Apoptosis and Autophagy
Apoptosis Overview Pathway
NIK
FLIP
Casp-8,-10
p53
RIP
TRAF2
JNK
HtrA2
NF-κB
[Ca2+]
Bcl-xL
ICAD
opla
Cyt
PI3K III
JNK
Phagophore
Chk1/2
Atg16 Atg12
FLIP
XIAP
NF-κB
Atg3
Atg10
p53
Puma
Noxa
DNA
Fragmentation DNA Damage
Bim
Inhibition of Apoptosis Pathway
Death Receptor Signaling Pathway
© 2002–2014 Cell Signaling Technology, Inc.
© 2003–2014 Cell Signaling Technology, Inc.
TAB
IKKγ cdc37
JNK
Casp-8/10
IKKγ
HSP70
Bid
tBid
XIAP
IκB
Casp-3,-6,-7
NF-κB Apoptosis
Casp-9
PDK1
Bax
Bax
p70S6K
Erk1/2
PKC
Jak
Src
JNK
Bim
Bcl-2
Bcl-xL
Bad
p90RSK
HSP27
Cyto c
Mcl-1
Mule
Bax
HSP70
PKA
Bim
FoxO1
CREB
Cyto c
Complex
IIa
RIP1
RIP3
Bcl-2
Casp-9
Stat1
IκB
NF-κB
DR3
APO-3
FADD
FADD
Erk1/2
cIAP1/2
PKA
IKKα
Smac
NF-κB
p100
IκB
Casp-3
Akt
Bad
14-3-3
FoxO1
Bcl-2
Bcl-xL
Bad
Cytosolic
Sequestration
GAS2
α-Fodrin
ROCK1
DFF45
Acinus
Calcineurin
Cell Shrinkage
Membrane Blebbing
Stat3
Pathway Diagram Key
Direct Stimulatory Modification
Multistep Stimulatory Modification
Direct Inhibitory Modification
Multistep Inhibitory Modification
Apaf-1
Arts
Pro-Apoptotic
Pro-Survival
DNA
Chromatin
Fragmentation Condensation
© 2002 – 2013 Cell Signaling Technology, Inc.
Caspase-3
CaMKII
Bax
Bax
Caspase-2
Bcl-2
RAIDD
Noxa
Puma
[NAD]
Bcl-2
Smac/
Diablo
Endo G
AIF
PIDD
SirT2
p53
Mule
HtrA2
ub
Bax
Bcl-2
Cyto c
Caspase-9
JNK
Hrk
DP5
Bcl-2
Bmt
Bax
Bax
XIAP
ATM/
ATR
ING2
Transcription
Death Receptor Signaling • created January 2002 • revised February 2012
Tentative Stimulatory Modification
Transcriptional Stimulatory Modification
Separation of Subunits or Cleavage Products
Tentative Inhibitory Modification
Transcriptional Inhibitory Modification
Joining of Subunits
Translocation
Kinase
Transcription Factor
Receptor
Pro-apoptotic
GAP
GTPase
Phosphatase
Caspase
Enzyme
Anti-apoptotic
GEF
G-protein, ribosomal subunit
Autophagy Signaling Pathway:
Inhibition of Apoptosis Pathway:
Apoptosis is a regulated cellular suicide mechanism characterized by nuclear condensation, cell shrinkage, membrane
blebbing, and DNA fragmentation. Caspases, a family of cysteine proteases, are the central regulators of apoptosis.
