Apoptosis

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APOPTOSIS

• Programmed cell death
• Specific biochemical signature (ex. phosphatidylserine "flips" to outer surface of plasma membrane)
• Does not induce inflammation

Important in a Variety of Processes

• Organism development
• Cell number and organ size
• Quality control during development
• Removal of damaged cells

Cellular Changes

• Cytoplasm condenses
• Nucleus becomes misshapen
• Chromatin condenses along the nuclear envelope
• Cell eventually fragments into blebs
• Phosphatidylserine on the blebs indicates to phagocytic cells that apoptosis is occurring
• Phagocytic cells clear the cellular debris

Two Paths of Apoptosis

1. Extrinsic Pathway (using the Fas pathway as an example)

a) Trimeric Fas ligand on another cell binds to Fas death receptor
b) Intracellular domain of Fas receptor recruits and activates FADD (Fas associated death domain)
c) Activated FADD recruits initiator procaspases such as procaspase-8 or -10 (complex is referred to as death-inducing signaling complex or DISC)
d) Complex formation activates the procaspases which then activate executioner caspases
e) Executioner caspase activation leads to apoptosis

2. Intrinsic Pathway

a) Apoptotic stimulus activates BH3-only protein
b) BH3-only protein blocks the activity of Bcl-2 protein
c) Without Bcl-2 activity, BH123 proteins are able to oligomerize and cytochrome c is released from the intermembrane space of mitochondria
d) Cytochrome c in the cytoplasm activates Apaf1 proteins which form a heptameric complex
e) Apaf1 complex recruits initiator procaspase-9
f) Activated caspase-9 activates executioner caspase
g) Executioner caspase activation leads to apoptosis

Bcl-2 Family of Proteins

• Based on which specific domains are present in the protein

1. Anti-apoptotic Bcl-2 proteins (ex. Bcl-2 or Bcl-XL)
2. Pro-apoptotic BH123 proteins (ex. Bax or Bak)
3. Pro-apoptotic BH-3 proteins

Full-Length Text

• Here we will learn about apoptosis, a type of programmed cell death.

• First, let's start a table to explore some key concepts of apoptosis.

• Denote that apoptosis is important for a variety of processes, which include: organism development, cell number and organ size, quality control during development, and removal of damaged cells.

• Denote that apoptotic cells have specific biochemical characteristics that allow them to be recognized as undergoing apoptosis.
  • This plays a role in specific processes that we will explore shortly.
  • They do not induce inflammation.

First, let's explore what apoptosis looks like.

• Draw a healthy cell.

• Include: the nucleus, a mitochondrion, Golgi stacks and endoplasmic reticulum.

• Show that the nucleus comprises diffuse chromatin.

• Indicate that the cell receives a signal to undergo apoptosis, and then let's explore the changes that occur.

• Show that the cell size shrinks; specifically indicate that the cytoplasm condenses.

• Redraw the organelles.

• Show that the nucleus is misshapen.

• Show that chromatin condenses along the inner side of the nuclear envelope.

Next, show that the cell further deteriorates as follows:

• Show that it further misshapes and fragments with what we identify as blebs that surround it.

• Indicate that there is breakup of the nuclear envelope, cell and nuclear fragmentation, and blebbing.
  • Within some of these blebs, draw organelles and bits of condensed chromatin.

• Introduce a phagocytic cell, which engulfs a bleb.

• Then, draw the phospholipid phosphatidylserine on the surface of the bleb being phagocytosed.

• Indicate that phosphatidylserine "flips" during apoptosis from the inner side of the plasma membrane, so it is now on the outer side.

• Write that phagocytic cells clear the cellular debris of apoptotic cells.

• The phagocytic cells use increased levels of phosphatidylserine on the surface of apoptotic cells as a marker.

Now let's explore the two main pathways of apoptosis, the extrinsic pathway and the intrinsic pathway.

• To begin, let's draw a section of a cell.

• On one end, draw a section of the plasma membrane.

• On the other, draw a section of a mitochondrion.

• We'll see that both pathways run through a series of steps that conclude with what we introduce, now, are:
  executioner caspases, which trigger apoptosis.

The extrinsic pathway is shorter than the intrinsic, so we'll begin with it.

