Cell Signalling Flashcards
What are the 3 stages of cell signalling?
Reception
Transaction
Response
What are 3 places signals originate?
Synaptic - release of neurotransmitter from pre-synaptic cleft
Paracrine - further in the distance
Endocrine-signalling molecules pass through vessels
Describe stage 1: reception
signal detected when a ligand binds to a receptor.
info transmitted by a conformational change
Describe stage 2: Transduction. What happens when PK1 is activated?
- signal transduction pathway
- amplification of the signal
- first messenger activated GPCR -> activates G protein -> activated membrane-bound enzyme
- enzyme catalyses the production of 2nd messenger -> activate the kinase
activated PK1 phosphorylates PK2, phosphorylates PK3, phosphorylates a protein, to either increase or decrease activity
What 3 receptors are involved in triggering the kinase cascade?
GCRPs, Guanylyl cyclases, catalytic receptors
What are heterotrimeric G-proteins? What are the 3 types of G-Alpha subunits and what do they do?
as - increase cAMP production (activates PKA which phosphorylates a cascade of enzymes)
ai/o - inhib of cAMP production (inhibits production of cAMP and PKA)
aq - increase DAG (which leads to kinase cascade) and IP3 production (which leads to increase calcium release).
How does Gαq Signalling lead to calcium release?
In inactive state G alpha q has GDP bound, when activated it releases the GDP protein and accepts GTP which causes dissociation of alpha subunit from gamma-beta subunit.
Activating G alpha q coupled receptor activates the phospholipase C Beta (PLCβ) enzyme which then cleaves the PIP2 molecule into diacylglycerol (DAG) and IP3 which are secondary messengers.
IP3 binds with the ligand gated ion channel in the endoplasmic/sarcoplasmic reticulum to release calcium. Meanwhile the diglyceride molecule activates phosphokinase C (PKC)
how much greater is ca2+ concentration outside than inside the cell?
10,000x
What causes calcium changes?
the result from agonist stimulation or depolarisation
What protein pumps move calcium out of the cytoplasm?
SERCA, PMCA, NCX
Activation of ion channels causes calcium to flow into the cytoplasm from:
Ca2+ stores (IP3P, RyR)
Extracellular fluid
Ligand-gated ca2+ channels
voltage-gated calcium channels
What is the mechanism (8 steps) of muscle contraction?
- GPCR activates Gαq → 2. IP3 produced, and opens IP3R on SR → 3. Ca2+ released from SR → 4. Ca2+ binds and activates calmodulin (CaCM) → 5. CaCM binds and activates the myosin light chain kinase (MLCK) → 6. The activated MLCK catalyses the transfer of phosphate to myosin heads, activating myosin head ATPases → 7. Phosphorylated myosin heads form cross bridges with actin, shortening occurs = 8. Muscle contracts
How does Gαs Signalling lead to muscle relaxation, by phosphorylation of MLC phosphatase?
GPCR receptor in its active conformation activates the bound Gαs coupled receptors = releases GDP replaced by GTP.
Gαs is released and interacted with adenylate cyclase which converts ATP into cyclic AMP (cAMP = second messenger).
cAMP activates protein kinase A (PKA).
PKA catalyses the transfer of phosphate to a phosphatase enzyme = activates the phosphatase = removes phosphate from the myosin light chain.
Therefore MLC is not active.
How does Gαs Signalling lead to muscle relaxation, by inhibiting the function of MLCK?
GPCR receptor in its active conformation activates the bound Gαs coupled receptors = releases GDP replaced by GTP.
Gαs is released and interacted with adenylate cyclase which converts ATP into cyclic AMP (cAMP = second messenger) .
cAMP activates protein kinase A (PKA). PKA catalyses the transfer of phosphate to myosin light chain kinase.
This prevents the kinase from working
What role does G-alpha-i play in smooth muscle?
Inhibits/ reduces the production of cAMP.
How does Nitric Oxide cause muscle relaxation?
Activated nerve terminal releases Ach, Acetylcholine binds to endothelial cells M3 muscarinic G-protein receptor.
Gaq activates PLC, which cleaves PIP2 into IP3 and DAG.
IP3 releases intracellular calcium from the ER.
Calcium activates CAM.
CAM in endothelial cells activate NO synthase, which converter argenine into NO.
NO can now diffuse into the smooth muscle fibre and activates guanylate cyclase.
This converts GTP into cGMP, which then activates PKG
PKG phosphorylates a phosphatase (MLCP)
MLCP takes a phosphatase off MLC
causing muscle to relax
What are 2 sources of NO?
NO endothelium and NO donors
Compare how we can therapeutically target smooth muscle with an agonist (oxytocin and in an inhaler), antagonist (Ligand-type-calcium-channel blocker)
Oxytocin induces labour by the Gaq pathway.
LTCC blockers dilate blood vessels to reduce high blood pressure.
Agonists in inhalers relax smooth muscle in asthma via the gas pathway.
What is desensitisation/tachyphylaxis/tolerance and what are 4 possible causes of GPCR desensitisation?
mechanism underlying loss of signal
- enzymes (like kinases) will add phosphate residues onto the receptor, inhibiting it
- down-regulation of the receptor (via internalisation and reduced expression)
- depletion of mediators
- Increased metabolic breakdown of ligand or secondary message
Draw the GPCR desensitisation pathway (7 steps)
- ligand binding
- g protein signalling
- arrestin recruitment
- b-arrestin recruitment
- receptor internalisation: goes into endosome, ligand dissociates, phosphate removed, arrestin dissociates
- receptor degradation or
- receptor recycling
How does desensitisation affect the drugs we use?
Morphine binds to opioid receptors –> arrestin binds to a receptor and leads to desensitisation
Methadone does not lead to tolerance as it triggers internalisation and recycling.