Initiator caspases (including caspase-2, -8, -9, -10, -11, and -12) are closely coupled to pro-apoptotic signals. Once
activated, these caspases cleave and activate downstream effector caspases (including caspase-3, -6, and -7), which
in turn execute apoptosis by cleaving cellular proteins following specific Asp residues. Activation of Fas and TNFR by
FasL and TNF, respectively, leads to the activation of caspase-8 and -10. DNA damage induces the expression of PIDD
which binds to RAIDD and caspase-2 and leads to the activation of caspase-2. Cytochrome c released from damaged
mitochondria is coupled to the activation of caspase-9. XIAP inhibits caspase-3, -7, and -9. Mitochondria release multiple
pro-apoptotic molecules, such as Smac/Diablo, AIF, HtrA2 and EndoG, in addition to cytochrome c. Smac/Diablo binds
to XIAP which prevents it from inhibiting caspases. Caspase-11 is induced and activated by pathological proinflammatory and pro-apoptotic stimuli and leads to the activation of caspase-1 to promote inflammatory response and apoptosis
by directly processing caspase-3. Caspase-12 and caspase-7 are activated under ER stress conditions. Anti-apoptotic
ligands including growth factors and cytokines activate Akt and p90RSK. Akt inhibits Bad by direct phosphorylation and
prevents the expression of Bim by phosphorylating and inhibiting the Forkhead family of transcription factors (Fox0). Fox0
promotes apoptosis by upregulating pro-apoptotic molecules such as FasL and Bim.
Macroautophagy, often referred to as autophagy, is a catabolic process that results in the autophagosomic-lysosomal
degradation of bulk cytoplasmic contents, abnormal protein aggregates, and excess or damaged organelles. Autophagy
is generally activated by conditions of nutrient deprivation but has also been associated with physiological as well as
pathological processes such as development, differentiation, neurodegenerative diseases, stress, infection and cancer.
The kinase mTOR is a critical regulator of autophagy induction, with activated mTOR (Akt and MAPK signaling) suppressing autophagy, and negative regulation of mTOR (AMPK and p53 signaling) promoting it. Three related serine/
threonine kinases, UNC-51-like kinase -1, -2, and -3 (ULK1, ULK2, UKL3), which play a similar role as the yeast Atg1,
act downstream of the mTOR complex. ULK1 and ULK2 form a large complex with the mammalian homolog of an
autophagy-related (Atg) gene product (mAtg13) and the scaffold protein FIP200 (an ortholog of yeast Atg17). Class III
PI3K complex, containing hVps34, Beclin 1 (a mammalian homolog of yeast Atg6), p150 (a mammalian homolog of
yeast Vps15), and Atg14-like protein (Atg14L or Barkor) or ultraviolet irradiation resistance-associated gene (UVRAG),
is required for the induction of autophagy. The Atg genes control the autophagosome formation through Atg12-Atg5
and LC3-II (Atg8-II) complexes. Atg12 is conjugated to Atg5 in a ubiquitin-like reaction that requires Atg7 and Atg10 (E1
and E2-like enzymes, respectively). The Atg12–Atg5 conjugate then interacts noncovalently with Atg16 to form a large
complex. LC3/Atg8 is cleaved at its C terminus by Atg4 protease to generate the cytosolic LC3-I. LC3-I is conjugated
to phosphatidylethanolamine (PE) also in a ubiquitin-like reaction that requires Atg7 and Atg3 (E1 and E2-like enzymes,
respectively). The lipidated form of LC3, known as LC3-II, is attached to the autophagosome membrane. Autophagy and
apoptosis are connected both positively and negatively, and extensive crosstalk exists between the two. During nutrient
deficiency, autophagy functions as a pro-survival mechanism; however, excessive autophagy may lead to cell death,
a process morphologically distinct from apoptosis. Several pro-apoptotic signals, such as TNF, TRAIL, and FADD, also
induce autophagy. Additionally, Bcl-2 inhibits Beclin-1-dependent autophagy, thereby functioning both as a pro-survival
and as an anti-autophagic regulator.
Cell survival requires the active inhibition of apoptosis, which is accomplished by inhibiting the expression of pro-apoptotic
factors as well as promoting the expression of anti-apoptotic factors. The PI3K pathway, activated by many survival
factors, leads to the activation of Akt, an important player in survival signaling. PTEN negatively regulates the PI3K/
Akt pathway. Activated Akt inhibits the pro-apoptotic Bcl-2 family member Bad, Bax, caspase-9, GSK-3 and FoxO1 by
phosphorylation. Many growth factors and cytokines induce anti-apoptotic Bcl-2 family members. The Jaks and Src phosphorylate and activate Stat3, which in turn induces the expression of Bcl-xL and Bcl-2. Erk1/2 and PKC activate p90RSK,
which activates CREB and induces the expression of Bcl-xL and Bcl-2. These Bcl-2 family members protect the integrity
of mitochondria, preventing cytochrome c release and the subsequent activation of caspase-9. TNF-α may activate both
pro-apoptotic and anti-apoptotic pathways; TNF-α can induce apoptosis by activating caspase-8 and -10, but can also
inhibit apoptosis signaling via NF-κB, which induces the expression of anti-apoptotic genes such as Bcl-2. cIAP1/2 inhibit
TNF-α signaling by binding to TRAF2. FLIP inhibits the activation of caspase-8.
