Signalling Mechanisms in Growth Flashcards
What is the quiescent phase?What organ cell is normally found in this?
G0, hepatic cells
Which proto-oncogene has a key role in the entry into the cell cycle
C-Myc
What is C-Myc
porto-oncogene that has a key role in the entry into the cell cycle
What is the concentration of C-Myc in G0
Low
When does the concentration of C-Myc rise rapidly
when cell division is triggered
What does C-Myc do
controls a lot of genes involved in the cell cycle progression
What are the key components of signalling pathways (3)
- Regulation of enzyme activity by protein phosphorylation (kinases)
- Adapter proteins
- Regulation by GTP-binding proteins – class of signalling molecules that are prevalent in many tumours.
what is a mitogenic GF? Example?
a growth factor that induces mitosis in a cell (e.g. hepatocyte growth factor)
Describe mitogenic GF stimulation of signalling pathways starting with a GF arriving and binding to its receptor
The growth factor arrives and binds to a receptor (usually tyrosine kinase type receptors)
It then acts via small GTP-binding protein (Ras)
This triggers a kinase cascade
This activates genes required for progression of the cell through the cell cycle- this is slower because it requires transcription and translation to take place
One of the genes triggered early in the kinase cascade is c-Myc
C-Myc is triggered by …
Growth factors activating RAS which triggers a kinase cascade trigger c-Myc
What is the end action of a phosphorylated receptor protein tyrosine kinase (RPTK)
Recruits adaptor and signalling protein
E.g. of an adaptor protein?
Grb2
Describe the process of phosphorylating a RPTK at a molecular level? Residues of what amino acid is phosphorylated?
- The receptors normally sit on the plasma membrane as monomers but most growth factors are dimeric
- When dimeric growth factor binds to two receptor tyrosine kinase molecules, it brings them closer together and their respective tyrosine kinase domains cross-phosphorylate one another
- Tyrosine kinases use the gamma phosphate of ATP to phosphorylate tyrosine residues in proteins
What do the phosphorylated tyrosine domains of RPTK serve as
docking sites for ADAPTOR proteins
What serves as docking sites for ADAPTOR proteins
The phosphorylated tyrosine domains of RPTK
what genes are mutationally activated or overexpressed in many breast cancers
EGFR/HER2
EGFR/HER2 mutationally activation/ overexpression is indicative of what
Breast cancer
What antibody treatment for breast cancer is there?
Anti-her2 receptor tyrosine kinase antibodies
What is Ras
Small GTP binding protein
Grb2 is an…
Adaptor protein
Domains of Grb2?
1 SH2 domain and 2 SH3
What does an SH2 domain bind to
phosphorylated tyrosine
What domain on a Grb2 binds to phosphorylated tyrosine
SH2
What does an SH3 domain bind to
Proline rich regions
What domain on a Grb2 binds to Proline rich regions
SH3
e.g. of a Small GTP binding protein
Ras
What is the on form of ras
GTP bound
What is the off form of ras
GDP bound
What does Sos catalyse
the exchange of GDP for GTP on Ras
the exchange of GDP for GTP on Ras is catalysed by what
Sos
How is Ras self-regulated
is self-regulated, it can hydrolyse GTP to GDP to turn itself off
- intrinsic GTP hydrolysis capability
What stimulates hydrolysis of the GTP on Ras
- intrinsic GTP hydrolysis capability and
GTPase activating proteins (GAPs)
What turns ras on
Sos
What turns ras off
itself and GTPase activating proteins (GAPs)
What can mutations of Ras cause and how
Cancer, mutated to be constantly on and GTP bound as Ras is the main GTP binding protein in GF stimulatory pathways
Which domains on Grb2 bind to the RPTK and which to Sos
SH2 to RPTK and SH3 to Sos
Where can you find Sos
bound to Grb2 (adaptor protein)
Describe RPTK signalling to Ras starting with phosphorylation of the receptor
- When the RPTK becomes activated you get phosphorylation of the receptor
- Then Grb2 binds to the phosphorylated tyrosine domains, bringing Sos close enough to the membrane to activate Ras
- Sos allows the exchange of GDP for GTP in Ras This changes the conformation of Ras, making it into an activate state that can signal downstream and can allow propagation the signal
What does Ras need to bind to to signal downstream
The plasma membrane
2 forms of ONCOGENICALLY ACTIVATE Ras?
