Transcript G-Protein Coupled Signal Transduction

CfE AH Biology: Unit 1
2. Proteins
ci) Signal Transduction
Learning Intentions
• To explain what is meant by the term signal
transduction.
• To describe the consequences of signal
transduction with reference to activation of
enzymes or G-proteins, a change in uptake or
secretion of molecules, rearrangement of the
cytoskeleton or activation of proteins that
regulate gene transcription.
Learning intention
To find out about signal transduction and its
possible results in a cell.
Success criteria
I can define’ signal transduction’
I can describe how cell surface receptor proteins connect an extracellular
signal to intracellular responses through a signal transduction pathway
I can describe the possible results of signal transduction to include
- enzyme activation
- G protein activation
- a change in secretion or uptake of molecules
- rearrangement of the cytoskeleton
- activation of proteins that regulate gene expression
Task
Use the scholar site to make notes of signal
transduction under the following headings;
• Cell surface receptor proteins
• enzyme activation
• G protein activation
• a change in secretion or uptake of molecules
• rearrangement of the cytoskeleton
• activation of proteins that regulate gene
expression
Organise your notes under the following
headings Your notes should attempt to
answer the following questions:
Signal Transduction
What is meant by signal transduction?
• Why is this called ‘signal transduction’
• Identify examples of signaling molecules.
• What can be inferred about the nature of signaling molecules that rely on signal
transduction?
• How are membrane proteins involved? (be specific)
G-Protein Coupled Signal Transduction
• Explain what a G-protein is.
• With reference to cAMP, describe, with the aid of annotated diagrams, the specific
cellular response to adrenaline.
Ion Coupled Signal Transduction
• What is an ‘action potential’?
• Explain the role of a named neurotransmitter in generating an action potential.
• Describe how ion-coupled signal transduction results in muscular contraction.
Signal transduction
Receptor proteins in
the cell membrane
convert an
extracellular chemical
signal to a specific
intracellular response
through a signal
transduction pathway.
Signal transduction
•
Where extracellular messages are hydrophilic, eg protein hormones
such as STH (human growth hormone) and insulin or neurotransmitters
such as acetylcholine, they are unable to pass directly through the lipid
bilayer.
•
They are not directly transported across membranes, but instead their
message is transduced, ie converted into an intracellular signal by the
act of binding to target receptors in the plasma membrane.
•
We are going to consider, using case studies, three such systems at the
cell surface:
– ion-channel-coupled receptors, eg ligand-gated ion channels
– G-protein-coupled receptors
– enzyme-coupled receptors.
G proteins: proteins which act as molecular switches - they allow signals from
outside the cell to be transmitted inside (they are involved in signal transduction);
their activity is regulated by their ability to bind and break down GTP (guanosine
triphosphate) to GDP (guanosine diphosphate) - when GTP is bound they are 'on'
and when GDP is bound they are 'off'
Ion channel coupling
•
The next slide illustrates how an action potential travelling along one nerve cell
can be transmitted to another.
•
The junctions between nerves or nerves and muscles are called synapses.
•
The transduction of the message is required.
•
The action potential initiates the fusion of neurotransmitters containing vesicles
with the presynaptic membrane. These chemicals diffuse across the synapse,
triggering ion-gated Na+ channels on the post-synaptic membrane, which initiates
the generation of an action potential in the target cell. This may result in action
potential propagation down the nerve or the triggering of myofibril contraction
in neuromuscular junctions.
•
Neurotransmitters are rapidly reabsorbed after release or broken down.
•
This class of transduction is regulated by controlling the influx and efflux of
ions with associated voltage-gated or ligand-gated channels.
Ligand-gated ion channels
+
Na
Na+
Na+
+
Na+
Na
Initial influx of Na+ depolarises the
membrane. This may trigger local voltagegated channels to open and cause a further
influx of Na+. Such rapid depolarisation
spreads to other Na+ channels, creating an
action potential that spreads to the entire
plasma membrane.
G-protein-coupled signal transduction
G-protein coupled
•
In this system, G-protein-coupled receptors (GPCR) activate a second protein, the Gprotein.
•
GPCRs have seven transmembrane alpha-helix domains.
•
The G-protein is a guanine triphosphate (GTP) binding protein and is bound into the
intracellular face of the plasma membrane.
•
On activation by the GPCR, GTP binds to and activates the G-protein complex.
