The document summarizes recent research findings on apoptosis regulation by Bcl-2 family members and XIAP. Key points include:
1) BID and BIM can mediate crosstalk from death receptors to mitochondria to induce apoptosis.
2) XIAP is a crucial factor in determining whether Fas induces type I or type II apoptosis.
3) FasL can trigger both apoptosis and necroptosis in neutrophils.
4) Non-apoptotic roles of IAPs and death receptors are becoming increasingly evident and important.
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Regulation of Apoptosis by BCL-2 Family Members (Prof. Thomas Kaufmann)
1. Valld’Hebron Research Institute
Nov 15 2012
Regulation of Apoptosis by Bcl-2 Family Members
and XIAP
Thomas Kaufmann
Institute of Pharmacology,
University of Bern , Switzerland
thomas.kaufmann@pki.unibe.ch
2. Necrosis vs. Apoptosis
passive, „accident“, active, energy-dependent,
alwayspathological physiological + pathophys.
cellsstayintact,
lysis of cells
clearedbyphagocytosis
whole (parts of) tissue/ oftendeath of individual
organaffected cells
no inflammation,
inducesinflammation
inducestolerance
2
3. PROGRAMMED CELL DEATH
Apoptosis
Necroptosis
(RIPK1/3)
Autophagic Cell Death NECROTIC
Pyroptosis CELL DEATH
Anoikis “accidents”:
(casp-1) - lack of energy
- physical damage
Cornification Pyronecrosis - chemical damage
(“keratinization”)
3
6. The Bcl-2 Protein Family Regulates the Integrity of the
Mitochondrial Outer Membrane
BH3-only
Bcl-2-like Bcl-2 Family
Bax-like
MOMP: mitochondrial outer
membrane permeabilisation MOMP
Smac/Diabl
cIAP1,2 Cyt.c + adaptor (APAF-1)
o
XIAP
Vaux &Silke Nat Rev MCB 2005
Caspase-3, -7 (, -6) Caspase-9
6
7. Only XIAP can directly block caspases-3 and -9
Riedl and Shi, Nat Rev MCB 2004
7
8. Selective Interactions Between BH3-Only and Bcl-2-Like Proteins
Bim, Bid, Puma ABT-737 (Oltersdorf et al
Nature 2005)
Obatoclax (Nguyen et al
PNAS 2007)
A1
Chen et al Mol Cell 2005
(modified)
8
9. The Bcl-2 Family – Still Many Open Questions
BH3-only
Bcl-2-like
Bax/Bak
Strasser, Nat Rev Imm 2005
9
10. BOK: A BAX/BAK-Like Protein?
BOK: BCL-2 related ovarian killer (Hsu et al. PNAS 1997)
BOK is widely expressed
Ke F. et al., CDD 2012
10
11. BOK is Deleted in Human Cancers with high Frequency
Beroukhim et al., Nature, 2010
11
12. BOK IS NOT A FUNCTIONAL BAX/BAK HOMOLOGUE
• BOK induces intrinsic apoptosis upstream of BAX/BAK
• BOK localises predominantly to non-mitochondrial sites:
– Golgi, ER/nuclear outer membrane
– nuclear compartment
• Bok-/- cells present with aberrant ER stress response (part. BFA)
+4-OHT (h):
Echeverry et al., in revision
12
14. APOPTOSIS IS ONLY ONE OF SEVERAL POSSIBLE OUTCOMES IN DR SIGNALING
FADD/C8 (?)
Proliferation
cIAP1/2
Apoptosis
Necroptosis
NFkB
TNFa
cell survival
14
15. TNF-R1: Not Meant to Kill
TNFa
TNF-R1
RIP1
Apoptosis cIAP1/2
Necroptosis
NFkB
anti-apoptotic genes
cytokines
=> cell survival
15
16. LPS plus GalactosamineInjectionModelof TNF-R1-MediatedFulminant Hepatitis
• Bacterial LPS -> TNFa(Macrophages, Neutrophils, NK T)
• Response via soluble, circulating TNFa TNF-R1
(Pfeffer et al 1993, Rothe et al 1993, Grivennikov et al 2005)
• Sensitisation by D-(+)-galactosamine (GalN)
Maeda S et al. Immunity 2003
Kaufmann et al. (2009)
16
17. Hepatocytesare Type IICells
TYPE I TYPE II
Bid
Caspase-8
tBid
Bid Caspase-8 Bcl-2-like
? X
Bax/Bak
Effector
Caspases
Effector Cyt.c
Caspases Apaf1/ Caspase-9
17
18. The BH3-Only Protein BIM IsRapidlyPhosphorylated in LPS/GalN-Induced Hepatitis
*
Kaufmann et al. Immunity (2009) 18
19. Both BID and BIM areInvolved in LPS/GalN-InducedHepatitis
Kaufmann et al. Immunity (2009)
19
20. BIM isActivatedby JNK MediatedPhosphorylation
A B
+/- D-JNKI1 (30 mg/kg, i.p.)
