression Therapy
  • Interleukin-12 Subunit p35 / metabolism*
  • Interleukins / immunology
  • Interleukins / metabolism*
  • Mice
  • Mice, Inbred C57BL
  • Mice, Inbred CBA
  • Mice, Transgenic
  • Microscopy, Electron, Transmission
  • Minor Histocompatibility Antigens / metabolism*
  • Receptors, Cytokine / metabolism*
  • T-Lymphocytes, Regulatory / immunology*
  • T-Lymphocytes, Regulatory / metabolism


  • Ebi3 protein, mouse
  • Forkhead Transcription Factors
  • Foxp3 protein, mouse
  • Interleukin-12 Subunit p35
  • Interleukins
  • Minor Histocompatibility Antigens
  • Receptors, Cytokine
  • interleukin-35, mouse