•
The G-protein then activates a separate target protein. This is usually an enzyme or
ion channel.
•
If the target protein is an enzyme, activation may start a cascade of small
intracellular signaling molecules within the cytoplasm, triggering intracellular events.
•
If the target protein is an ion channel, activation may result in the mediation of
plasma membrane permeability.
Struggling to understand?
• Try this website:
• http://www.nature.com/scitable/topicpage/gpcr14047471
G-protein-coupled signaling case study
Production of cyclic AMP (cAMP)
•
Binding of an extracellular signaling molecule to a G-protein-coupled receptor
(GPCR) stimulates a G-Protein. The G-protein in turn stimulates or inhibits
production of a secondary signaling molecule, cAMP.
•
cAMP is made from ATP in the cytoplasm by a membrane bound enzyme called
adenyl cyclase . Adenyl cyclase activity is stimulated or inhibited by different
CPCRs.
•
cAMP is a second messenger, used for intracellular signal transduction, such as
transferring into cells the effects of hormones like glucagon and adrenaline,
which cannot pass through the plasma membrane.
•
Hormones such as adrenaline, glucagon and thyroid-stimulating hormone (TSH)
act on their appropriate receptor, triggering G-protein and switching on adenyl
cyclase, which is plasma membrane bound, raising cAMP levels.
•
cAMP is destroyed by cAMP phosphodiesetrase.
G-protein-coupled signalling case study
•
cAMP can activate a second group of enzymes called cAMP-dependant kinases.
•
Once bound to cAMP these kinases are activated, releasing active catalytic
subunits. These can diffuse through the nuclear pores where, for instance, they
can phosphorylate gene regulatory proteins called cAMP response element
binding proteins (CREBs), which can then stimulate gene transcription.
•
Raised cAMP can also trigger some of the effects of the following ligands when
stimulated through the appropriate GCPR/G-protein activation.
– Adrenaline is a hormone that has many different effects depending on what
target receptor it stimulates. Many different classes of adrenaline receptor
exist and depending on what type the target cell expresses the effect may
be very different. For example, myocardial tissue in the heart will increase
the rate and contraction of its cells when triggered by adrenaline. Adipose
cells (fat cells) are triggered to break down triglycerides and release fatty
acids and glycerol into the bloodstream.
– TSH acts on thyroid cells, stimulating thyroid hormone synthesis and
secretion.
– Glucagon stimulates liver cells to break down stored glycogen to glucose.
G-protein-coupled signalling case study
Cascades
• At each step the generation of multiple linked intracellular messages creates
interlinked signal systems called cascades, eg molecule 1 stimulates production of
molecule 2 etc.
•
Cascades can greatly magnify the response. Initial binding of one signal molecule
to its GPCR can generate many copies of intracellular signal molecules such as
cAMP or IP3.
G-protein-coupled signalling case study
An example of ion channel regulation by G-protein coupled signalling
•
Acetylchoine, a
neurotransmitter,
can reduce cardiac
muscle activity.
•
Efflux of K+ balances
influx of Na+ into
cytoplasm from
outside, and Ca+ from
intracellular
compartments.
•
This makes it more
difficult for the
target membrane to
depolarise and
trigger cell
contraction.
•
These acetylcholine
GPCR are different
to the ligand-gated
channels found at
neuromuscular
junctions in skeletal
muscle and nerve-tonerve synapses.
http://www.slideshare.net/dominicalwala/si
gnal-transduction-and-second-messengers
Organise your notes under the following
headings Your notes should attempt to
answer the following questions:
Signal Transduction
What is meant by signal transduction?
• Why is this called ‘signal transduction’
• Identify examples of signaling molecules.
• What can be inferred about the nature of signaling molecules that rely on signal
transduction?
• How are membrane proteins involved? (be specific)
G-Protein Coupled Signal Transduction
• Explain what a G-protein is.
• With reference to cAMP, describe, with the aid of annotated diagrams, the specific
cellular response to adrenaline.
• What is meant by a cascade? Explain how the above is an example of a cascade.
• How does acetylcholine cause heart rate to slow down?
Ion Coupled Signal Transduction
• What is an ‘action potential’?
• Explain the role of a named neurotransmitter in generating an action potential.
• Describe how ion-coupled signal transduction results in muscular contraction.