Death Receptor Signaling Pathway:
Apoptosis can be induced through the activation of death receptors including Fas, TNFαR, DR3, DR4, and DR5 by their
respective ligands. Death receptor ligands characteristically initiate signaling via receptor oligomerization, which in turn
results in the recruitment of specialized adaptor proteins and activation of caspase cascades. Binding of FasL induces
Fas trimerization, which recruits initiator caspase-8 via the adaptor protein FADD. Caspase-8 then oligomerizes and is
activated via autocatalysis. Activated caspase-8 stimulates apoptosis via two parallel cascades: it can directly cleave and
activate caspase-3, or alternatively, it can cleave Bid, a pro-apoptotic Bcl-2 family protein. Truncated Bid (tBid) translocates to mitochondria, inducing cytochrome c release, which sequentially activates caspase-9 and -3. TNF-α and DR-3L
For Research Use Only. Not For Use in Diagnostic Procedures. © 2014 Cell Signaling Technology, Inc., Cell Signaling Technology® is a registered trademark of Cell Signaling Technology, Inc.
DNA Damage
Genotoxic Stress
Apoptosis
Survival
Apoptosis Overview Pathway:
www.cellsignal.com/pathways | www.cstj.co.jp
Bim
Hrk
DP5
Bcl-xL
HSP60
p52 RelB
PARP
DFF40
Stat1
JNK
Bmt
Bak
Bak
p70 S6K
14-3-3
p65/ NF-κB
RelA p50
FAS
Bim
Microtubules
Bad
Casp-7
Apoptosis
14PSTCORPAPOP0133ENG_00
Bid
Bcl-2 Family
FLIP
XIAP
A20
Mcl-1
Bcl-xL
p90RSK
RelB
XIAP
Necroptosis
CREB
LC8
Mule
tBid
Processing
Actin
PI3K
PKC
NIK
HSP27
Lamin A
LC8
Caspase-8,-10
Casp-8,-10
RIP1
Bim
tBid
Apaf1
Casp-6
Fas/
CD95
DR4/5
RIP
TRADD TRADD
FADD
TRAF3
Casp-8
MLKL
XIAP
Stat3
[cAMP]
s
cleu
NF-κB
Complex
IIb
Bcl-2
m
Nu
tBid
MKK1
Akt
FLIP
Bid
las
ytop
C
Daxx
ASK1
Casp-8, -10
RIP
TRAF2 TRAF1
cIAP1/2
IKKα/β/γ
APO-2L/
TRAIL
Death Stimuli:
Survival Factor Withdrawal
FasL
Survival Factors:
Growth Factors, Cytokines, etc.
Mcl-1
HSP90 HSP70
Raf
TRADD
RIP1
TRAF2
CYLD
cIAP1/2
FADD
HSP90
TNF-R2
Bim
Apaf-1
Akt
PI3K
FoxO1
Smac/
Diablo
NF-κB
sm
Ras
PIP3
GSK-3
HSP90
IκB
Fas/
CD95
Cytopla
HSP90
IKKα IKKβ
IKKβ HSP90
APO-3L/
TWEAK
TRAF1
FLIP
TAK1
TNF-R1
PTEN
A20
HSP27
NIK
TNF-α
TRAF2
RIP
FADD
TRADD
TRAF2
cIAP
CYLD
Mitochondrial Control of Apoptosis Pathway
© 2002–2014 Cell Signaling Technology, Inc.
FasL
TNFR1
Autophagolysosome
Lysosome
LC3
Death Receptor Signaling
TNF-α
Fusion
LC3-I
FADD
Survival Factors:
Growth Factors,
Cytokines, etc.
