9 - Control of Cell Death Flashcards
Types of cell death
- Programmed cell death
- Necrosis
Programmed cell death
- A physiological process where cells are eliminated during development and other normal biological processes
- E.g. Apoptosis, Autophagy
Apoptosis
Type 1 cell death
Autuphagy
Type 2 cell death
Necrosis
Pathological process after exposure to serious physical or chemical insult
Anoikis
Delayed cell death associated with build up of autophagy vesicles
Cornification
Epithelial cell specific process to produce outer (dead) layer of the skin
Pyroptosis, pyronecrosis
Infection induced death
of macrophages
Necroptosis
‘regulated’ necrosis
Morphological features of apoptosis
- No loss of membrane integrity
- Aggregation of chromatin at the nuclear membrane
- Shrinking of the cytoplasm and condensation of nucleus
- Fragmentation of cell into apoptotic bodies
- Leaky mitochondria due to pore formation
Biological features of apoptosis
- Strictly regulated process
- Energy (ATP) dependent
- Ladder pattern of DNA fragmentation (non random)
- Prelytic DNA fragmentation
- Alteration in membrane asymmetry
Physiological significance of apoptosis
- Evoked by physiological stimuli (growth factors etc)
- Affects individual cells
- Phagocytosis by macrophages or adjacent cells
- No inflammatory response
Morphological features of necrosis
- Loss of membrane integrity
- Swelling of cytoplasm and mitochondria
- Total cell lysis
- No vesicle formation
- Disintegration (swelling) of organelles
Biological features of necrosis
- Loss of regulation of ion homeostasis
- No energy requirement
- Smear pattern of DNA (random digestion)
- Postlytic DNA fragmentation
- Possibility for recovery after reversible injury (unlike apoptosis)
Physiological significance of necrosis
- Evoked by non-physiological disturbance
- Affects groups of cells
- Phagocytosis by macrophages
- Significant inflammatory response
What mediates the events associated with apoptosis
Caspases
Pathways to apoptosis
- Can be extrinsic or intrinsic
- Apoptosis induction –> Initiator caspase activation –> Effector caspase activation –> Death substrate cleavage –> Apoptosis
Regulation of apoptosis
- Positive modulators (pro-apoptosis)
- Negative modulators (anti-apoptosis)
BCL-2 Family
- Inhibitors of apoptosis (anti-apoptosis): Bcl-2, Bcl-xL
- BH3 only (pro-apoptosis): Bid, Bim Bad
- Effectors (pro-apoptosis): Bax, Bak, Bok
Anti-apoptotic Bcl-2 family structure
Four Bcl-2 homology domains (BH) as well as a putative trans
-membrane domain (TM) responsible for their preferred localization at inner membranes.
Effector Bcl-2 family structure
Bax subfamily resemble Bcl-2
closely in structure possessing three out of four (multiple) BH domains
BH3 only Bcl-2 family structure
Only share one BH3 domain with all other Bcl-2 family members.
Balance between anti-apoptotic and pro-apoptotic Bcl-2 family members
Determines if effector members are free to initiate apoptosis
Positive modulators of apoptosis (inducers)
- Cytochrome c
- Apoptosis protease activating factor 1 (APAF1)
- Caspases
- Apoptosis inducing factor (AIF)
- Endonuclease G (Endo G)
- Granzyme A (GrA)
Cytochrome-c
Activates APAF1
APAF1
Critical component of apoptosome, cleaves caspase 9
AIF
Induces chromatin condensation and DNA degradation
EndoG
Facilitates chromatin condensation with AIF
GrA
Serine protease released by cytotoxic T cells
E2F
- Transcriptionally activates many pro-apoptotic genes
- E.g. Bax, Bad, APAF1
Negative modulators of apoptosis (inhibitors)
- Bcl-2 family genes
- Inhibitor of apoptosis proteins (IAPs) that block caspase activation
- Pro-thymosin-α (ProTα) that blocks apoptosome formation
- E1B: acts like Bcl2 to bind Bcl2 family effectors
NF-kB
- Activation leads to enhanced
survival - Inhibition of NF-kB promotes
apoptosis
Reasons for studying apoptosis
Utilised in studies of:
- Immunology
- Embryology
- Aging
- AIDS
- Neurology
- Cancer
Methods for studying apoptosis
- Protease activity (caspase 3)
- Membrane alterations
- DNA fragmentation assay
- DNA strand breaks
Membrane alterations
- Phosphatidylserine translocation to outside of cell when cell undergoes apoptosis
- Annexin V (a phospholipid-binding protein with a high affinity for phosphatidylserine)
DNA fragmentation assay
- Apoptotic DNA ladder
- Gel electrophoresis
DNA strand breaks
TUNEL assay (terminal deoxynucleotidyl transferasemediated dUTP nick end labelling)
Internal pathways of how cancer cells avoid apoptosis
- p53 loss of function
- Rb inactivating mutations
- Myc gene amplification
- Bcl2 activation
- Bax inactivating mutations
- Caspase inactivating mutations
External pathways of how cancer cells avoid apoptosis
- Loss of pro-apoptotic signaling molecules
- Viral infection can prevent apoptosis (e.g. HPV E6 protein blocking p53 function)
- Interaction with other cells (hide from CTLs)
- Interaction with chemicals
Apoptosis and cancer therapy
- Inducing apoptosis of cancer cells is an ideal therapeutic approach as it prevents inflammation and damage due to necrotic cell death and harnesses the cells own apoptotic machinery
- But cancerous cells are resistant to apoptosis
- Need to know affected pathways to produce targeted
therapies