V12Ras
L61Ras
How is V12 Ras oncogenic
prevents GAPs from binding to Ras, meaning Ras can’t turn off easily
How is L61 Ras oncogenic
mutation inhibits the intrinsic GTPase activity of the Ras protein preventing GTP hydrolysis
Which form of Ras does this describe: mutation inhibits the intrinsic GTPase activity of the Ras protein preventing GTP hydrolysis
L61 Ras
Which form of Ras does this describe: prevents GAPs from binding to Ras, meaning Ras can’t turn off easily
V12 Ras
What does GTP bound Ras do
- GTP-bound Ras binds to a kinase and activates it and that kinase activates several other kinases
- The kinase cascade involved in the growth stimulatory signalling is called the ERK cascade (extracellular signal-regulated kinase cascade)
- This type of kinase cascade is also known as a MAPK cascade (mitogen-activated protein kinase cascade)
What starts the ERK cascade (extracellular signal-regulated kinase cascade)
- GTP-bound Ras binds to a kinase and activates it and that kinase activates several other kinases
What starts the MAPK cascade (mitogen-activated protein kinase cascade)
- GTP-bound Ras binds to a kinase and activates it and that kinase activates several other kinases
In the ERK/MAPK cascade, what is the first kinase activated by and what is it called
First kinase activated by Ras is called Raf or MAPKKK
In the ERK/MAPK cascade, what is the second kinase activated by and what is it called
Activated by Raf or MAPKKK
Second is MEK or MAPKK
In the ERK/MAPK cascade, what is the third (final) kinase activated by and what is it called
Activated by MEK or MAPKK.
The final one is ERK or MAPK
What does the ERK/MAPK cascade achieve
Phosphorylates a number of proteins and changes their activity
What is the key group proteins ERK/MAPK cascade phosphorylates
gene regulatory proteins (transcription factors)
What do gene regulatory proteins (transcription factors) do once phosphorylated
regulate gene expression
What phosphorylates gene regulatory proteins (transcription factors)
ERK/MAPK cascade
one of the most important genes activates by the ERK/MAPK cascade
c-Myc
What are Myc and Ras genes examples of
ONCOGENES
What’re Cyclin-dependent kinases (Cdks)?
serine-threonine dependant kinases
what do cyclins do
Activate cdks
What happens to the cyclin once a cdk is activated? what is this a form of
it degrades, form of regulation
when do cyclin levels rise
become expressed during mitosis and then go away (degraded), and then come up again in the next mitosis
What activates Cyclin-dependent kinases (Cdks)?
Cyclin
explain the variety of cyclins
- There are different cyclin-Cdk complexes that trigger different events in the cell cycle
What is cyclin that promotes mitosis
mitotic cyclin
What is cyclin that promotes DNA replication
S phase cyclin
what is the m-phase promoting factor
controls the progression through mitosis- the Cdk is being activated by a mitotic cyclin
Cdk is being activated by a mitotic cyclin is called …
m-phase promoting factor
What is an example of an activated CDK
m-phase promoting factor, MPF
or start kinase
What does an activated M phase promoting factor do
- MPF phosphorylates proteins involved in mitosis e.g. nuclear lamins (cause breakdown of the nuclear envelope)
What do nuclear lamins do
- Breakdown of the nuclear envelope is caused by phosphorylation of nuclear lamins
What does an M phase promoting factor consist of
Cdk1 and mitotic cyclin
What does Cdk1 and mitotic cyclin form
MPK
What forms start kinase
Cdk2 and an G1 cyclin (e.g. cyclin E)
What does start kinase do, what’s its most important function
phosphorylates substrates needed for that phase, most important protein that is phosphorylated by start kinase is RETINOBLASTOMA
is a Cdk cyclin complex fully active?