STOP HERE
G-protein-coupled signalling case study
Regulation of the cytoskeleton by G-protein-coupled signalling
•
Two subfamilies of G-proteins, G12/G13 , are involved in cytoskeletal remodelling by
promoting actin–myosin contraction.
•
An associated family Gi is involved with actin fibre polymerisation and membrane
protrusion.
•
Two pathways exist. Stimulation of the G1-linked receptor stimulates the Rac pathway
via the production of the intracellular signal for phosphatidyl inositol triphosphate
(PIP3), which induces polymerisation of actin filaments and the extrusion of the cell
towards the stimulus.
•
Simultaneously, stimulation of the G12/G13 linked receptor stimulates the Rho
pathway, causing actin–myosin contraction.
•
All the intermediate messengers are short lived so the stimulus is polarised to the
surface in contact with the stimulating ligand.
•
The Rho and Rac pathways appear to be antagonistic so if Rac is stimulated at the
front Rho can only be stimulated at the rear. This allows directed movement towards
a stimulus with contraction of cytoskeleton behind and extrusion at the front.
•
The movement of neutrophils towards bacteria has been characterised in this manner.
G-protein-coupled signalling case study
Olfactory sensitivity
•
Olfactory receptors in animals are good examples of GPCRs that are sensitive to
odours.
•
The receptors are localised on modified cilia protrusions from the olfactory
neurons.
•
Different classes of olfactory sensors exist for many different molecules. Each
is encoded by different genes. What they all have in common is their mode of
action.
•
Each GPCR activates an olfactory G-protein, which in turn stimulates the
production of cAMP by adenyl cyclase.
•
Increased cAMP levels stimulate associated cAMP-gated Na+ channels, resulting
in an influx of Na+ ions depolarising the target membrane. If depolarisation is
strong enough an action potential is set up that causes the nerve to fire and send
the action potential down the axon, where it generates a nerve impulse. These
impulses send sensory information about smell to the central nervous system.
G-protein-coupled signalling in
photoreception
•
Vertebrate vision is mediated by photoreceptors coupled to G-proteins.
•
The photoreceptor is coupled to a G-protein that switches off production of
GMP cyclase and results in falling cGMP. This closes associated cGMP-gated
cation channels and results in reduced neurotransmitter release from associated
neurones when the photoreceptor is illuminated.
•
cGMP-gated cation channels are kept open by high levels of cGMP, inhibiting
depolarisation. As cGMP levels fall these channels close.
•
The postsynaptic neurones lose their inhibition and an action potential is
generated along the neurone as depolarisation is promoted. The neurone is said
to be excited. A nerve impulse is then sent along sensory nerves to the CNS.
•
Light detection and its magnification are discussed in detail in a separate
presentation.
Enzyme-coupled signalling
In this form of signal
transduction the binding of a
ligand stimulates a
transmembrane enzyme directly.
These molecules span the
bilayer, existing singly or as two
proteins stimulated by dimers.
Ligand binding stimulates an
enzyme directly rather than via
a G-protein.
The cytoplasmic portion
containing the catalytic site may
have additional domains.
Alternatively, a transmembrane
receptor in permanent
association with an enzyme
undergoes a conformational
change on ligand binding which
results in enzyme activation.
Enzyme-coupled signalling case study
Receptor tyrosine kinases (RTKs)
•
Many enzyme-coupled signalling systems fall into these families.
•
On stimulation by their appropriate ligand they are able to auto-phosphorylate
tyrosine amino acids on their cytoplasmic domains through the activation of their
catalytic site.
•
The phosphorylated tyrosines may in turn provide binding sites for intracellular
signals, which are then activated themselves by becoming phosphorylated or by
undergoing a conformational change through binding to the phosphorylated
tyrosine residues.
•
In turn this may start an intracellular cascade by activated intracellular signals,
including the stimulation of some of the same intracellular signals as G-proteinlinked systems, eg phopholipase-C-linked cascades.
•
Once stimulated and the signal transduced, the receptor–ligand complex is
internalised by receptor-mediated endocytosis, switching off the response. The
recycled receptor minus ligand is then returned to the plasma membrane.
Enzyme-coupled signalling case study:
insulin
–
This hormone triggers many intracellular messengers. Binding to its receptor stimulates
phosphorylation of its receptor and associated signal molecules, triggering PIP3
production and its associated cascade. This gives rise to expression of glucose
transporter Glut 4 on the plasma membrane, increasing intracellular glucose levels.
