Cell Biology Lecture No. 24: Apoptosis
Monday April 8 , 2013
Ultrastructural Features Of Apoptosis:
- Apoptosis (also known as programmed cell death) is a key process that ensures normal embryonic
development and proper tissue homeostasis in adult animals. Programmed cell death is an active
cellular process which requires energy as opposed to necrosis (the passive form of cell death) where
cells simply swell up and die. One of the key features of apoptosis is chromatin condensation as the
nucleus starts to morphological changes. Apoptotic cells tend to shrink somewhat as well, and blebs
(irregular bulges in plasma membrane) start to become visible. As this progresses, the blebbing
continues and the nucleus actually breaks down in smaller fragments known as apoptotic bodies, which
are recognized and engulfed whole by phagocytic cells in order to minimize cellular debris produced.
Apoptosis also involves the cleavage of DNA by DNAases (ICAD) that cut in between nucleosomes
(periodicity of about 200 base-pairs), yielding 200 base-pair fragments.
-C. elegans is present as two sexes (male or hermaphrodite) and has been important for helping with
discoveries about programmed cell death. As a model organism, C. elegans is incredibly advantageous
due to: its small size, transparency (visible under differential interference contrast (DIC) microscopy),
fully-sequenced genome, short reproductive development, and the fact that it has a defined number of
cells for each sex that can be traced through a lineage (ever single cell has been mapped).
Mutations In The Ced-3 Gene Block Apoptosis:
-It was the mutations in the ced-3 gene that were really responsible for shedding light on the process of
programmed cell death. In the early stages of C. elegans development, it was discovered that out of
1,090 newborn cells, 131 cells die during development, resulting in a nematode with exactly 959 cells.
These dead cells are highly refractile and can be detected by DIC microscopy. In certain mutants of the
ced-3 and ced-4 gene (ced = Cell Death Abnormal), it was observed that they contained “un-dead” cells
(more than usual). They also determined that the ced-9 mutants prevented death in cells needed to
Role Of Capasases In Apoptotic Pathways:
-Cysteine-dependent aspartate-directed proteases, or caspases, play key roles in apoptotic pathways.
There are two forms of caspases present: initiator caspases (that cleave inactive pro-forms of effector
caspases, thereby activating them) and effector/executioner caspases (that cleave other protein
substrates within the cell, to trigger the apoptotic process). Effector caspases like ced-3 influence a
variety of features associated with apoptosis: DNA fragmentation (by allowing for DNAase ICAD
activation), fragmentation of nucleus (by cleaving nuclear lamins), membrane blebbing (through
cytoskeletal disruption of important proteins), and fragmentation of Golgi (by influencing Golgi matrix
proteins). The Apoptosome:
-The Apoptosome (also known as the 1.4 megadalton “Wheel of Death”) is a large, disk-shaped
heptamer of Apaf-1 (apoptotic protease activating factor 1), a protein that assembles in response to
apoptosis signals and serves as an activation machine for initiator and effector caspases. Normally, Apaf-
1 exists inactive in the cytosol, but upon binding of cytochrome c (released from mitochondrial pores),
Apaf-1 will form the Wheel of Death. The Apaf-1 heptamer recruits inactive caspase-9 (ced-9 initiator
caspase) monomers to bind to the apoptosome in order for active caspase-9 dimers to be formed.
Caspase-9 can now cleave inactive forms of downstream effector caspases (like ced-3), activating them
for the process of apoptosis to be underway.
The Bcl-2 Family Of Proteins:
-The BcI-2 family is made of proteins that contain functional BcI homology (BH) domains (BH 1-4) and
are divided into three classes: pro-survival members (have extra BH4 domain; form channels in
mitochondrial outer membrane), pro-apoptotic members (missing BH4 domain) and BH3-only proteins
(regulate activity of the other two classes, particularly Bcl-2 and Bax/Bak). Bcl-2 (resembles ced-9) is a
pro-survival protein that inhibits apoptosis and whose high expression was found in human B-cel