2. REAPPRAISAL OF BAFF IN AUTOIMMUNE CONDITIONS Pierre Youinou European University of Brittany, Brest, France

3. T NF-like WEAK inducer of apoptosis ( TWEAK ) A Pr oliferation I nducing L igand ( APRIL ) B -cell A ctivation F actor of the TNF F amily ( BAFF ) TUMOR NECROSIS FACTOR FAMILY

5. SETTING THE SCENE FOR BAFF KNOWN ACTIVITIES OF BAFF REGULATION OF BAFF PRODUCTION

6. THE CELL SOURCES OF BAFF Monocytes Macrophages Astrocytes Synoviocytes Dendritic cells Activated T lymphocyte Epithelial cells Neutrophils

7. BR3 TACI BCMA HSPG BAFF SPECIFIC AND NON-SPECIFIC RECEPTORS APRIL BAFF BAFF APRIL APRIL Simone Forest 2010

8. A VARIETY OF MOLECULES Furin BAFF APRIL TWEAK-PRIL TWEAK Daridon, Youinou et al, Autoimmun Rev 2007

9. OVEREXPRESSION OF BAFF IN AUTOIMMUNE DISEASES Number of sera tested Proportion with increased level 94 192 126 52 57 2 Systemic lupus erythematosus patients Autoimmune disease controls Normal controls

10. BAFF IS NOT Stohl et al, Arthritis Rheum 2003 Day of observation Day of observation Persistently elevated serum BAFF Intermittently elevated serum BAFF CONSTANT

12. Menbrane-bound Cell-free BAFF APRIL Glycosylated or not - + PNGaseF 28 21 Anti-BAFF Complexes Virus-like clusters Rochas, Youinou et al, Arthritis Rheum, 2009 Roschke et al, J Immunol, 2004 Le Pottier, Youinou et al, submitted Liu et al, Cell, 2002 BAFF ? Receptor for BAFF Alternative splices 28 17 TFMS - + Rabbit - + Goat BAFF IS NOT RECOGNIZED

13. IN-HOUSE ELISA FOR BAFF Le Pottier, Youinou et al, Clin Chem 2009 28 21 S#1 S#2 Rabbit Ab S#1 S#2 Mouse Ab 28 21 Absorbance (492nm) 0 1 2 3 BAFF (  g/L) 0.1 1 10 100 Goat / Rabbit-biot Goat / Goat-biot Rabbit / Rabbit-biot Rabbit / Goat-biot

14. SETTING THE SCENE FOR BAFF KNOWN ACTIVITIES OF BAFF REGULATION OF BAFF PRODUCTION

15. BAFF-MEDIATED B-CELL HOMEOSTASIS PROLIFERATION BAFF Tg TNF-/- BAFF Tg DIFFERENTIATION CD40-dependent CD40 +IL-4 BAFF CD40-independent BAFF no BAFF yes SSC SURVIVAL ACTIVATION

16. BT1 BT2 High affinity BCR Low affinity BCR Lymphoid follicle germinal center Marginal zone Pre-B MARGINAL ZONE B CELLS AT RISK OF BEING ACTIVATED Apoptosis BAFF

17. MICE TRANSFECTED WITH cDNA FOR HUMAN BAFF Groom, Mackay et al, J Clin Invest 2002 1/1280

18. Walters et al, J Immunol, 2009 BAFF-INDUCED REGULATORY T CELLS WT BAFF-Tg CD4 + Foxp3 +

19. SETTING THE SCENE FOR BAFF KNOWN ACTIVITIES OF BAFF REGULATION OF BAFF PRODUCTION

20. DISCOVERY OF A NEW mRNA FOR BAFF 3’ RACE-PCR ? Exon 5 Exon 3 748 862 Δ 4 BAFF 1 2 4 3 5 6 BAFF Δ BAFF φ BAFF

21. Δ 4 (D->124) LOCATION OF THE NEW PROTEIN GFP Δ 4 RAMOS BAFF pIRES Δ 4 WT 21 kDa 17 kDa Mouse anti-BAFF Rabbit anti-BAFF BAFF Δ 4 BAFF

22. Cold probe (20x) Mouse anti-BAFF Nuclear extract - - - - + - - + - - - + - + - + Rabbit Anti-BAFF - - + + Supershift Shift Δ 4BAFF IS A TRANSCRIPTION FACTOR

23. TRUNCATED BAFF TRANSPORTS TRANSCRIPTION FACTOR(S) FOR FULL-LENGTH BAFF INTO THE NUCLEUS Le Pottier, Tob ón, Mackay, Youinou and Pers , submitted Rabbit Mouse -1600 ATG CD40 NF- κ B subunits Negative control Positive control  actin Mouse Plas Δ 4 wt Mouse/ β -actin 0.85 0.90 1.24 0.86 Rabbit Rabbit/ β -actin 0.83 0.90 1.57 0.70 1 1

24. mRNA for Δ 4 BAFF Δ 4 BAFF protein product Full-length BAFF Transcription factor