What do caspases look like?

Caspases have an overall structure consisting of two heterodimers of subunits.

First of all there are a few points about the structure of caspases to consider:

They have been evolutionarily conserved and can be tracked back to nematodes and insects from C.elegans to humans _[18][19].
They are therefore homologous in other species and have similar functions _[18][19]
A cysteine residue resides in the caspase active site _[20]
Their mission is to cleave at specific aspartic acid residues on other proteins _[20]
The last two points show how they got their name - Cysteine Aspartic acid Specific Proteases _[20]
Caspases are synthesised as procaspases which consist of two subunits:
- A large subunit - p20
- A small subunit - p10 _[20]
Active caspases are produced by cleaving these procaspases
An active caspase consists of two heterodimers of p10/p20 with two active sites _[20]
However the caspase 8 above has been kept in procaspase state and is composed of p12 and p18 subunits
Caspase 8 is an initiator caspase and contains a Death Effector Domain (DED) which contains CD95 apoptosis receptor and CD95 ligand _[21].

Functional aspects of Caspases

Executioner otherwise known as effector caspases need an autocatalytic process in order to become active
They function by travelling throughout the cytoplasm cleaving several molecules_[21]
Examples of structure they can cleave are:
- The cytoskeleton e.g. actin
- Enzymes e.g. poly-ADP-ribose polymerase
- Signalling molecules e.g. Akt-1
- Inhibitors of endonucleases e.g. ICAD_[21]
Effector caspase processes are the cause of cell shrinkage and what eventually kills the cell_[21]

Receptors of Death!

There are 8 death receptors and each one functions in a different way _[22].

Some are described below:

Death receptor 1 is known as tumour necrosis factor receptor 1 (TNFR1)
It has many biological effects such as:
- inflammatory responses,
- cytotoxicity,
- hepatocyte proliferation,
It is induced by TNF therefore is dependent on its signalling
However TNF's activity is limited on TNFR2.[22]

Death receptors possess an internal death domain and use this to employ adapter proteins

Examples of the adapter proteins it employs are shown below:

TNFR-associated death domain (TRADD),
Fas-associated death domain (FADD),
caspases such as caspase-8, which when activated induces apoptosis _[23].

There are two states the death domain can be present itself and these are membrane-bound or soluble. These state in which it presents itself is very important in regards to Fas and TNF-related apoptosis inducing ligand (TRAIL) _[24].

In terms of soluble presentation FasL can be cleaved from the membrane to form a soluble compound (sFasL). This lowers cytotoxic activity yet does not prevent the death receptor binding to it [24].

However if sFasL is cross-linked with anti-flag antibodies to resemble a membrane bound ligand it results in a drastic increase in cytotoxicity. It has been shown that more than 3 Fas molecules need to be combined to begin apoptosis efficiently _[25].

The Cascade!

The caspase cascade is a big process you may already have heard of depending what year of study you are in.

There is a video on this website situated on the signalling mechanisms page.

This section will explain the cascade in more detailed information so you can gain a better understanding.

First of all there are two pathways that are interlinked as the image shows on the homepage.

There is the intrinsic pathway is activated by various stimuli such as DNA damage or cytotoxicity. This pathway acts through the mitochondria, managed by the Bcl-2 family the Bcl-2 family that inhibit apoptosis prevent Bak and Bax (pro-apoptotic proteins) from piercing the membrane of the mitochondria _[26].

However if there is inhibition of Bcl-2 then oligomerisation of Bak and Bax is caused and they form a channel in the mitochondrial membrane which in turn is used to release cytochrome c by exocytosis into the cytosol. Once released cytochrome c pairs with APAF-1 and ATP. This amalgamation of molecules creates a landing platform for the activation of procaspase-9, known as the apoptosome _[17].

After the apoptosome is formed procaspase-9 is cleaved and therefore caspase-9 is activated which then travels through the cytosol to cleave procaspases-3,-6 and -7 known as the downstream effector caspases. These caspases are essential for the recipe of apoptotic cell death and they go on to break down the cell to form membrane enclosed vesicles to be disposed of by macrophages in the process of phagocytosis _[27].

A video of the intrinsic pathway is embedded below it is around 30 minutes long however it gives a good explanation and goo visual representations.

_[28].

The extrinsic pathway is a different story this begins outside the cell and is stimulated by death receptors. induction of apoptosis in this pathway is by the death-inducing signalling complex (DISC). Within DISC is FADD which draws in initiator caspases such as caspase-8 or -10 _[29].

TNFR1 has a different mechanism to Fas and TRAIL-R and instead leads to the formation of two complexes [29]. The first complex is created within the cell membrane. TNFR1 is then brought into the cell by endocytosis and complex 2 is formed, which is analogous to DISC _[30].

When caspase-8 or -10 are activated they move on to activate the effector caspases. Bid a pro-apoptotic protein is then cleaved by caspase-8 which in turn kickstarts the mitochondrial apoptosis pathway [30].