Signal Transduction in Biological Membranes Flashcards
What are G-Protein Coupled Receptors?
A family of receptors
How do G-protein coupled receptors act?
By altering the activity of effectors (e.g. enzymes / ion channels)
How do G-protein achieve alteration of effectors?
Via the activation of one or more types of guanine nucleotide binding proteins (G-proteins)
What are G-proteins responsible for?
A diverse range of cellular functions, including muscle contraction, stimulus-secretion coupling, catabolic and anabolic metabolic processes and light, smell, and taste perception
What is meant by G-proteins being heterotrimeric?
They are made up of three distinct subunits, alpha, beta and gamma.
Which sub-units in G-proteins bind tightly together?
ß and gamma
How do the ß and gamma sub units in G proteins function?
As a single unit
What does the α sub-unit of G proteins have?
A guanine nucelotide-binding site
What does the guanine nucleotide-binding site do?
Binds GTP and slowly hydrolyses it to GDP (GTPase activity)
How is G-protein present under basal conditions?
At the inner face of the plasma membrane, predominantly in its heterotrimeric form, with GDP bound to the α-subunit
What has a high affinity for G-protein under basal conditions?
Activated receptor (agonist bound)
What occurs when the activated receptor binds with the G-protein?
A protein-protein interaction occurs, leading to GDP being released by the α-subunit and binding GTP in it’s place
What does the activated G-protein receptors act as?
A guanine nucleotide exchange factor (GEF)
What happens once GTP has bound to the α-subunit of the G-protein?
The affinity of the receptor for both α-GTP and the ßγ subunits is decreased
What is the result of the decreased affinity of α-GTP and ßγ subunits?
Both are subsequently released, and are able to interact ith effectors
How is the effector interaction terminated?
By the intrinsic GTPase activity of the α-subunit hydrolysing GTP → GDP
What happens once effector interaction has been terminated?
The affinity of the α-subunit for the ßγ subunit increases, and the ßγ-subunit increases, and the αßγ heterodimer is reformed and awaits reactivation by an agonist-activated receptor to reinitiate cycle
What can G protein can be thought of as?
On/off switches and timers
What is the ‘on switch’ of G proteins?
Receptor-facilitated GDP/GTP exchange
What is the ‘timer/off switch’ of the G proteins governed by?
The length of time taken for GTP hydrolysis
What did our understanding of G-protein mediated systems first come from?
Attempts to understand how hormones such as adrenaline brought about the formation of the second messnger cyclic AMP
Where does the G-protein have an intermediate role?
Where the stimulatory Gs stimulates adenylyl cyclase to produce cAMP
How are other pathways similar to the Gs pathway?
There are other, similar transduction pathways that employ a similar 3-component transduction pathway
What exists antagonistically to Gs pathways?
Inhibitory (Gi) pathways
What do Gi pathways do?
Reduced cAMP levels by inhibiting adenylyl cyclase
How are Gi proteins like Gs proteins?
They have additional effects independent of adenylyl cyclase inhibition, including effects on ion channels and signalling pathways involved in cell growth and differentiation
Have G-protein families that exert their actions on effectors other than adenylyl cylase been discovered?
Yes
What do Gq proteins do?
Preferentially interact with the membrane bound enzyme phospholipid phosphatidylinositol 4,5-bisphosphate (PIP 2 ) to generate the second messengers 1,4,5-triphosphate (InsP 3 ) and diacylglycerol (DAG)
Where is rhodopsin present?
In mammalian retinal rod cells
What is the function of rhodopsin?
Light-sensing
What does rhodopsin do?
Activates a G protein (called transducing, or Gt), which in turn activates a phosphodiesterase enzyme that hydrolyses cylic GMP to 5’-GMP
What is the receptor for adrenaline/noradrenaline?
ß-adrenoreceptor
What is the G-protein for adrenaline/noradrenaline?
Gs
What is the effector for adrenaline/noradrenaline?