Additionally, glycolysis and fatty acid synthesis are promoted, as is glycogen synthesis.
–
Diabetes mellitus is a condition associated with impaired insulin function either through
insufficient insulin production (type 1/insulin dependant) or insulin resistance related to
insensitivity or density of insulin receptors (type 2/non-insulin-dependant, NIDDM).
–
Some rare mutations of insulin receptors can give rise to conditions such as Donohue
syndrome (leprechaunism). In this case the receptors lose their affinity for insulin
because of an inherited recessive mutation, giving rise to complete resistance to insulin.
This condition is universally fatal, with most sufferers dying within 2 years of birth.
–
Other mutations have been identified which relate to reduction in insulin sensitivity by
mutations retarding the ability of the cell to recycle the receptor back to the plasma
membrane. This may provide a partial model for loss of insulin sensitivity in type 2
(NIDDM) diabetes.
Enzyme-coupled signalling case study
Insulin action
Effect of insulin on glucose uptake and metabolism
1. Insulin binds to its receptor, resulting in activation of tyrosine
kinase.
2. Activation of intracellular message cascades trigger the
following:
• glut-4 transporter to the plasma membrane and influx of
glucose
• glycogen synthesis
• glycolysis
• fatty acid synthesis.
Enzyme-coupled signalling case study
Cell growth factors
Examples: epidermal growth factor (EGF), fibroblast growth factors (FGFs)
•
Most cell growth factors stimulate cell proliferation and cell growth through
increasing mitosis and DNA activity.
•
The genes encoding the growth factor and their receptors are mainly protooncogenes as mutation of the same can give rise to oncogenes, which promote
uncontrolled cell division.
Enzyme-coupled signalling case study
Epidermal growth factor
• Coded for by a proto-oncogene.
• Its receptor EGFR is an RTK which is also
coded for by a proto-oncogene.
• The diagram shows the multitudinous
ways that the stimulated receptor homo
dimer can generate intracellular signals
and their potential outcomes.
• One effect of EGFR stimulation is the
transcription and expression of EFGR
genes.
• Over-expression of EFGR and EGF is
linked to cancer and is an expression of
oncogenic mutations.
Enzyme-coupled signalling case study
Fibroblast growth factors
•
This family of ligands is involved in the control of cell growth and cell
differentiation, vascularisation, wound healing and embryo growth.
•
They act through an associated family of RTKs called fibroblast growth factor
receptors (FGFRs) in a similar manner to EGFRs.
•
One subclass of receptor, FGFR3, has been associated with bone growth as
FGFR3 stimulation seems to inhibits the formation of bone from cartilage.
•
Mutations in FGFR3 have been associated with achondroplasia, the most common
form of dwarfism (~1 in 25000 live births world wide), which results from
retarded long bone growth.
•
Achondroplasia is a non-sex-linked dominant condition in which sufferers inherit
an allele for mutated FGFR3 that is overly active.
Enzyme-coupled signalling case study
Molecular medicine
•
Current advances in molecular biology are focusing on targeting specific RTKs as
those related to cell growth factors are often over-expressed in cancer cells.
•
Drugs and monoclonal antibodies are in development to try and block such
receptors.
•
Herceptin® and Erbitux® are commercial monoclonal antibodies that block two
types of EGF receptor (Her-1 and Her-2) that can be over-expressed in some
types of breast cancer.
•
Iressa® is a tyrosine kinase inhibitor that may block the action of the overexpressed EGF receptors on some lung cancer cells.
Enzyme-coupled signalling case study
JAK-STAT pathways receptors
•
This is another class of enzyme-coupled signal receptors.
•
They are receptors that always exist as two identical single-pass transmembrane
proteins that exist as homo dimers. Each of their cytoplasmic tails is bound to a janus
kinase (JAK) .
•
Tyrosine residues are phosphorylated on the cytoplasmic domains on ligand binding, as
are associated signal transducer activator of transcription (STAT) molecules.
•
These STAT molecules dissociate to form cytoplasmic dimers, which enter the nucleus
and bind to specific DNA sequences in the promoters of genes that begin
transcription.
•
They differ from RTKs in the directness of their response as few intermediary
messengers are utilised.
•
Growth hormone, erythropoietin and interleukins act by these pathways.
Enzyme-coupled signalling case study
JAK-STAT pathways of growth hormone (STH)
If you are looking for questions go to slide
number 19.