PE
Atg4
Atg12
c-Jun
p53
Lysosome
Atg7
Atg7
JNK
FoxO1
www.cellsignal.com
TNF-α
Autophagosome
p53
Atg16
Endo G
LC3-II
Atg5
Atg5
AIF
CAD
Atg1
Membrane
Nucleation
Apoptosis
• Cell shrinking
• Membrane
blebbing
leus
Nuc
Autophagy
Induction
Beclin1
Bcl-2
APP
sm
ULK complex
p150
Sequestration
ATM/ATR
CAD
ER Stress
PRAS40
Casp-9
Apaf-1
ROCK
GβL mTOR Raptor
hVPS34
Cellular Stress
FoxO1
p53 / Genotoxic
Stress
Amino
Acids
Akt
Endo G
AIF
PI3K
cdc2
14-3-3
Puma
Mcl-1
Mule
Casp-3,-6,-7
Casp-7
Bad
Cell Cycle
Noxa
Cyto c
IκBα
Erk1/2
Bim
Smac/
Diablo
XIAP
Casp-9
Calpain
NF-κB
Bak
tBid
Arts
IκBα
Bcl-2
Bax
Bid
Casp-12
PKC
p90RSK
SIR2
MAPK / Erk1/2
Signaling
AMPK
Signaling
ASK1
RIP RAIDD
PIDD
Casp-2
TRADD
FADD
IKKβ
PI3K-I / Akt
Signaling
© 2007–2014 Cell Signaling Technology, Inc.
FADD
RIP
Itch
CYLD
TAK1
IKKγ
c-IAP
TRAF2
TRADD
RIP1
T RADD
FADD
oplasm
IKKα
Trophic Factors
TRAF2
c-IAP
Cyt
Autophagy Signaling Pathway
TNF, FasL, TRAIL
© 2002–2014 Cell Signaling Technology, Inc.
can deliver pro- or anti-apoptotic signals. TNFαR and DR3 promote apoptosis via the adaptor proteins TRADD/FADD and
the activation of caspase-8. Interaction of TNF-α with TNFαR may activate the NF-κB pathway via NIK/IKK. The activation
of NF-κB induces the expression of pro-survival genes including Bcl-2 and FLIP, the latter can directly inhibit the activation
of caspase-8. FasL and TNF-α may also activate JNK via ASK1/MKK7. Activation of JNK may lead to the inhibition of
Bcl-2 by phosphorylation. In the absence of caspase activation, stimulation of death receptors can lead to the activation
of an alternative programmed cell death pathway termed necroptosis by forming complex IIb.
Mitochondrial Control of Apoptosis Pathway:
The Bcl-2 family of proteins regulate apoptosis by controlling mitochondrial permeability. The anti-apoptotic proteins Bcl-2
and Bcl-xL reside in the outer mitochondrial wall and inhibit cytochrome c release. The proapoptotic Bcl-2 proteins Bad,
Bid, Bax, and Bim may reside in the cytosol but translocate to mitochondria following death signaling, where they promote
the release of cytochrome c. Bad translocates to mitochondria and forms a pro-apoptotic complex with Bcl-xL. This
translocation is inhibited by survival factors that induce the phosphorylation of Bad, leading to its cytosolic sequestration.
Cytosolic Bid is cleaved by caspase-8 following signaling through Fas; its active fragment (tBid) translocates to mitochondria. Bax and Bim translocate to mitochondria in response to death stimuli, including survival factor withdrawal. Activated
following DNA damage, p53 induces the transcription of Bax, Noxa, and PUMA. Upon release from mitochondria, cytochrome c binds to Apaf-1 and forms an activation complex with caspase-9. Although the mechanism(s) regulating mitochondrial permeability and the release of cytochrome c during apoptosis are not fully understood, Bcl-xL, Bcl-2, and Bax
may influence the voltage-dependent anion channel (VDAC), which may play a role in regulating cytochrome c release.
Mule/ARF-BP1 is a DNA damage activated E3 ubiquitin ligase for p53, and Mcl-1, an anti-apoptotic member of Bcl-2.
Revised March 2014