Cell Signalling and Cancer Flashcards
1
Q
define intracellular signalling
A
a set of linked biochemical events connecting a stimulus with a response
2
Q
what does communication between cells control
A
cell behaviour and survival
3
Q
what occurs when signals bind to cell receptors (3)
A
- we have extracellular signals that binds to a receptor on the cell membrane
- this results in different signalling events that lead to:
- proliferation
- Quiescence/ differentiation
- cell death (apoptosis).
- We can also have survival signals which block cell death.
4
Q
what are the two types of signals
A
Signal molecules can bind intracellular or cell surface receptors
- 2 types of signals:
- Small Hydrophobic Signals: that can bypass the cell membrane and go directly into the cell and bind to a receptor within the cell (e.g. hormones)
- Hydrophilic Signals: These bind to the receptors on the cell-surface membrane (majority)
5
Q
how do signalling pathways work and what are the final effects
A
- A signal binds to a receptor
- This activates lots of downstream proteins which then:
- Activate multiple downstream factors to allow for multiple different effects.
- Acts as signal amplifiers to allow for greater effects.
- One of the ultimate effects can be gene expression, metabolism or an effect on the cell cytoskeleton etc
6
Q
what is phosphorylation and what is it mediated by
A
- One key event in cell signalling is the phosphorylation of proteins/lipids
- Phosphorylation is mediated by kinases which act to catalyse phosphorylation
7
Q
what is the process of phosphorylation an example of
A
Phosphorylation is an example of post-translation modification of the protein.
8
Q
what removes phosphorylation
A
Phosphatases remove phosphorylation to return the protein back to normal
9
Q
which 3 residues does phosphorylation normally occur on
A
- Phosphorylation can occur mainly on 3 residues:
- Threonine residues
- Tyrosine residues
- Serine residues.
10
Q
- what are the effects of phosphorylation (3)
A
- Phosphorylation can act as a binding site on the protein for phospho reader proteins.
- It might increase/supress the activity of the protein
11
Q
examples of growth factors that bind to receptor tyrosine kinases (9)
A
- Platelet-derived growth factor (PDGF)
- Epidermal growth factor (EGF)
- Insulin
- Insulin-like growth factor (IGF)
- Transforming growth factor (TGFα & β)
- Nerve growth factor (NGF)
- Vascular endothelial growth factor (VEGF)
- Macrophage-colony-simulating factor (MCSF)
- Ephrins
12
Q
- what are RTKs
A
- These are membrane bound receptors for a certain kind of signal.
- They are tyrosine kinases so they phosphorylate things on tyrosine residues.
13
Q
- describe the basic shape of RTKs
A
- Outside of cell: Bit that binds to a signal - various shapes
- Inside of cell: A domain that acts as a tyrosine kinase.
14
Q
- how do RTKs exist in an inactive state vs active state
A
- When in an inactive state they exist as monomers.
- When in active state they exist as dimers.
- Most are homodimers (identical monomers e.g. 2 EGF receptors) but some can heterodimerize (e.g. a EGF & HERR 2 monomer)
- When in active state they exist as dimers.
15
Q
what are the steps following gf binding to RTK (4)
A
- Growth factor binding to receptor tyrosine kinase causes:
- Conformational change of the receptor
- Dimerisation
- Self-activation of tyrosine kinase activity of the cytoplasmic domain
- Trans-phosphorylation: the tyrosine kinases receptor on the left phosphorylates the receptor on the right and vice versa.
16
Q
what further activates the RTK
A
After transphosphorylation you additionally get autophosphorylation of the RTK to further activate tyrosine kinase activity.
17
Q
- what is a molecule that can activate the MAPK pathway
A
lots of signals e.g. EGF (epidermal growth factor)
18
Q
what does MAPK pathway drive
A
MAPK pathways drive cell growth and one of the signals that drive it is EGF
19
Q
describe a general overview of the MAPK pathway
A
- EGF → binds to EGF receptor → activated → can recruit downstream proteins such as GRB2
- GRB2 acts to activate other downstream proteins such as RAS
- RAS become activated and binds to other factors which results in a downstream cascade allowing for the activation of other kinases → impact on cell growth.