Stimulates adenylyl cyclase
What is the physiological response to adrenaline/noradrenaline?
Glycogenolysis, lipolysis
What is the receptor for acetylcholine?
M3-Muscarinic
M2-Muscarinic
What is the G-Protein for acetylcholine when a M3 receptor is used?
Gq
What is the effector for acetylcholine when a M3 receptor is used?
Stimulates phospholipase C
What is the physiological response to acetylcholine when a M3 receptor is used?
Smooth muscle contraction
What is the G-protein for acetylcholine when a M2 receptor is used?
Gi
What is the effector for acetylcholine when a M2 receptor is used?
Inhibits adenylyl cyclase
Stimulates K channel
What is the physiological response to acetylcholine when a M2 receptor is used?
Slowing of cardiac pacemaker
What is the receptor for light?
Rhodopsin
What is the G-protein for rhodopsin?
Gt
What is the effector for rhodopsin?
Stimulates cyclic GMP phosphodiesterase
What is the physiological response to light?
Visual excitation
What are the adrenergic receptors?
α1
α2
ß1
ß2
What G-protein α-subunit does the α1 receptor utilise?
Q
What G-protein α-subunit does the α2 receptor utilise?
I
What G-protein α-subunit does the ß1 receptor utilise?
S
What G-protein α-subunit does the ß2 subunit utilise?
S
What does stimulation of the α1 receptor do?
Stimulates phospholipase C
What does stimulation of the α2 receptor do?
Inhibits adenylyl cyclase
What does stimulation of the ß1 receptor do?
Activates adenylyl cyclase
What does stimulation of the ß2 receptor do?
Activates adenylyl cyclase
What are the cholinergic receptors?
M1
M2
M3
What G-protein α-subunit does the M1 receptor utilise?
Q
What G-protein α-subunit does the M2 receptor utilise?
I
What G-protein α-subunit does the M3 receptor utilise?
Q
What does stimulation of the M1 receptor do?
Activates phospholipase C
What does stimulation of the M2 receptor do?
Inhibits adenylyl cyclase
What does stimulation of the M3 receptor do?
Activates phospholipase C
How many Gα proteins does the human genome encode for?
20
How many Gß proteins does the human genome encode for?
5
How many Gγ proteins does the human genome encode for?
12+
How many combinations of Gα-γ are possible?
Over 1000
How many receptor types that can interact with Gα subtypes are there?
At least 800
How many enzyme/ion channel effectors can be activated/inhibited by interactions with Gα subunits and receptors?
10 or more
How do G-proteins bring about a specific cellular response?
An extracellular signal works via a specific GPCR to activate a single, or small sub-population of G-proteins and effectors in the cell
What can be used in the experimental manipulation of the G-protein cycle?
Cholera Toxin (CTx) and Pertussis Toxin (PTx)
What do CTx and PTx do?
They are enzymes that ADP-ribosylate specific G-proteins
What does CTx do specifically?
Eliminates the GTPase activity of G s α
What is the result of the elimination ofthe GTPase activity of G s α?
G s α is irreversibly activated
What does PTx do specifically?
Interferes with the GDP/GTP exchange on G i α
What is the result in the interference withthe GDP/GTP exchange on G i α?
G i α becomes irreversibly inactivated
What can genetic changes in GPRCs result in?
Loss of function or gain of function mutations
Give 3 examples of conditions caused by mutations in GPCRs
Retinitis Pigmentosa
Nephrogenic Diabetes Insipidus
Familial Male Precocious Puberty
What causesRetinitis Pigmentosa?
A loss-of-function mutation to rhodopsin
What causesNephrogenic Diabetes Insipidus?
A loss-of-function mutation to V2 vasopressin receptor
What causesFamilial Male Precocious Puberty?
A gain-of-function mutation (receptor active without ligand) to the LH receptor
What is adenylyl cyclase?
