Outline of presentation

• Introduction of apoptosis and the

induction pathway;
• Common structure and features of
caspases;
Caspases and Apoptosis • Classification of caspases;
• Activation pathway of initiator caspases
―the signaling systems of proteases and effector caspases;
• Substrates of caspases;
Yun Ding MCDB
• Regulation of caspases;
4/21/2005 • Relations between caspases-mediated
apoptosis and human diseases.

“Death” is important to the “life” Apoptosis VS Necrosis

―Distinct morphological and biochemical features
• Apoptosis (programmed cell death): a Apoptosis:
physiological process of cellular a. Cytoplasmic shrinkage
autodestruction, or cell suicide. b. Disrupt of cellular organelle

c. Membrane blebbing

• Strictly controlled in response to integrity d. Nuclear fragmentation

of prodeath signaling. e. Intranucleosomal DNA

fragmentation

f. Phosphatidylserine exposure
• Critical roles of apoptosis in development, g. Apoptotic bodies formation
maintenance of homeostasis and host g. Engulfed by macrophages or
defense in multicellular organisms. other cells
h. No inflammatory responses

Apoptosis-”falling off petals or leaves from a flower and trees in autumn” Marco Vitale et al.

Induction of apoptosis:
DISC

• Cell surface death receptor mediated

pathway (extrinsic)

apoptosome

• Mitochondrial-initiated pathway
(intrinsic)

C.J.Zeiss Vet Pathol 40: 481-495 (2003)

1
 General features of Caspases Caspase structures
• Human ICE (interleukin-1βprocessing enzyme,
1992) and C. elegans ced-3 gene (1993) were the
first identified caspases;
• So far, 11 members from human, 4 from C. elegans
and 7 from Drosophila have been identified with
obvious conservation in evolution.
• Caspase (Cystein aspartate-specific protease)
a. active cystein residue in the catalytic site;
b. specificity in cleavage after an Asp residue of
the substrate;
c. synthesized as inactive zymogens
Caspase-3

William C. Earnshaw et al. Annu. Rev. Biochem. 68: 383-424 (1999)

Alexei Degterev et al. Oncogene 22: 8543-8567 (2003)

Classification of Caspases Activation of Caspase-8 through cell surface death receptor

mediated pathway

―DISC (death inducing signaling complex) formation

Alexei Degterev et al. Oncogene 22: 8543-8567 (2003) Avi Ashkenazi et al. Science 281: 1305-1308 (1998)

Activation of Caspase-9 through mitochondria initiated

Molecular mechanism of Caspase-8 activation
pathway

―Apoptosome formation
―Induced proximity for self-activation

Howard Y. Chang et al. Microbiology and Molecular Biology Reviews 821-846 (2000) Imawati Budihardjo et al. Annu. Rev. Cell Dev. Biol. 15: 269-290 (1999)

Helen Blanchard et al. Structure 7: 1125-1133 (1999) Alexei Degterev et al. Oncogene 22: 8543-8567 (2003)

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 Molecular mechanism of Caspase-9 activation Activation of effector Caspases-3,6,7 through
transactivation by initiator caspases cleavage

―allosteric regulation
Procaspase-7 activation:

1. Proteolytic cleavage is
not required for Caspase-9
activiation.

2. Caspase-9 dimer has

only one active site.

Alexei Degterev et al. Oncogene 22: 8543-8567 (2003) Stefan J. Riedl et al. Nature Molecular Cell biology Reviews. 5: 897-907 (2004)

Substrates of Caspases


• Apoptotic and inflammatory regulators;
• Protein kinases and other signal
transduction regulators;
• Cytosolic and nuclear structural proteins;
• Repair and housekeeping enzymes;
• Cell cycle regulators;
• Disease related factors.


Alexei Degterev et al. Oncogene 22: 8543-8567 (2003)

Regulation of Caspases Bcl-2 family: involved in mitochondria-mediated apoptosis

• Gene expression;
• Phosphorylation, nitrosylation
a. Inactivation of human caspase-9 by Akt;
b. Direct inhibition of caspase activity
through active site nitrosylation
• Compartmentalization (caspase-1,8,12);
• Bcl-2 protein family: pro- vs. anti-; Alternative model for Bcl-2
• Caspase inhibitors: family to regulate cytochrome c
release from mitochondria
a. CrmA and p35;
b. FLIPs;
c. IAPs

Jerry M. Adams et al. Science 281: 1322-1326 (1998)

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 FLIPs inhibition
CrmA and p35: suicide substrates
• v-FLIPs: contain two DED domains and
p35-mediated inhibition of caspase-8
compete with recruitment of procaspases to
the death receptor complex;

• c-FLIP:
a. c-FLIPS: similar to v-FLIP;
b. c-FLIPL: contains NH2-terminal DEDs
and COOH-terminal caspase-like domain

Stefan J. Riedl et al. Nature Molecular Cell biology Reviews. 5: 897-907 (2004)

IAPs: inhibitors of apoptosis

IAP-mediated inhibition
of effector caspase

IAP-mediated inhibition
of initiator caspase

These inhibition does not need cleavage by caspases.