- I.e. MAPK pathways drive cell growth and one of the signals that drive it is EGF
20
Q
what binds to the activated receptor/what does this receptor phosphorylate
A
- When we get activation of the receptor, we get binding of Grb2.
- Grb2:
- Growth factor-receptor bound protein 2
- Contains SH2 domain that binds P-Tyr residues on RTKs
21
Q
what does Grb2 bind to after it has been phosphorylated, what is the effect of this downstream
A
- Grb2 then binds to Sos
- The Assembly of the receptor-Grb2-Sos complex enables recruitment & activation of Ras downstream
- If you do not have Sos then you don’t get recruitment of RAS.
- If you don’t have phosphorylation of RTK then you do not recruit either RAS or Sos.
22
Q
describe the 3 types of RAS
A
- there are 3 types of RAS, encoded by 3 RAS genes:
- k-ras
- h-ras
- n-ras.
- RAS is the most commonly mutated oncogene in tumours.
- It is implicated in ~20 – 30 % of human cancers
- As a result there are lots of efforts to try and drug RAS.
23
Q
describe RAS structure
A
- RAS is a “G-proteins” (GTP-coupled signal transducing protein)
- It contains small lipid group that attaches it to the cell-surface membrane.
- it’s an oncogene (drives cell growth)
24
Q
how is RAS activated
A
- In its inactive form it binds GDP.
- If you get a signal, it exchanges the GDP to GTP and becomes active.
- It can then pass the signal downstream.
25
describe the RAS-GDP → RAS-GTP cycle
- Inactive RAS binds to GDP.
- When we have a growth signal e.g. EGF, it results in RAS exchanging GDP for GTP.
- It does this with help of other proteins called Guanine nucleotide Exchange Factors (GEFS) which help speed up the exchange.
- This results in activation of RAS and the signal is then transduced allowing for an effect e.g. gene expression.
- Then RAS turns itself off.
- It is able to do this because RAS has a GTPase which hydrolyses GTP for GDP
- This is stimulated by other proteins called GTPase-Activating Proteins. (GAPs)
- A lot of mutations found in RAS stop the ability to inactivate itself→ cancer.
26
what helps RAS go from inactive to active form
Guanine nucleotide Exchange Factors (GEFS) which help speed up the exchange of GDP to GTP.
27
what helps RAS go from active to inactive form
intrinsic GTPase which hydrolyses GTP for GDP (stimulated by GTPase-activating proteins)
28
importance of Grb/SOS
The important thing about the Grb/SOS receptor is that they act as a GEF (Guanine Nucleotide Exchange Factor) for RAS so i.e. they activate RAS and so we get signal transduction downstream.
29
what does activated RAS do
- Activated RAS binds another protein called Raf.
- Raf allows the cell to activate downstream kinases such as MEK kinase via phosphorylation.
- MEK then goes on to phosphorylate other signals e.g. it phosphorylates ERK.
- Activation of these 2 sets of kinases (MEK & ERK) ultimately leads to activation of lots of transcription factors e.g. C-Myc, C-jun, C-fos which then act to drive cell proliferation.
30
what type of kinases are MEK and ERK
MEK and ERK are both serine or Threonine (not tyrosine) kinases.
31
how can the MAPK pathway be switched off (4) and the effect
- effect = stops cell growth
- 4 main ways to turn off pathway
1. Remove the signal e.g. stop expressing EGF.
2. Switch off the receptor:
- Therefore, even if the signal is present there is no effect.
- This is done using Tyr phosphatases that remove the phosphorylation signal so receptor becomes inactive
3. If you express lots of GTPase activating proteins then you drive inactivation of RAS and therefore stop the signal.
4. Or you can dephosphorylate the downstream kinases using Serine or Threonine phosphatases.
- In tumours this is often deregulated e.g. RAS mutation
32
which cancers are RAS mutations common in (2)
- RAS is mutated in 90% of prostate cancers.