An integral plasma membrane enzyme that can either be activated (via Gs) or inhibited (Gi) by activation of different receptors
What does adenylyl cyclase do?
Hydrolyses cellular ATP to generate cyclic AMP
What does cyclic AMP do?
Interacts with a specific protein kinase, which in turn phosphorylates a variety of other proteins within the cell to affect activity
What specific protein kinase does cyclic AMP interat with?
Cyclic AMP-dependent protein kinase (PKA)
What can receptors that activate adenylyl cyclase, and therefore increased cellular cyclic AMP levels, cause?
Increased glycogenolysis and gluconeogenesis in the liver
Increased lipolysis in adipose tissue
Relaxation of a variety of types of smooth muscle
Position inotropic and chronotropic effects in the heart
What is phospholipase C?
An enzyme that hydrolyses the membrane phospholipid (PIP 2 ) to IP 3
What is phospholipase C activated by?
Gq
How does IP 3 exert its effects?
By interacting with specific intracellular receptors on the endoplasmic reticulum to allow Ca 2+ to leave the lumen of the ER and enter the cytoplasm
What is cyclic GMP phosphodiesterase?
A specialised mechanism found in the photoreceptive cells of the retina
What doescyclic GMP phosphodiesterase do?
Regulates the breakdown of the second messenger cyclic GMP phosphodiesterase by Gt
What happens once a receptor has productively interacted with a G-protein?
The binding of the agonist is weakened, and agonist-receptor dissociation is likely to occur
What is the receptor susceptible to whilst activated?
A variety of protein kinases that phosphorylate the receptor and prevent it from activating further G-proteins
What is the problem with the activated receptor being susceptible to phosphorylation?
It compromises an important part of the receptor desensitisation phenomenon observed for most of the GPCR
What may the active lifetime of α-GTP be limited by?
Cellular factors that stimulate the intrinsic GTPase activity of the Gα subunit
What does the enzymatic activities in the cell favour?
The basal state
What is the result of enzymatic activities in the cell being such that the basal state is favoured?
Cells contain high activity enzymes that metabolise second messengers, rapidly returning their levels to the basal
How is the effect of second messenger/protein kinase activation opposed?
Enzymatic cascades are activated downstream that oppose the effect
What can the rate at which the sinoatrial node fires an action potential be affected by?
Ach release by the parasympathetic nerves
What is the predominant receptor in regulation of chronotropy in the heart?
M2 muscarinic cholinoceptors
What does activation ofM2 muscarinic cholinoceptors cause?
An increase in the open probability of K+ channels via Gi
What does an increase in plasma membrane permeability to K+ cause?
Hyperpolarisation, slowing the intrinsic firing rate, resulting in a negative chronotropic effect
What is inotropy?
The force of heart contraction
What can influence inotropy?
Sympathetic innervation of the cardiac ventricles (and/or circulating adrenaline)
How is a positive inotropic effect in the heart bought about?
Activation of the ß-adrenoreceptors (predominantly ß1) increases the open probability of voltage operated calcium channels via Gs.
Gs both interact directly with the VOCCs, and indirectly via cyclic AMP → PKS → phosphorylation and activation of VOCCs
The influx of Ca brings about a positive inotropic effect
How is arteriolar vasoconstriction bought about?
Sympathetic release of noradrenaline acts on α1-adrenoreceptors to stimulate phospholipase C and IP3 production via Gq
IP3 releases ER Ca, and initiates a contractile response
What can pre-synaptic GPCR influence?
Neurotransmitter release
Give an example of where pre-synaptic GPCR influence neurotransmitter release?
Pre-synaptic µ-opiod receptors can be stimulated, either by endogenous opiods or by analgesics such as morphine to couple Gα1 proteins
How is neurotransmitter release reduced by GPRCs?
The Gßγ subunits are liberated from the heterodimer and interact with VOCCs to reduce Ca entry, thus reducing neurotransmitter release
