Signal Transduction (14-19) Flashcards
Do cells live in isolation?
No
→ all cells interact with their environment and neighbours
→ receive and act on signals from beyond their plasma membrane
signalling controls all aspects of our behaviour: growth, differentiation and development, metabolism
Do bacteria signal?
Yes - they have membrane proteins that act as information receptors
signals: pH, osmotic strength, food, oxygen, light, noxious chemicals, predators, competitors
What is the universal signalling pathway?
single → receptor → response
What does each stage of signal transduction mean?
Signal → information from beyond the plasma membrane
Receptor → information detector
Amplification → small signals are (usually) amplifies within the cell to give a large response
Response(s) → chemical changes and/or changes in gene expression
→ you can’t just turn a system on - feedback pathways are required
What are ligands?
A chemical messenger produced by signalling cells that bind to receptors in or on target cells
agonists → ligands that stimulate pathways - most natural ligands (e.g. serotonin)
antagonists → ligands that inhibit pathways - most drugs (e.g. antihistamines)
What is direct contact (type of signalling)?
A protein (ligand) on the signalling cell binds to a protein (receptor) on the target cell
→ target cell response
→ common in tissue development (e.g. cell-cell contact controls eye development in Drosophila)
What is gap junction (type of signalling)?
Gap junctions: exchange small signalling molecules and ions, co-ordinating metabolic reactions between cells
→ ability of cells to inform other cells of intracellular content
→ gap junctions are made and broken during embryo development
→ electrical synapses use gap junctions between neurones for rapid transmission
What do gap junctions do in electrical synapses?
Clusters of gap junctions connect the interior of two adjacent neurones
→ enable the passage of electrical current carried by ions + intracellular messengers + small metabolites
(connections between pre- and post- synaptic membranes)
What is autocrine signalling?
The ligand induces a response only in the signalling cell (self stimulation)
→ signalling cell expresses ligand and receptors
→ most autocrine ligands rapidly degrade in extracellular medium
e.g. eicosanoids: autocrine ligands derived from fatty acids and exert complex control (aggregation of platelets in immune system, integration of pain/inflammatory response, contraction of smooth muscle)
also common feature of cancers → auto-production of growth hormones stimulates cell proliferation
What is paracrine signalling?
The ligand induces a response in target cells close to the signalling cell
→ architecture of cells limits ability of ligand to diffuse
→ destroyed by extracellular enzymes and internalised by adjacent cells
e.g. paracrine signalling occurs at neuromuscular junctions
What occurs during paracrine signalling at neuromuscular junctions?
- a nerve impulse is transmitted
- stimulates movement of synaptic vesicles, which fuse with the cell membrane
- acetylcholine is released
- acetylcholine stimulates channel opening, allowing ion exchange
- the muscle twitches, acetylcholinesterase degrades acetylcholine
(response to receptor binding → opening of ion channels)
What is endocrine signalling?
The ligand is produces by endocrine cells and is carried in the blood, inducing a response in distant target cells (ligands often called hormones)
→ human endocrine tissues: pituitary, thyroid and adrenal glands, pancreas, ovaries, testes
Is the distinction of signals always absolute?
No → ligands can belong in more then one class
e.g. acetylcholine → neurotransmitter in neuromuscular junction - paracrine manner, as a hormone - endocrine manner
How is signalling specificity provided through cell-type specific expression?
a. Certain receptors are only present on certain cells
b. Molecules downstream of the receptor are only present in some cells
How is signalling specificity provided through high-affinity interactions?
There is a precise molecular complementarity between ligand and receptor, mediated by non-covalent forces
(affinity: ability of a molecule to find and interact with another molecule)
How is the rate of receptor ligand interactions determined?
R (receptor) + L (ligand) → RL (receptor-ligand couples)
association rate → 2 reactants so defined by second order, conc. of both reactants = k+[R][L]
units = M^-1s^-1
dissociation rate → 1 reactant so defined by first order rate, conc. of the one reactant considered = k-[RL]
units = s^-1
At equilibrium what happens to the rate of association and dissociation?