No
What happens to make a Cdk cyclin complex become fully active (3)
phosphorylation by Cdk activating kinase
inhibitory phosphorylation by Wee 1
removal of the inhibition by Cdc25
What puts an activating phosphorylation of Cdk1
CAK puts an activating phosphorylation on Cdk1
What puts an inhibitory phosphorylation of Cdk1
Wee1
What takes off the inhibitory phosphorylation of Cdk1
Cdc25
What does CAK do
puts an activating phosphorylation on Cdk1
What does Cdc25 do
Takes off the inhibitory phosphorylation of Cdk1
What does Wee1 do
puts an inhibitory phosphorylation of Cdk1
When is Cdk1 dephosphorylated. to activate it
end of interphase
What positive feedback effect does MPF have
- Active MPF is able to phosphorylate Cdc25 to increase its activity
- This is a form of positive feedback that drives mitosis
- As soon as you get some active Cdk1 then you activate more Cdc25, which in turn leads to more dephosphorylation etc.
What two features of CDK cyclin complexes/cell cycle allow the same Cdk to be used in different stages of the cell cycle
Different cyclins change the complex:
when cyclin binds to Cdk is actually changes the substrate specificity so that it can phosphorylate different substrates depending on which cyclins are bound to it
- It also changes substrate accessibility – The substrates available in G1/S will be different to those available in S
What CDKs cyclin complex is G1/S
Cdk2 and cyclin E
Cdk2 and cyclin E regulated which stage of the cell cycle
G1/S
What CDKs cyclin complex is S
Cdk2 and cyclin A
Cdk2 and cyclin A regulates which stage of the cell cycle
S
What is one of the key jobs of c-Myc
stimulate transcription of CYCLIN D
What does Cyclin D do
CYCLIN D Triggers cell cycle by ACTIVATing Cdk4 AND Cdk6
What does cyclin D bind to
Cdk4 and Cdk6
What part of the cell cycle is Rb present
All of it
What does Rb have bound to it in it’s inactive state
transcription factors called E2F
What does the Cdk4/Cdk6-Cyclin D complex target
Rb
What causes the formation of the Cdk4/Cdk6-Cyclin D complex
Myc induction
What does the Cdk4/Cdk6-Cyclin D complex do to Rb
starts to phosphorylate the Rb protein and as it becomes phosphorylated it loses affinity for E2F and releases E2F
What does E2F do, what is one of its main targets
- E2F transcription factors can then bind to proteins of genes in the nucleus that are involved in cell cycle progression
- One of the E2F targets is the gene for cyclin E- the next cyclin that is required for cell cycle progression
Rb acts as a ‘X’ in the cell cycle….
Brake
Summarise the activity of Rb, starting with Myc being activated
- Myc turns on cyclin D, which complexes with Cdk4/6
- This Cdk4/6-cyclin D complex phosphorylates Rb and allows E2F to start being released into the cytoplasm and then this stimulates the production of cyclin E
- Cyclin E forms a complex with Cdk2 which further phosphorylates Rb leading to further increase in E2F
- This increase in E2F concentration means that it can now bind targets with lower affinity
- This cycle continues throughout the cell cycle
Names of the 2 CDK inhibitor families?
- INK4
- CIP/KIP
When are INK4 CDK inhibitors active
in G1- they inhibit Cdk4/6 by displacing cyclin D
How do INK4 CDK inhibitors work
When are INK4 CDK inhibitors active
When are CIP/KIP CDK inhibitors active
S Phase- they inhibit Cdk/cyclin complexes by binding to them
How do CIP/KIP CDK inhibitors work
S Phase- they inhibit Cdk/cyclin complexes by binding to them
Genes that are commonly lost in cancers are ….
tumour suppressor genes
Genes that are commonly overexpressed/over-produced in cancers are ….
oncogenes
