Flashcards in Cellular Signaling I Deck (22):
What is the general order of signaling events that occur in a cell
3. G protein effectors that are linked to second messenger generation
4. Regulator of G protein signaling (RGS)
G Protein Coupled Receptor
It resides in the plasma membrane with parts of the protein coming into the cell and sticking out of the cell.
They are the target of the ligands or many ligands interact with their part that sticks out of the membrane
How does signaling takes place in GPCR
A ligand binds to the receptor, causes a conformational change to form a heterotrimer.
This conformation to heterotrimer promotes nucleotide exchange which activates the G protein receptor.
Normally in deactivated state the GPCR has GDP bound to it. But when activated the GDP is released and GTP is now bound to the receptor. This is what activates the protein.
What happens when the GPCR is activated
The alpha intially is bound to the beta and gamma units but when GTP binds to the alpha unit it then dissociates from the beta and gamma subunits. It is now only bound to the GTP and acts downstream on its effector molecules
What are the 2 downstream events
Cellular transcription or cellular response
What is about the inactive bound state that the alpha, beta and gamma subunits do not interact with the effector molecules
Their active sites are shielded away as the active sites of the three subunits are not acting with each other (remember the fist and palm model)
How are GPCRs specific
There are more than 800 GPCRs. They are very specific. They have different alpha, beta and subunits.
Name the different alpha subunits
Galpha s - Activates adenylate cyclase
Galpha i 0 inhibits adenylate cyclase signalling
These two are antagonistic
Galpha q - activates the phospholipase C and finally Galpha 12/13 - activates small G proteins
How can the same receptor produce different affects
Different ligands, different ligand concentrations can produce different results from the same receptor. This has to do with the Kd of the receptor with different ligands, it has different affinities for different ligands. This allows the cell to not respond in a binary fashion where the cell is simply on or off but in a differential fashion, in a graded manner
How do you turn off G protein signalling
It is important to know that when alpha subunit is bound to GDP it is off and when it is bound to GTP subunit it is on.
The signal is turned off as the alpha subunit is also an eznyme and it cleaves the GTP unit into a GDP unit, releasing Pi and then it can reassociate with the beta and gamma subunits.
The activity is very slow.
There are also RGS proteins which bind to the alpha subunit and speed up the GTP hydrolysis.
Basically what is a GPCR
It is a GEF, Guanine nucleotide Exchange Factor
What is an RGS
GAPs, GTP Activating Proteins, any protein that speeds up the rate of hydrolysis of GTP is considered a GAP.
What happens if the ligand is still bound to the receptor
The signalling cascade is initiated again
What are the 2 types of desensitization events. What is the difference in the 2
Hetero and homologous desensitization of the receptors.
In homologous the mechanism only works on the receptors that generated the signal whereas heterologous will work on broad classes of receptors, even on different receptors.
In heterologous desensitization, the signalling doesnt have to come from GPCR but it can come from any signalling cascade that leads to the activation of PKA and PKC that phosphoryltes the GPCRs.
What does a kinase do
Uses ATP to transfer a phosphate group on the substrate
What happens in heterologous desensitization
PKA and PKC phosphorylates the GPCR which inhibits reassocaciation of the alpha, beta and gamma subunits. They are sterically hindered by the phosphorylation sites on the GPCRs.
Also the phosphorylated receptor now has a much lower affinity for the ligand.
What happens in the homologous desensitization of the receptors
It is also a kinase mediated event, it is triggered by the recruitment of proteins called GRKs that phosphorylate that specific receptor.
When the surface of beta and gamma are exposed the interaction between the GRKs are favorable so the GRK binds to these 2 subunits and then it phosphorylates the receptor (at different sites than PKA and PKC).
Then arrestins bind to the phosphorylated receptors and sterically inhibits the reassocaition of alpha, beta and gamm subunits.
Arrestins also cause the internalization of the GPCR, it can either take the receptor off of the cell membrane or the ligand and the whole complex is degraded.
3rd thing can happens is that the receptor is remvoed, the G protein is dephosphorylated and then placed back into the membrane and now the alpha, beta and gamma subunits bind to the receptors in their inactive forms.
Beta Andrenergic Receptors
Binds with the norepinephrine and epinephrine. These are the ones that trigger the fight or flight response.
What does beta blocker do
It binds to the beta Andrenergic receptor. It physically the receptor, sounds like it is non competitive.
What does cholera does to you
Leads to excessive diarrhea. This is done by the the cholera toxin. It ADP ribosylates the Galpha s receptor. In this from the receptor has no ligand bound to it but it is still activates. The alpha subunit however cannot hydrolyze the GTP molecule so this leads to a continuous signalling cascade resulting in excessive diarrhea.
What happens in whooping cough
Remember that it is caused by Pertusis Bordetella. It releases the Pertusis toxin. This toxin leads to ADP ribosylation of the Galpha I (inhibitor) protein, so the alpha subunit cannot bind to the inhibitor molecule (or the receptor). This leads to downstream signal cascading events leading to whoping cough.