They are equal
k+[R][L] = k-[RL] → k+/k- = [RL]/[R][L]
Keq = k+/k- = [RL]/[R][L]
units = M^-1
How can the ‘dissociation equilibrium constant’ (Kd) be described?
The reciprocal of Keq →
Kd = k-/k+ = [R][L]/[RL]
units = M
→ favoured by biologists due to familiar units (affinity can be described in terms of conc.)
What is the meaning of low/high affinity?
High affinity → specific interactions
e.g. sptretavidin (Kd ~ 10^-14M for biotin) can scavenge biotin when its very low in conc. - strongest non-covalent bond in nature
Low affinity → less specific interactions
e.g. bovine (Kd ~ 10^-6M for plastic) will interact non-specifically with most things - ideal blocking agent
Is binding just two molecules sticking together and remaining together?
No → binding is a dynamic process - a mixture of association and dissociation
A + B ⇌ AB
off ⇌ on
→ higher affinity = longer time spent together
How are signals amplified?
Enzyme cascades
→ produce amplifications of several orders of magnitude within milliseconds
What is signalling desensitisation?
When a signal is present continuously the signal transduction pathway becomes desensitised
→ when the signal falls below a threshold system regains sensitivity
e.g. walk from bright sunlight into a dark room
What is signalling cross-talk?
Many signalling pathways share common components leading to potential cross-talk
What is signalling integration?
If multiple signals are given, the cell produces a unified response
+ response % - response → net response depends on the integrated output of both receptors
combination of cross-talk and integration → signal responses can be very complex
What are the classes of receptors?
Receptors with intrinsic enzyme activity
Receptors linked to protein kinases
Receptors coupled to target proteins via a G protein
Intracellular receptors
Receptors that are ion channels
What are receptors with intrinsic enzyme activity?
Some receptors are enzymes - binding of ligand activated the enzyme activity
→ prototype for this group is the insulin receptor
What hormones regulate blood glucose levels?
Following intake and ingestion of food, blood glucose can rise dramatically
(pancreatic)
Insulin → lowers blood sugar levels
Glucagon → raises blood sugar levels
(adrenal)
Epinephrine → raises blood sugar levels
Cortisol → raises blood sugar levels
Where are insulin and glucagon produced?
In the pancreas
→ acing cells have digestive function
→ the islets of Langerhans secrete hormones
α cells: glucagon
β cells: insulin
δ cells: somatostatin
How is the insulin receptor made?
IR is made as a single protein, from a single gene
→ following translation the subunits
1. enter ER membrane
2. associate into dimers
3. exported to cell surface, via Golgi
4. during intracellular transport: processed by cleavage into α and β subunit
5. at plasma membrane - displayed as trans-membrane proteins
How does insulin activate the insulin receptor (IR)?
Insulin signalling starts at the plasma membrane
→ insulin binding stimulates allosteric change in IR
→ brings cytosolic domains close
→ leads to auto-phosphorylation
→ results in activation
What is a first/primary messenger/ligand?
An extraceullar substance that binds to a cell-surface receptor and initiates signal transduction that results in a change in intracellular activity
receptor → a protein that binds and responds to the first messenger
What is the first step in insulin signalling?
Activated IR phosphorylates and activates the insulin receptor substrate-1 (IRS-1)
→ signal has been transduced from the extracellular side of membrane to the intracellular side + has been transferred to a soluble protein in the cytosol
What does activated IRS-1 do?
Activated IRS-1 is bound by adaptor molecules Grb2 and Sos
→ signal transferred to Sos - a guanine nucleotide exchange factor (GEF)
What does Sos do?
Sos converts (GDP-bound) Ras to active (GTP-bound) Ras
→ activates Ras
What does activated Ras do?
Activated Ras recruits Raf kinase to the membrane - activates its protein kinase activity
→ Raf phosphorylates and activates MEK kinase
→ MEK kinase activates mitogen-activated protein kinase (MAPK)
→ signal amplified across the cytosol through a MAPK cascade
What does ERK (MAP kinase) do?