*
Kaufmann et al. Immunity (2009) 20
21. Both BID and BIM canMediate a Crosstalk fromDeathReceptors to Mitochondria
JNK
Corazza et al. JCI (2006)
Bim JNK mediated BIM-activation
downstream of TRAIL
21
22. Fas/CD95/Apo-1
• FasL mainly expressed on activated T cells and natural killer cells.
• Critical role in the control of the immune system
• Fas or FasL-mutant mice develop lymphadenopathy and SLE (systemic
lupus erythematosus)-like disease and are predisposed to lymphoma
development
• Many ALPS patients have heterozygous inherited mutations in the
Fasgene. (Fisher et al. Cell 1995, Rieux-Laucat et al. Science 1995)
• Only the membrane bound form of FasL is inducing cell death (O’Reilly et
al. Nature 2009)
22
23. Type I or Type II Fas-Induced Apoptotic Pathway
FasL FasL
TYPE I TYPE II
Fas Fas
? Bid
Caspase-8
tBid
Bid Caspase-8 Bcl-2-like
? X
Bax/Bak
Effector
Caspases
Effector Cyt.c
Caspases Apaf1/ Caspase-9
23
36. Neutrophils: - most frequent leukocyte in human blood
- major role in innate immunity
- end-differentiated, short-lived
- apoptotic clearance of activated neutrophils essential
36
37. Primary Neutrophils Die By Classical Apoptosis When Cultured In Vitro
C57BL/6 WT
100
80
Survival (%)
60 untreated
GM-CSF 1ng/ml
G-CSF 10ng/ml
40
Q-VD oph 20uM
20
0
0 24 48 72 96 Time (h)
37
38. Neutrophils die by Apoptosis in Response to High Doses of TNFa
100
80 wt untreated
bid-/- untreated
Survival %
60
wt TNFa
40 bid-/- TNFa
20
0
0 24 48 72 96 Time (h)
100
80
60 wt TNFa
TNFα 50 ng/ml bid-/- TNFa
Q-VD oph 20 μM 40
wt TNFa + Q-VD-oph
20 bid-/- TNFa + Q-VD-oph
0
0 24 48 72 96 38
47. SUMMARY
• Several poorly characterised BCL-2 family members
• Besides BID, BIM can mediate a crosstalk from DR to
mitochondria
• XIAP is a crucial discriminator between type I and
type II Fas-induced apoptosis
• FasL triggers mix of apoptosis and necroptosis in
neutrophils
• Non-apoptotic roles of IAPs and DRs become
increasingly evident (important)
47
48. ACKNOWLEDGMENTS
University of Bern (CH) WEHI, Melbourne (AU) • Philipp Jost(Munich DE)
• NohemyEcheverry •Andreas Strasser • MadsGyrd-Hansen (Kopenhagen, DK)
• UrsinaGurzeler •Francine Ke • ChristophBorner (Freiburg i.Brsg., DE)
• Tatiana Rabachini •Paul Ekert
• Daniel Bachmann • Georg Häcker(Freiburg i. Brsg., DE)
•John Silke
• Simone Wicki • Thomas Brunner (Konstanz, DE)
•David Huang
• Nicole Tochtermann • Frank Essmann(Tübingen, DE)
•UeliNachbur
• LaetitiaRoh
• Julia Fernandez-Rodriguez
(Gothenburg, SE)
• Hans-Uwe Simon
• Mario Tschan • Kurt Ballmer (Villigen, CH)
• ShidaYousefi
• Clemens Dahinden
48