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Bhlhe40 Sustains TH17 Cell Pathogenicity in EAE

This study investigates the role of the transcription factor Bhlhe40 in driving the pathogenicity of autoreactive CD4+ T cells in experimental autoimmune encephalomyelitis (EAE) and multiple sclerosis (MS). The findings suggest that Bhlhe40 is crucial for the pathogenic function of TH17 cells in EAE.

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Bhlhe40 Sustains TH17 Cell Pathogenicity in EAE

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  1. The Transcription Factor Bhlhe40 Sustains TH17 Cell Pathogenicity in EAE Brian T. Edelson Dept. of Pathology and Immunology Div. of Laboratory and Genomic Medicine Washington University School of Medicine St. Louis, MO USA 1 February 2018 Disclosure: Honorarium, Pfizer, Inc (June 2017)

  2. Bhlhe40 DNA binding & dimerization • Basic helix-loop-helix (bHLH) family, member e40 (aka DEC1, Stra13) – first cloned in 1997 in differentiated chondrocytes and as an RA-induced gene in P19 embryonal carcinoma cells • Evolutionarily conserved: present in zebrafish, xenopus. • Expression: non-hematopoietic cells; some macrophages, dendritic cells, granulocytes, activated T cells, iNKT cells • Function: generally transcriptional repression, some examples of transcriptional activation; binds class B E box (CACGTG) • In vivo: circadian rhythm, muscle repair, neuronal excitability, immune cell function • Key question: what drives the pathogenicity of autoreactive CD4+ T cells in EAE and MS

  3. Bhlhe40-/- Mice Are Protected from EAE c d IFN-γ IL-17A GM-CSF IL-10 Lin et al., 2014, Nat. Commun.

  4. Pertussis Toxin (PTX) Induces Bhlhe40 Expression in TH Cells During EAE ±MOG35-55/CFA ±Pertussis toxin (WT or mutant) GFP (Bhlhe40) and cytokine expression by CD4+ T cells in the lymph nodes 7 days Bhlhe40GFP reporter mice Lin et al., 2016, J. Exp Med.

  5. IL-1β Enhances Bhlhe40 Expression in Polarized TH17 Cells A B DLN cell cultures O/N (taken day 7 post EAE induction) Lin et al., 2016, J. Exp Med.

  6. Bhlhe40flox/flox Mice Bhlhe40fl/fl (n=10) Bhlhe40fl/fl (n=6) LysMCreBhlhe40fl/fl (n=4) MRP8CreBhlhe40fl/fl (n=7)

  7. Deletion of Bhlhe40 in T Cells or IL-17Apos Cells Bhlhe40fl/fl (n=12) Bhlhe40fl/fl (n=14) Cd4CreBhlhe40fl/fl (n=14) Il17aCreBhlhe40fl/fl (n=10) Note – Delayed onset and reduction in EAE severity seen here using Il17aCre-mediated deletion of Bhlhe40 is more significant than seen after Il17aCre-mediated deletion of Tbx21 or Rorc (Brucklacher-Waldert et al., 2016, J. Immunol.)

  8. Bhlhe40 Expression in IFN-γposIL-17Aneg “exTH17” and IFN-γposIL-17Apos cells Il17aCre x R26-LoxP-Stop-LoxP-tdTomato x Bhlhe40GFP mice D7 DLN, ICS after P/I

  9. Bhlhe40 Expression in IL-17A Fate-Mapped Cells IL-17A fateposBhlhe40-GFPnegCD4+ T Cells IL-17A fateposBhlhe40-GFPposCD4+ T Cells IFNγpos IFNγposGM-CSFpos IL-17Apos Triple neg GM-CSFpos IL-17AposGM-CSFpos IL-17AposIFNγpos D7 DLN, ICS after P/I (n=4) IL-17AposIFNγposGM-CSFpos

  10. A B 3355 peaks Bhlhe40 ChIP-Seq on WT and KO TH17 cells 10 kb 10 kb C WT Bhlhe40 KO 10 kb WT Il10 KO Csf2/Il3 WT KO

  11. BHLHE40 Expression in Human TH Subsets (RNA-Seq) – Sorted from PBMCs (healthy donors) CCR6+CXCR3+ = TH1/17 Arlehamn C. L. et al., 2014, J. Immunol.

  12. Role of Bhlhe40 in Pathogenic CD4+ T Cells Lin et al., 2014, Nat. Commun. Lin et al., 2016, J. Exp. Med. Ronchi et al., 2016, Nat. Commun. Mufazalov et al., 2016, EMBO

  13. Acknowledgments Chih-Chung “Jerry” Lin Tara R. Bradstreet Elizabeth A. Schwarzkopf Nicholas Jarjour Melissa Cook Washington University School of Medicine Maxim Artyomov Takeshi Egawa John Russell Gregory Wu National University of Singapore Reshma Taneja

  14. BACKUP SLIDES

  15. Transcriptional Analysis of Polarized Bhlhe40-/- TH Cells

  16. Bhlhe40-Expressing CD4+ T CellsAreAbundantintheCNS During EAE Spinal cord (d15) CD4 GFP GFP (Bhlhe40) expression by CD4+ T cells in the spleen and CNS MOG35-55/CFA Pertussis toxin (co-adjuvant) Active EAE induction 14-15 days Bhlhe40GFP reporter mice Lin et al., 2016, J. Exp. Med.

  17. IL-1β Enhances Bhlhe40 Expression in Polarized TH17 Cells A B C EAE Pathogenic gene signature  

  18. Pertussis Toxin (PTX) Induces IL-1β Production by Myeloid Cells A B (Monocytes/monocyte-derived cells) DLN, Day 7 post EAE induction

  19. Identification of human TH subsets (gating strategy) 1: TB-specific (tetramer-positive) T cells 2: naïve CD4 T cells 3: “TH1/17” cells (6+X3+) 4: TH17 cells 5: TH1 cells 6: TH2 cells Modified from Arlehamn C. L. et al. J Immunol 2014

  20. Zhang H et al., J Allergy Clin Immunol 2013 (GSE43005) Arlehamn C. L., et al. J Immunol 2014 (GSE56179)

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