Migrates to the nucleus
→ alters gene expression modulating expression of ~100 insulin responsive genes + cyclins/CDKs required for cell division
→ insulin is a growth factor
How is the insulin pathway linked to the epidermal growth factor (EGF) signalling pathway?
Grb2 and Sos are common to both EGF and insulin signalling
→ activation of EGFR and IR recruits the same MAPK cascade - the same genes are modulated in the downstream response
How is insulin receptor substrate-1 (IRS-1) bi-functional?
It binds to Grb2 → MAPK cascade - gene expression changes (Ras dependant)
+
recruits and activates phosphoinositide 3-kinase (PI-3K) to the cytosolic face of the plasma membrane → glucose regulation (Ras independent)
→ growth and glucose metabolism co-ordinated via insulin signalling - not much point growing without food supply
What does PI-3K do?
Phosphorylates the membrane lipid PIP2 to produce PIP3
→ PIP3 is a second messenger
second messenger: never a protein, a small metabolically unique molecule whose conc can change rapidly - relay signals from receptors to target molecules in the cytoplasm or nucleus
What does PIP3 recruit?
PDK1 (PIP3-dependant protein kinase)
→ activates protein kinase B (PKB) aka Akt
What are the 2 responses to insulin?
Growth factor:
Phosphorylation IRS-1 (amplifies) → Grb2 + Sos adaptors recruit Ras → Signal transduction via amplifying MAPK cascade → gene expression changes
Glucose regulator:
Phosphorylation IRS-1 (amplifies) → signal propagation + amplification - conversation of membrane lipids → amplification - lipid dependant kinase activation of PKB → up regulation of glucose entry into cells + glycogen production
What are the cellular responses to insulin?
Within minutes → increased uptake of glucose into muscle cells and adipocytes + altered glucose metabolism by modulation of enzyme activities
→ don’t require new protein synthesis, occur low insulin levels ~10^9 - 10^10M
Within hours → increased expression of: liver enzymes that synthesise glycogen, adipocyte enzymes that synthesis triacylglycerols, genes involved in mitogenesis in some cell lines
→ require continuous exposure to ~10^8M insulin
How is the Ras-independent insulin signalling pathway terminated?
A PIP3-specific phosphatase (PTEN) removes the phosphate at the 3 position of PIP3 - converting it into PIP2
→ PDKI and PKB can no longer be recruited to the plasma membrane, shutting of signalling through PKB
What is diabetes mellitus?
When insulin signalling goes wrong
Type I → Insulin dependent diabetes mellitus
→ deficiency in insulin production, early onset, responds to insulin injection
Type II → Non-insulin dependent diabetes mellitus
→ failure to respond to insulin, typically late onset, associated with obesity
symptoms of both: excessive thirst, frequent urination (polyuria), excretion of large amounts of glucose in the urine (glycosuria)
results in high blood sugar levels
What does activated PKB do?
(in muscle and adipose tissues)
Activated PKB stimulates the movement of the glucose transporter GLUT4 from internal membrane vesicles to the PM
→ increasing uptake of gluocose
also mediates the conversion of excess glucose into glycogen (in the liver/muscles) and to triacylglycerols (in adipose tissue)
Do continual high sugar levels desensitise insulin signalling?
Study done on reversal of type 2 diabetes
→ suggested in part that desensitisation of insulin responses followed prolonged exposure to high sugar levels
What are protein kinases?
Phosphorylate their targets
How is body mass measured?
Estimated by Body Mass Index (BMI) = weight (kg) / height x height (m^2)
BMI > 30 obese
BMI 25-30 overweight
BMI < 25 normal
→ gives poor estimates for tall/short and athletic people, nevertheless we’re stuck with it
What is the approximation of obesity?
Obesity is the result of intake of excess calories, more than are consumed by the body’s activities
excess food + low exercise = obesity
What is the lipostat theory?
