Signal transduction Flashcards
What are the two main types of kinases?
- Tyrosine kinase
* Serine/threonine kinase
What are the two types of GTP binding proteins?
- Trimeric G proteins
* Monomeric GTPases
Describe how G protein coupled receptors work
Ligand binding activates a G protein which in turn activates or inhibits another protein - often this is an enzyme that generates a specific second messenger
Describe the structure of G protein coupled receptors
All G protein coupled receptors hace 7 membrane spanning regions with their amino termini on the extracellular face and their carboxy termini on their cytoplasmic face of the plasma membrane
• Many cell surface receptors are coupled to trimeric signal transducing G proteins that bind to either GDP or GTP
What is the mechanism of action of a G protein coupled receptor?
- Binding of a ligand to the receptor changes the conformation
- The receptor binds to Gα protein
- GDP becomes displaced and GTP binds to Gα
- This triggers Gβγ dissociation, activating downstream pathways
- GTP on Gα hydrolyses to GDP in seconds
- Re-association of Gα with Gβγ and the inactivation of adenylate cyclase
What is needed by a system
- Receptor
- A transducer (G protein)
- An amplifier (adenylate cyclase) that generates large amounts of second messenger
How many types of Gα subunit are there?
20
Give 3 examples of Gα subunit
- Gαq (Gq) Stimulates phospholipase C
- Gs: Stimulates adenylate cyclase, increase in cAMP
- Gi: Inhibits adenylate cyclase, decrease in cAMP
What are phospholipase isoforms?
Proteins which possess distinct domain structures but catalyse the same reaction (liberation of IP3 and DAG from PIP2)
What is the resting calcium level?
~100nM
What is active calcium level?
0.5-1 μm
What is protein kinase C?
- At least 12 isoforms
- Most are catalytically inactive, soluble proteins in the cytoplasm
- Phosphorylates a wide variety of substate proteins on the threonine and serine residues
- Has substrates in the cytoplasm and some can translocate to the nucleus to phosphorylate nuclear proteins
Describe how PKC is activated
- A rise in cytosolic calcium causes PKC to bind to the cytosolic leaflet of the plasma membrane
- One bound, it can be activated by the membrane associated DAG and/or Ca2+
Which receptor does adrenaline bind to?
β2 adrenergic receptor
Describe the effect of adrenaline binding to the receptor
- Intracellular concentration of cAMP is increased as the receptor couples to Gas
- cAMP is synthesised within cells from aTP by the enzyme adenylate cyclase
- cAMP is degraded by the enzyme cAMP phosphodiesterase
What does signal transduction result in?
An amplification of a signal
How can PKC act in the nucleus?
- PKA can activate transcription of genes containing the cAMP response elements of CREs in their promoter
- CREB binds to this sequence and activates transcription of downstream genes
- When CREB is unphosphorylated it is inactive, only when it is phosphorylated can it activate transcription
What is the cholera toxin?
- Oligomeric complex which after cleavage becomes active and enters the intestinal epithelial cells to stimulate Gas
- Causes an over production of cAMP
- results in a release of water and ions including Na+, K+, Cl- and HCO3- into the lumen of the small intestine
- This leads to rapid fluid loss, causing dehydration
How does the pertussis toxin act?
• Inhibits Gi, leading to an overproduction of cAMP in th lung epithelia
What activates RTKs to control cell proliferation?
Insulin like growth factors
Describe how RTKs are activated
- Signal molecule in a dimer form binds
- Kinase activity is stimulated
- Tyrosine is phosphorylated
- Signal is relayed by activated signalling proteins into the cells interior (bound to phosphorylated tyrosine)
Describe how RTKs activate Ras
- Adaptor proteins bind to the phosphorylated tyrosine
- Causes Ras activating protein to activate Ras
- GDP dissociates and GTP binds, activating Ras
What does Ras do?
Regulates cellular proteins:
• Proliferation
• Cytoskeletal dynamics
• Membrane trafficking/vesicular transport \
What targets Rac/Rho subgroups of Ras?
Clostridial cytotoxins
Cancer and Ras
Ras mutated to be constantly bound to GTP in 20% of cancers
What does Ras activate? and what does this cause?
The MAPK pathway
• Changes in protein activity
• Changes in gene expression
What is the EGFR receptor pathway?
- EGFR (epidermal growth factor receptor) is a RTK activated by TGFα (Transforming growth factor alpha)
- The receptor activates Ras via Grb2/SOS proteins
- Ras activates Raf (kinase), which in turn stimulates gene transcription via other kinases (MEK and ERK).
What are mutations in EGFR, Ras and Raf associated with?
Tumourgenesis
These cause overexpression and/or hyperactivation of the respective proteins
