Signalling via intracellular receptors Flashcards
Which signals can pass through the membrane to interact with intracellular receptors?
- Small molecules
- Hydrophobic molecules
What do cell-surface receptors do?
Bind hydrophillic signals (can’t cross the membrane) and TRANSDUCE the signals into an intracellular response
How do molecules that bind intracellular receptors get into the cell?
Diffuse straight through the membrane
OR
Use carrier proteins
Which 4 molecules bind to nuclear receptors?
1) Steroid hormones (testosterone, cortisol, estradiol)
2) Thyroid hormones (thyroxine)
3) Retinoids (retanoic acid - Vitamin A)
4) Vitamin D
What gases bing to intracellular receptors?
Nitric oxide (NO) Carbon monoxide (CO)
What are 3 properties of NO?
1) Very small (only 2 atoms)
2) Gaseous
3) Free radical
What is a free radical and what can it do?
Unpaired electron in the outer shell
Very reactive
Can damage other proteins
What are other nitric gases that are quite stable?
Nitrogen Dioxide (NO2) Nitrous Oxide (N2O)
How is NO made?
- Substrate of L-arginine (an amino acid)
- 2 successive oxidations
- Producing L-citrulline, NO and water
What enzyme produces NO from L-arginine?
NOS (Nitric oxide synthase)
What is important about NOS?
Comes in various varieties, which are specific for function
What is cNOS?
Constitutive NOS - when transcribed in the cell they are expressed all of the time
How is the enzymatic activity of cNOS induced?
How long does the activity last for?
By Ca2+/Calmodulin
Activity is short-lived
How much NO does cNOS produce?
Very small amounts (picomoles)
What are the 2 representatives of cNOS?
eNOS (endothelial)
nNOS (neural)
Where is eNOS present?
At the CELL MEMBRANE
Expressed in:
- Endothelium
- Cardiac myocytes
- Osteoblasts/osteoclasts
- Platelets
What is nNOS present?
In the CYTOSOL
Expressed in:
- CNA
- NANC neurons
- ENS
- Retina
What is eNOs important in?
Vasculature
What is iNOS?
Inducible NOS
How is iNOS different to cNOS?
Expression is inducible but the enzymatic activity is constitutive
Where as cNOS - the expression is constitutive but the enzymatic activity is regulated
iNOS also has a long-lasting effect and produces more NO
When is iNOS transcriptionally induced?
In response to pathological stimuli (LPS, IFN-g, IL-1)
Where is iNOS expressed?
- Macrophages and Kupffer cells
- Neutrophills
- Fibroblasts
- Vascular smooth muscle and endothelial cells
What activates eNOS?
What does this cause?
Ach release from autonomic nerve terminals
Causes:
1) eNOs to act on L-arginine to make L-citrulline and NO
2) NO binds to guanylyl cyclase and converts GTP to cGMP
3) cGMP then binds to cGMP-dependant protein kinase (PKG) to acitvate it
4) PKG acts on myosin light chain phosphatase
4) MLCP removes phosphate from myosin light chain to cause RELAXATION OF THE MUSCLE
What does NO released from eNOS inhibit?
Platelet aggregation and vascular smooth muscle proliferation
In the CNS, what is nNOS tethered close to in the cytosol?
Why?
NMDA-type glutamate receptor on the postsynaptic membrane
So that nNOS can respond to Ca2+ increases near the open channels
What is the process of nNOS activation and what does this cause?
1) Glutamate produced by presynapse
2) Glutamate bind NDMA glutamate receptor - releases Ca2+
3) Ca2+ acitvates nNOs - synthesises NO
4) NO diffuses into the presynaptic terminal to activate glutamate production
5) Forms a positive feedback loop
What does the positive feedback loop of NO in the brain cause?
Long-term potentiation and memory
synaptic plasticity
What does NO do in the immune system?
- Kills bacteria an parasites
- Programmed cell death
What does iNOS do?
Triggers death at very high levels of NO
Cytostatic and cytotoxic agent
What causes rheumatoid arthritis, Crohn’s disease and asthma?
- Inappropriately elevated levels of NO - causing inflammatory responses
- Necrosis
What can nitroglycerine be used as? (2 things)
1) Dynamite (when mixed with fine clay)
2) Angina treatment
How does nitroglycerine treat angina?
Administered as a spray
Rapidly breaks down in vivo to generate NO - relaxes blood vessels and lowers blood pressure
What does PDE5 do?
Recycles cGMP into GMP and then GTP, even when there is NO present (causes GTP–>cGMP)
This prevents the build up of cGMP (which normally binds to cGMP-dependant protein kinase to produce smooth muscle contraction)
What is an inhibitor of PDE5?
What is the consequence of this?
Viagra
- cGMP builds up
- Smooth muscles relax
- More blood flow to the blood vessels
What is PDE5?
Phosphodiesterase 5
What makes the signalling molecules that bind to nuclear receptors hydrophobic?
Many carbon rings
What is the structure of a nuclear receptor?
1) Transcriptional activating domain
2) DNA-binding domain
3) Regulatory domain/ligand binding domain
What does the regulatory domain of a nuclear receptor bind?
1) Inhibitory proteins
2) Bind the ligand
What does binding of the regulatory domain to an inhibitory protein do? (nuclear receptor)
Holds the receptor in an inactive state
What does binding of the regulatory domain to the ligand do? (nuclear receptor)
- Removes the inhibitory protein
- Resulting in a conformational change
- Binding coactivators
What drives the different effects between ligands in different tissues, in regards to a nuclear receptor?
How?
Coactivators:
- Nuclear receptors are the same in each tissue
- Coactivators are tissue specific and modify the activit of the ligand
What does the DNA binding domain of the estrogen receptor consist of?
Zinc fingers
What are the biding sites divided into?
What receptors bind here?
Inverted repeat (palindromic) - homodimers
Direct repeat (tandem) - heterodimers
How do glucocorticoid and estrogen receprtors bind to DNA?
- Bind as symmetric homodimers to an inverted repeat DNA site
- Bind palindromic sequence
- 2 proteins face each other (mirror image of binding sequence)
- Sequence separated by 3 nucleotides
How do receptors for Vitamin D, retanoic acid and thyroid hormone bind to DNA?
- As heterodimers to tandem repeats
What do the nuclear family members that form heterodimer bind to?
What is important about this factor
RXR (retanoid X receptor)
RXR is present and active all of the time, but can only activate transcription when there is a pairing of the second receptor
What is important in the response element of heterodimers?
The spacing between direct repeats
Where are homodimeric receptors found?
Why are they found here?
In the cytoplasm in the absence of the ligand (in their inactive state)
Anchored to the cytoplasm by inhibitor proteins, including Hsp90
What does hormone binding to homodimeric receptors cause?
Release of inhibitor protein - allowing them to enter the nucleus
Where are heterodimeric receptors located?
Exclusively in the nucleus at their binding site
How do heterodimeic receptors act in the absence and presence of the ligand?
How do they do this?
Absense - Repressors by recruiting histone deacetylases
Presence - activators by recruiting histone acetylases
How do the nuclear receptor family function?
How is this different to insulin receptors?
Entirely by transcriptional regulation
Insulin receptors also have a faster response - translocation
What is the response of nuclear receptors broken down into?
Describe each stage
1) Early primary response
- Receptors activate transcription of primary response genes
2) Delayed secondary response
- Some primary response genes encode transcription factors and activates secondary response genes to create secondary response proteins
What are primary response genes?
Genes that are directly activated by the nuclear receptor
When does the early response occur?
1-6 hours after activation
When does the delayed response occur?
6-48 hours after activation