Postulates that fat-borne factors act on the brain to regulate energy homeostasis, controlling levels of adiposity
→ eating behaviour is inhibited when body weight exceeds a certain value (‘the set point’)
→ energy consumption increases above the set point
inhibition of eating behaviour and increased energy should therefore reduce body mass back to the set point
Feeding → fat synthesis → adipose tissue grows → beta-oxidation of fats → energy and heat
adipose tissue → feedback signals (I’m full) inhibits feeding
→ feed forward signals (let’s use it) stimulate oxidation of fatty acids
What is the evidence for the Lipostat model of body mass regulation?
A soluble factor called LEPTIN is released into the bloodstream by adipose tissue
→ leptin binds leptin receptors in the hypothalamus and changes feeding behaviour
→ action at a distance - endocrine signalling
How was leptin discovered?
First identified in mice
→ leptin corrected the Lepob/Lepob (mutant) obese gene phenotype
What does leptin do?
Leptin is released by adipose tissue
→ carries a message that fat reserves are sufficient (enough food)
→ binds receptors in specific neurons (the anorexigenic or appetite-reducing neurons) in the hypothalamus - stimulates signalling cascade that results in release of a hormone (alpha-MSH) that modulates nervous transmission
effects: suppression of appetite, stimulation of the sympathetic nervous system (do more) - increases blood pressure, heart rate, thermogenesis
What is the first part of leptin signalling?
2 leptin bind to the extracellular binding domains of Lep-R
→ dimerises Lep-R
→ generates sites for the recruitment of JAK
JAK: soluble kinase
What is Janus kinase (JAK)?
Cytosolic, non-receptor tyrosine kinases that transduce cytokine-mediated signals via the JAK-STAT pathway
→ possess two near-identical phosphate-transferring domains
→ one domain - kinase, other regulates the kinase activity of the first
What does JAK do?
Phosphorylates Lep-R
→ phosphorylated Lep-R recuits STATs
→ JAK phosphorylates the STATs
STAT proteins: (Signal Tranducer and Activator of Transcription) latent transcription factors
What does phosphorylated STAT do?
The phosphorylated STATs dimerise, exposing their nuclear localisation signals (NLS)
→ dimer enters nucleus - active transcription factors that modulate gene expression
precursor for alpha-MSH is made + processed + produced
→ signals to the brain ‘stop eating’
What is the cross-talk between the insulin and leptin signalling pathways?
Leptin signals to brain - ‘stop eating’ but also:
signals in liver and muscle cells; making them are sensitive to insulin (IRS-2 link)
What is erythropoietin (EPO)?
A home cytokine that controls the development of erythrocytes from precursor cells in the bone marrow
→ produced in the kidneys
→ kidney measures haematocrit (vol of blood occupied by RBC) - normal 45%
Under hypoxic conditions → erythropoietin secreted from kidneys to increase production of red blood cells
How is erythropoietin used?
Analogues with iron injections - possible treatment for anaemia caused by cancer treatment only in:
→ women receiving platinum-based chemo for cancer of ovaries - blood haemoglobin level of 8mg/100ml or lower
→ people who have very severe anaemia and can’t receive blood transfusions
…believed to have been used in cycling e.g. 1998 Stuart O’Grady Tour de France - doping with EPO
How does erythropoietin signal?
Via a JAK-STAT pathway using STATs
→ following JAK autophosphorylation EPO signalling can access a Ras-dependnat pathway
→ modulation of gene expression in the nucleus
→ growth and development can be co-ordinated
What is the basic structure of G-protein coupled receptors?
Extracellular domains → E1 and loops E2-4
Trans-membrane domains → TM1-7
Cytosolic domains → loops C1-C3 and C4 tail
C4 has a lipid anchor
(G-protein: guanine-nucleotide binding proteins → family of proteins that act as molecule switches - involved in transmitting signals)
How do ligands bind to G-protein coupled receptors?
GPCRs fold into a tertiary structure resembling a barrel
→ the 7 helices forming a cavity within the plasma membrane
→ cavity is a ligand-binding domain for small ligands (often covered by loop E2)
Ligand binding changed the relative orientation if the trans-membrane helices → a twisting motion
→ also reveals amino acids in the cytosolic domains for activating heterotrimeric G-proteins
Bulky ligands (proteins and peptides) may also bind to the extracellular loops or the N-terminus
What is a heterotrimeric G-protein?