- It is commonly mutated in ovarian cancer as well.
33
which type of mutation do RAS mutations in cancer always involve and what sites
- RAS mutations in cancer always involve point mutations at specific site (codons 12, 13 or 61)
- Examples:
- codon 12 mutation results in G/T transversion (Glytine → Valine):
- This is typical of carcinogen-induced mutations in smokers or workers with occupational carcinogen exposure
- **Raf** mutations are common in melanoma (80%), lung and colorectal cancers.
- Raf mutations usually occur at V600DE
- Here the mutants stimulate cell proliferation about 10-fold, independent of upstream signals.
- This is because the mutation elevates the activity of RAF to turn on downstream kinases:
- Basal level kinase activity elevated 2-12-fold
- This is why is lots of people try to target RAS and RAF in tumours using small molecules such as inhibitors.
34
MAPK pathway simplified
EGF → RTK → Grb2+Sos → RAS+GTP → RAS+GTP+Raf → MEK+Pi → ERK+Pi → transcription factors ⇒ proliferation
35
what can turn the PI3K pathway on
This pathway can be turned on by lots of signals e.g. insulin.
36
what is PI and what activates PI3K
- PI (Phosphatidylinositol) is a phospholipid found in eukaryotic cell membranes *(on the left in picture)*
- PI is phosphorylated to form PIP, which is in turn is phosphorylated to form PIP2.
- PI3-K catalyses phosphorylation from PIP2 to PIP3
- Then we can recruit other stuff.
- baso: PI → PIP → PIP2 → PIP3 (PI-3k involved in orange part)
- **PI3-K is activated by binding to phosphorylated Tyr residues on RTKs.**
37
Role of PI3K
PI3 Kinase phosphorylates PIP2 to give it PIP3 – 3 phosphorylations on the same lipid.
38
Role of PIP3
acts as a docking site for PDK1 and AKT
39
How is AKT deactivated
Phosphorylation
40
What does activated AKT do
AKT is then able to activate a number of targets with the ultimate effect of driving cell growth, protein synthesis and switching off apoptosis
41
which molecules cause phosphorylation + activation of AKT (2)
- Upon binding, PDK1 phosphorylates and activates AKT.
- mTOR (as part of mTORC2) also plays an important role in activating Akt
- ⇒ so PDK1 + mTORC2 needed to activate AKT
42
when do cells want to switch this pathway off
Normal cells would want to turn off PI3-kinase pathway once they have synthesized the appropriate protein or undergone cell growth.
43
how can we switch off this pathway (4) and the result of this
- The way the pathway is turned off is by either by:
- Remove signal
- Switch off receptor by Tyr phosphatases
- Dephosphorylate PIP3 back to PIP2, how?
- This involves a lipid phosphatase called PTEN.
- PTEN deletion is seen in 20% of tumours – it drives tumour genesis as it means you can’t turn off this pathway at this stage as you get unrestricted PI3 kinase activity.
- Switch off AKT (Ser/Thr phosphatases)
- End result – cell growth goes back to normal.
44
summary image of PI3K pathway
PI → PIP → PIP2 → PIP3 (PI3K) → recruit AKT & PDK1 → protein synthesis & cell gorowth
45
key difference between MAPK and PI3K pathway
PI3K pathway phosphorylates lipids instead of proteins
46
key difference between MAPK and PI3K pathway
PI3K pathway phosphorylates lipids instead of proteins
47
48
why is learning about pathways useful
If we know these pathways are commonly deregulated in tumours how can we target them to kill of tumour cells or improves prognosis.
49
what is stratified/ personalised medicine q
A more effective way of treating cancer by grouping patients according to their genetic mutation(s), and then targeting the signalling pathway to which it contributes.