As an example, we'll use the Fas death pathway.

• Show the trimeric Fas death receptor traverse the plasma membrane.

• Draw the plasma membrane of another cell with the trimeric Fas ligand and show the Fas ligand bind the Fas death receptor.

• Show that this causes the intracellular domain of the Fas receptor to recruit the FADD (Fas associated death domain) adaptor protein to bind to the intracellular tail of the Fas receptor.

• Show that this causes the FADD proteins to recruit initiator procaspases such as procaspase-8 or -10.

• Show that we use the term DISC (for death-inducing signaling complex) to refer to the FADD/procaspase complex.

• Show that formation of this complex activates the procaspases, which then cleave and activate the executioner caspases.

• The activation of the executioner caspases leads to apoptosis.

Now let's explore the intrinsic pathway of apoptosis.

• Within the intermembrane space of the mitochondrion, draw molecules of cytochrome c, which are important signal molecule during the apoptotic pathway.
  • Other molecules exist in the intermembrane space, which we ignore for simplicity.

Let's take a moment to learn the Bcl-2 family of proteins, which is integral to the intrinsic pathway of apoptosis.

• Write that it is made up of three classes of proteins based on the specific domains present in the protein: the anti-apoptotic Bcl-2 proteins, the pro-apoptotic BH123 proteins, and the pro-apoptotic BH-3 only proteins.

• Draw a few molecules of BH123 protein traversing the outer membrane of the mitochondrion: the main BH123 proteins in mammalian cells are Bax or Bak.

• Draw these proteins joined together (oligomerized).
  • This is the active form of the proteins which increases the permeability of the mitochondrial membrane, though whether this oligomer functions as a channel or in some other way is still debated.

• Draw an anti-apoptotic Bcl2 protein.
  • These proteins, such as Bcl2 or Bcl-XL, inhibit the oligomerization of BH123 proteins to block apoptosis from occurring.

• Now indicate that the cell recognizes an apoptotic stimulus.
  • This stimulus will activate the pro-apoptotic BH3-only proteins.

• Draw the activated BH3-only protein.
  • There are many different BH3-only proteins which respond to different apoptotic stimuli.

• Indicate that the activated BH3-only protein inhibits the activity of the Bcl2 protein.

• Indicate that without the blocking action of the Bcl2 proteins, the BH123 proteins are able to oligomerize.

• Draw molecules of cytochrome c in the cytoplasm, and for simplicities sake, indicate that they traveled through the BH123 "channels".
  • Cytochrome c activates the Apaf1 protein which causes it to form a heptameric complex.

• Draw the activated Apaf1 complex.
  • This complex recruits the initiator caspase, procaspase-9.

• Draw the Apaf1 complex with procaspase-9.
  • This causes the activation of caspase-9.

• Indicate that the activated caspase cleaves and activates executioner caspases.

• Indicate that these activated executioner caspases, which are different from the executioner caspases activated during the extrinsic pathway, lead to apoptosis.

This ends our diagram on apoptosis.

UNIT CITATIONS:

1. Campbell, N. A. & Reece, J. B. Biology, 7th ed. (Pearson Benjamin Cummings, 2005).

2. Alberts, B., Johnson, A., Lewis, J., Raff, M., Roberts, K. & Walter, P. Molecular Biology of the Cell, 5th ed. (Garland Science, 2008).

3. Alberts, B., Bray, D., Hopkin, K., Johnson, A., Lewis, J., Raff, M., Roberts, K. & Walter, P. Essential Cell Biology, 3rd ed. (Garland Science, 2010).

4. Lodish, H., Berk, A., Kaiser, C. A., Krieger, M., Scott, M. P., Bretscher, A., Ploegh, H. & Matsudaira, P. Molecular Cell Biology, 6th ed. (W. H. Freeman and Company, 2008).

5. Marieb, E. N. & Hoehn, K. Human Anatomy & Physiology, 10th ed. (Pearson, 2016).

6. Polinsky, K. R. Tumor Suppressor Genes. (Nova Publishers, 2007).

7. Coleman, W. B. & Tsongalis, G. J. The Molecular Basis of Human Cancer. (Springer Science & Business Media, 2001).