A trimer of α, β and γ subunits (Gα, Gβ and Gγ)
→ inactive when bound to GDP (no guanine nucleotide) but active when bound to GTP
Ligand binding alters receptor shape which induces nucleotide exchange (GDP of Gα replaced with GTP)
What occurs in G-protein coupled receptors following activation of Gα?
Gα activation → G-protein dissociates from the receptor producing:
→ Gα-GTP monomer
→ Gβγ dimer
(these modulate activity of other intracellular proteins)
How are G-protein coupled receptors regulated?
Gα has slow hydrolysis activity → regenerates the inactive from of the α-subunit (Gα-GDP) allowing re-association with the Gβγ dimer to form the resting G-protein (which can bind to a GPCR and await activation)
What is the difference between a type 1 and 2 error?
Type 1 error → believing a falsehood
Type 2 error → rejecting a truth
What is the flight or fight response?
A very rapid (acute stress) response to:
→ a perceived harmful event
→ an attack
→ a threat to survival
physiological responses include:
→ acceleration of heart and lung action, paling or flushing, general effect on sphincters, dilation of blood vessels in muscles, inhibition of tear and saliva production, dilation of pupils and tunnel vision, shaking, liberation of metabolic energy sources (particularly fat and glycogen) for muscular action
How is the flight or fight response stimulated?
Release of epinephrine (adrenaline) and cortisol from the adrenal glands
Cortisol → increases blood sugar through gluconeogenesis, surpasses the immune system
Epinephrine → hormone & neurotransmitter acts on nearly all body tissues, binds to a variety of adrenergic receptors (adrenergic receptors are GPCRs)
→ binding to α-adrenergic receptors inhibits insulin secretion by the pancreas, stimulates glyconenolysis (liver & muscle), stimulates glycolysis in muscle
→ binding to β-adrenergic receptors triggers glucagon secretion in the pancreas, increased lipolysis by adipose tissue
Together these effects lead to increased blood sugar and fatty acids providing substrates for energy production within cells throughout the body
What is the second messenger of the flight or fight response?
cAMP → epinephrine binds β-adrenergic GPCR receptor, Gαs activates + stimulates adenylate cyclase
→ results in increase of cAMP levels
epinephrine (via cAMP 2nd messenger)
→ induces glycogen breakdown in skeletal muscle (more glucose available)
→ induces contraction in cardiac muscle (increased heart rate)
2 principle aspects of flight or fight mediated by cAMP levels
How do cAMP and Protein Kinase A (PKA) react together?
cAMP has a variety of target proteins - one is PKA
→ PKA binds 2 cAMP and becomes activated
→ activated PKA phosphorylates target proteins like: transcription factors, ion channels and a variety of other enzymes
pathways acting through cAMP usually give the same response - major control being cell-type specific expression of cAMP targets
→ epinephrine and glucagon both raise blood glucose levels and induce triglyceride breakdown - increasing availability of fuel sources
How is epinephrine signalling terminated?
Adenylate cyclase acts as a GAP on Gαs
(GAP: GTPase-activating protein)
→ adenylate cyclase converts the Gα to its inactive form - the response of adrenaline’s short-lived
Why are G-protein coupled receptors important?
→ largest class of cell-surface receptors
→ the human complement is >1000
→ >200 have been identified formally, and functions are ascribed to >100
→ GPCRs mediate a very wide variety of responses
→ >500 known molecular targets (downstream)
→ ~60% of all drugs target GPCR-mediated pathways
→ current market > US$900billion/year
What is glucagon signalling?
Increases blood glucose conc.
→ via a GPCR and a second messenger (cAMP)
→ stimulated glycogen breakdown (liberates more sugars)
How does cholera toxin (CTx) traffic to the endoplasmic reticulum of target cells)
Binds to cell surface of intestine cells via GM1
→ retrograde trafficking via endoscopes and the Golgi complex to the ER
→ disulphide bond between CTxA1 and CTxA2 is broken by PDI
→ BiP keeps CTxA1 soluble until it dislocates across the ER membrane in an unfolded form
→ CTxA1 refolds in the cytosol
How is CTxA1 (cholera toxin) toxic?