Stefan J. Riedl et al. Nature Molecular Cell biology Reviews. 5: 897-907 (2004) Stefan J. Riedl et al. Nature Molecular Cell biology Reviews. 5: 897-907 (2004)

Comparison of the main caspases-mediated pathway in Roles of caspases and apoptosis in

C. elegans, D. melanogaster and mammals human diseases
• Neurodegenerative diseases;
1. Huntington’s disease: polyQ
proteins and caspase-3;
2. Alzheimer’s disease: β-amyloid
production and caspase-3,12

• Viral infection;
• Autoimmunity;
• Cancer.

Bruce A.Hay et al. Nature Genetics Reviews. 5: 911-922 (2004)

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 References
• C. J. Zeiss The Apoptosis-necrosis continuum: insights from
genetically altered mice. Vet Pathol 40:481-495 (2003)
• William C. Earnshaw et al. Mammalian caspases:structure,
activation, substrates,and functions during apoptosis. Annu. Rev.
Biochem 68:383–424 (1999)
• Alexei Degterev et al. A decade of caspases. Oncogene 22: 8543–
8567 (2003)
• Avi Ashkenazi et al. Death receptors: signaling and modulation.

“To be or not to be, •

Science 281: 1305-1308 (1998)
Howard Y. Chang et al. Proteases for cell suicide: functions and
regulation of caspases. Microbiology and Molecular Biology Reviews
that is a question. •
821-846 (2000)
Helen Blanchard et al. The three-dimensional structure of
caspase-8: an initiator enzyme in apoptosisStructure 7: 1125-1133
(1999)
• Imawati Budihardjo et al. Biochemical pathway of caspase
activation during apoptosis. Annu. Rev. Cell Dev. Biol. 15: 269-290
(1999)

A. Philchenkov J. Cell. Mol. Med. 8(4): 432-444 (2004)

• Stefan J. Riedl et al. Molecular mechanisms of caspase regulation

during apoptosis. Nature Molecular Cell biology Reviews. 5: 897-
907 (2004)
• Jerry M. Adams et al. The Bcl-2 protein family: arbiters of cell
survival. Science 281: 1322-1326 (1998)
• Bruce A.Hay et al. The genetics of cell death: approaches, insights
and opportunities in Drosophila. Nature Genetics Reviews. 5: 911-
922 (2004)
• A. Philchenkov Caspases: potential targets for regulating cell death
J. Cell. Mol. Med. 8(4): 432-444 (2004)

5
 References
• C. J. Zeiss The Apoptosis-necrosis continuum: insights from
genetically altered mice. Vet Pathol 40:481-495 (2003)
• William C. Earnshaw et al. Mammalian caspases:structure,
activation, substrates,and functions during apoptosis. Annu. Rev.
Biochem 68:383–424 (1999)
• Alexei Degterev et al. A decade of caspases. Oncogene 22: 8543–
8567 (2003)
• Avi Ashkenazi et al. Death receptors: signaling and modulation.
Science 281: 1305-1308 (1998)
• Howard Y. Chang et al. Proteases for cell suicide: functions and
regulation of caspases. Microbiology and Molecular Biology Reviews
821-846 (2000)
• Helen Blanchard et al. The three-dimensional structure of
caspase-8: an initiator enzyme in apoptosisStructure 7: 1125-1133
(1999)
• Imawati Budihardjo et al. Biochemical pathway of caspase
activation during apoptosis. Annu. Rev. Cell Dev. Biol. 15: 269-290
(1999)
• Stefan J. Riedl et al. Molecular mechanisms of caspase regulation
during apoptosis. Nature Molecular Cell biology Reviews. 5: 897-
907 (2004)
• Jerry M. Adams et al. The Bcl-2 protein family: arbiters of cell
survival. Science 281: 1322-1326 (1998)
• Bruce A.Hay et al. The genetics of cell death: approaches, insights
and opportunities in Drosophila. Nature Genetics Reviews. 5: 911-
922 (2004)
• A. Philchenkov Caspases: potential targets for regulating cell death
J. Cell. Mol. Med. 8(4): 432-444 (2004)