CTxA is an ADP-rebosylase that transfers a ribose group onto a specific arginine on Gαs
→ modifies Gαs locked ON permanently (cannot degrade its GTP) → adenylate cyclase is turned ON permanently
Cellular [cAMP] rises >100 fold above normal
→ activates CFTR membrane channels
→ increased efflux of Na+ and water into intestine (diarrhoea) - death from dehydration often follows
How do we receive light?
In the vertebrate eye, light passes through the neural layer through the cell bodies of the light receptor cells (rods and cones) and acts as a signal in the discs of photoreceptive membrane in the ‘outer segment’ of the retina
How are light receptors structured?
The inner and outer segments of a photoreceptor cell is a primary cilium (primary cilia extend from the surface of most vertebrate checks - and act as signalling organelles)
Rods: outer segments contains ~1000 discs not connected to the plasma membrane
→ each disc is a closed sac of membrane with embedded photosensitive rhodopsin (GPCR) molecules
What is rhodopsin?
A visual pigment
→ specialed GPCR made of opsin (the GPCR protein component)
→ linked to 11-cis-retinal - a prosthetic group that is the chromophore or light-absorbing group
How does retinal capture light?
Stimulates conformation change → cis-trans isomerisation
1. alternating single and double bonds form a ‘polyene’ with a longe unsaturated network of e- that can absorb light energy
2. light absorption causes cis-trans isomerisation around the C12 and C13 bond
3. the N of the lysine moves 5 A (0.5nm)
light energy is converted into atomic motion within a few picoseconds
How does light capture activate the GPCR?
Light absorption by retinal alters the conformation of the GPCR (inactive rhodopsin → activated metarhodopsin II)
Metarhodopsin stimulates nucleotide exchange on the alpha-subunit of a specific hetertrimeric G protein called transducin
What does transducin do?
Transducin Gαt activates cGMP phosphodiesterase
→ removes cGMP from cGMP-gated ion channels at surface of retinal cells
What happens when the cGMP-gated ion channels on the surface of retinal cells close?
Light closes the cGMP gated ion channels reducing influx of Ca++
→ closing hyper-polarises (more negative) the membrane
→ a light stimulus has been converted to a change in the electrical charge (potential) across a membrane
How sensitive are mammalian rhodopsin?
Really very sensitive!
→ peak absorbance at 500nm
→ a rod cell can respond to a single photon
→ ~5 such responses lead the brain to register a flash of light
Under high sensitivity are rod cells more of less sensitive?
Less
→ under high light intensity, rod cells are inhibited, and less sensitive to small changes in light sensitivity
→ there are 7 phosphorylation sites, the higher the light density the more sites are phosphorylated - higher phosphorylation the lower the ability to activate transducin (less cGMP)
What does arresting do?
Binds fully phosphorylated rhodopsin
→ stops activation of tranducin
(rhodopsin kinase and arresting also inhibit other GPCRs - not just rhodopsin)
What 3 mechanisms make rods insensitive to high light?
- prolonged cGMP-gated channel closure
- phosphorylation of opsin reduces tranducin activation
- arrestin binding to phosphorylated opsin stops transducin activation
Summarise rod phototransduction:
- cis→trans isomerisation of retinal, light energy → atomic motion, activating rhodopsin (receptor stimulated)
- signal transduction activates transducin - each metarhodopsin activates ~500transducins (first signal amplification)
- ~500 cGMP phosphodiesterase are activated
- ~10^5 cGMP (secondary messenger) molecules are removed - ion channel closure (second signal amplification)
- loss of cGMP leads to closure of ~100-250 Na+ channels, up to 10^7 Na+ ions no longer enter cytoplasm
- end result: light energy converted to a change in membrane potential - stimulates nerve impulse
How do rod cells and cone cells differ?
Rod cells → respond to low light intensities, peak abs 500nm
Cone cells → respond to higher light intensities and different wavelengths
human colour vision relies on 3 visual pigments with peak abs at: 414-426nm (blue), 530-532nm (green) and 560-563nm (red)
each cone cell expresses only one visual pigment
Which species has the record for receptors for colour sensitivity?
Mantis shrimps (Neogondactylus oestedii)
→ 12 receptors for colour sensitivity; others for intensity and polarisation (20 in total)
How to cephalopod and human eyes differ?
Cephalopod (octopus)
→ light strikes retina directly
→ there is no blind spot
→ retina has only rod cells
→ wide (w-shaped) pupils - focus wavelengths by changing the depth of their eyeball (altering distance between lens and retina)
Human eye
→ light strikes retina indirectly
→ there is a blind spot
→ retina has rods and cones
→ round pupils - can contract to give us sharp vision, will all colours focused on the same spot
What are dichromats?
Colour blindness
→ we typically see 3 colours
→ dichromats have difficulty distinguishing similarly sized objects where lightness varies in an unpredictable manner
What did John Dalton postulate?
The vitreous humour in his eyes was tinged with a blues pigment that absorbed red light - normal vision = clear humours
→ when he dies eyes dissected - vitreous ‘perfectly pellucid’: colourblindness not arise due to blue pre-retinal filter
In 1995 retinal samples taken + PCR + sequenced
→ Dalton had a deletion of the gene encoding the MW (green_ visual pigment - genetic dichromat
Is there selective pressures for human trichomacy?
Did colour vision evolve with production of yellow, orange and red pigments
→ dichromats can’t pick ripe fruit - selection pressure
Counter-selection?? - spotting things trichromats can’t
rationele for why colour blindness maintained in a population
What is sildenafil citrate structure?
Treats erectile dysfunction
→ structural similarity between sildenafil and the secondary messenger cGMP
→ potent inhibitor of cGMP phosphodiesterase PDE-5
→ also inhibits PDE-6, which regulates blue-green colour discrimination in the retina
side-effect of sildenafil citrate can be blue-tinged vision
→ pilots warned not to fly within 6 hours of taking
How do intracellular receptors that are enzymes work?
- an extracellular ligand diffuses across the plasma membrane
- ligand binds and activates its receptor
- the activated receptor converts its substrate into product
- the activity of downstream targets is altered
How does nitric oxide signal?
It activates granulate cyclase (intracellular receptor)
1. the soluble gas nitric oxide (NO* free radical) diffuses across the plasma membrane
3. binds and activates its receptor: granulate cyclase
4. the activates receptor converts GTP into cGMP
5. cGMP is a second messenger that alters the activity of its target proteins
dilation of blood vessels stimulated by endothelium cells
How is angina treated today?
Glycerol trinitrate (nitroglycerine)
→ releases NO* - blood vessels dilate
(previously with nitroglycerine - withdrew)
How do blood vessels dilate?
Blood vessels dilate in response to high blood pressure
→ increased volume = lower bp
relaxation of smooth muscle - dilation of blood vessel
NO* production is stimulated by high blood pressure, what does it do?
- autonomous nerves in blood vessel walls respond to high bp and releases acetylcholine (Ach)
→ Ach binds its receptors on p membrane of endothelial cells - increases cytosolic [Ca++]
→ high [Ca++] activates nitric oxide synthase - catalyses arginine → citrulline + NO
→ NO* is unstable - converted to nitrate within 10s (short life - paracrine signalling to smooth muscle)
→ NO* activates soluble granulate cyclase by binding its haemorrhage group causing conformational change
→ granulate cyclase converts GTP to cGMP
→ cGMP is a secondary messenger
What does cGMP activates by NO* do?
Activates protein kinase G in smooth muscle
→ PKG is a cGMP-dependant protein kinase - phosphorylates myosin light chain
→ muscle cells relax - dilation of blood vessels
→ increases blood vol = lower bp
How is blood pressure homeostasis co-ordinated?
By multiple signalling pathways
Signal 1: high shear → Ach release
Signal 2: Ach binding → [Ca++] increase
Signal 3: NO* → cGMP - myosin light chain phosphorylation
physiological response: blood vessel dilation, bp reduction
Where is nitric oxide used in neurotransmission?
NO* used in many signalling pathways e.g.
→ control of capillary dilation
control of blood vessel dilation
→ control of peristaltic movement though the gut
→ regulation of glomerular capillary pressure
→ regulation of blood flow in the adrenal glands
→ neurones of the corpus cavernous (erectile tissue in penis): regulation of muscle contraction and blood flow
processes are similar but not identical
→ not all responses are via cGMP-dependant protein kinase
What is amyl nitrate inhalation spray?
Commonly prescribed for a ‘weak’ heart
→ rapidly acting vasodilator
→ vaporises to generate NO*
→ NO* dilates vascular smooth muscle
→ results in coronary vasodilation + decreases systemic vascular resistance (lower bp)
also available as ‘Poppers’ or ‘Rush’ → effects are rapid but short lived
What are phosphodiesterases (PDEs)?
A super family of metallophosphydrolases that specifically cleave the 3’,5’-cyclic phosphate moiety of cAMP and/or cGMP to produce corresponding 5’ nucleotide
Phosphodiesterase type 5 (PDE5) specifically cleaves cyclic guanine monophosphate (cGMP)
→ the tissue distribution of PDE5 is relatively restricted - in lungs, vascular and tracheal smooth muscle, and platelets
→ PDE5 is the target for sildenafil citrate
What does sildenafil (viagra) inhibit?
cGMP phosphodiesterase
→ most active on PDE5 - the principle PDE in the corpus cavernosum, the erectile tissue of the penis
→ PDE5 - blood vessels become narrower, keeps muscle contracted
side effects: lowers bp - stimulates headaches, (also PDE3) heart palpitations and increased heart rate (tachycardia), blue-tinged vision
What are oestrogen receptors (ERs American spelling)?
Intracellular receptors that are transcription factors
→ they are cytosolic
What is the structure of oestrogen receptors (ER)?
The ER has an N-terminal transactivation domain, a DNA-binding domain and a hormone-binding domain that can bind oestrogen
→ its stores in he cytosol in complex with a dimeric chaperone protein called Hsp90
→ Hsp90 binds near the ligand-binding site and maintains the ER in a soluble state
→ the Hsp90:ER complex is too large to enter the nucleus (can’t diffuse across membrane - inactive)
What is the oestrogen-bound ER (oestrogen receptor)?
A transcription factor
1. oestrogen diffuses across PM + binds ER
2. ER is released from Hsp90
3. ER-oestrogen complex enters the nucleus + binds oestrogen response elements (EREs) as a dimer
4. oestrogen-responsive genes and transcribed
What is unusual about steroid hormone signalling?
Theres no amplification
→ no secondary messengers or protein cascades
→ one protein is receptor and effector
e.g. ER is receptor for oestrogen → oestrogen-actiavted ER binds DNA and directs transcription of oestrogen response genes (no amplification via protein cascades pr 2nd messengers)
What are the physiological roles of oestrogen receptors?
Reproduction
Cardiovascular system
Immune system
Central nervous sytem
Skeletal system
→ how can it regulate so many responses?
How can oestrogen receptors (ER) regulated so many processes?
There are multiple isoforms of the ER
→ 2 different forms of the ER (α and β) each encoded by separate gene (ESR1 and ESR2)
→ they can form ERα (αα) or ERβ (ββ) homodimers or ERαβ heterodimers
+ splicing variants → at lease 3α and 5 isoforms are known - large combinatorial repertoire
What happens when oestrogen binds GPER?
Multiple pathways are stimulated
1. ligand-dependant activation of ER
2. release of EGF via Ca++ as a second messenger
3. stimulation of the MAPK signalling pathway via interaction with Grb2
What is tamoxifen?
Used for treatment of ER+ breast cancer
→ a non-steroidal ER antagonist larger than estradiol (ER agonists) so the loops cannot take on their active formation
→ oestrogen signalling via ERs is inhibited
→ also causes cells to remain in the G0 and G1 phases of the cell cycle
