Week 6 - Receptors in cell signalling, principles of receptor-mediated endocytosis

Question 1

How can chemical signals be classified?

Answer 1

According to their function
- Hormones (signalling between cells in different tissues via the circulation)
- Neurotransmitters (signalling at specialised cell junctions in the nervous system, synapses)
- Local chemical mediators (signalling between adjacent cells in the same tissue)
A single molecule may fall into more than 1 of these categories depending on where it is synthesised and released, and it’s site of action

Question 2

What is a ligand?

Answer 2

Any small molecule that binds specifically to a receptor site

Question 3

What is an antagonist?

Answer 3

A ligand that does not cause activation upon binding

- It opposes the action of the ligand

Question 4

What are partial agonists?

Answer 4

Agonists which stimulate a receptor but are unable to elicit the maximum cell response possible

Question 5

What is a receptor?

Answer 5

A molecule that recognises specifically a second molecule (ligand) or family of molecules, and which in response to ligand binding brings about regulation of a cellular process
- In the unbound state, it is functionally silent

Question 6

What are some examples of the roles of receptors in cellular physiology?

Answer 6

• Signalling by hormones/local chemical mediators
• Neurotransmission
• Cellular delivery
• Control of gene expression
• Cell adhesion
• Modulation of the immune response
• Sorting of intracellular proteins
• Release of intracellular calcium stores

Question 7

What is the difference between the binding affinity of enzymes and that of ligands?

Answer 7

Affinity of ligand binding at receptor sites is generally much higher than binding of substrates and allosteric regulators to enzyme

Question 8

How can receptors be classified?

Answer 8

According to the specific physiological signalling molecule (agonist) that they recognise
- Sub-classification: affinity of a series of antagonists

Question 9

What is an acceptor?

Answer 9

A receptor whose basic function can be carried out without the interaction of a ligand
- Ligand binding alone produces no response

Question 10

How do cells respond to a chemical messenger?

Answer 10

It must produce specific receptor proteins

• These recognise and produce a response to the signalling molecule
• Interaction of the signalling molecule with its specific receptor must then result in the activation of a cellular process

Question 11

What is signal transduction?

Answer 11

Most signalling molecules are unable to cross the plasma membrane, so there are some common mechanisms to transduce an extracellular hydrophilic signal into and intracellular event:

• Membrane-bound receptors with integral ion channels
• Membrane-bound receptors with integral enzyme activity
• Membrane-bound receptors which couple to effectors through transducing proteins
• Intracellular receptors for hydrophobic ligands

Question 12

What are some similarities between receptor binding sites and the active sites of enzymes?

Answer 12

• Binding at both receptor sites and enzyme sites is specific
• The specificity of binding is governed by the shape of the binding cleft in the receptor/enzyme site
• The specificity of binding confers specificity to the regulation of processes in which receptors are involved or the specificity for substrate of an enzyme
• Binding is most often reversible
• Binding induces a conformational change and a change in the activity of the molecule
• No chemical modification of the ligand

Question 13

What is a difference between receptor binding sites and the active sites of enzymes?

Answer 13

The affinity of ligand binding at receptor sites is generally much higher

Question 14

What are the 4 major classes of receptors involved in cellular signalling?

Answer 14

• Membrane-bound receptors with integral ion channels
• Membrane-bound receptors with integral enzyme activity
• Membrane-bound receptors with no integral enzyme or channel activity
• Intracellular receptors

Question 15

How do membrane-bound receptors with integral ion channels work?

Answer 15

Agonist binding to ligand-gated ion channels results in a change in conformation and opening of a gated channel

• This permits the flow of ions down an electrochemical gradient
• This transduces the signal into an electrical event at the plasma membrane

Question 16

What is the subunit structure of the ‘classical’ ligand-gated ion channel family?

Answer 16

They have similar pentameric subunit structures

- Subunits have 4 transmembrane domains, 1 of which forms the lining to the channel pore

Question 17

What are some examples of ligand-gated ion channels?

Answer 17

• Nicotinic acetylcholine receptors
• Gamma amino-butyric acid receptors
• Glycine receptors

Question 18

How do membrane-bound receptors with integral enzyme activity work?

Answer 18

Agonist binding to the extracellular domain of these receptors causes a conformational change
- This activates an intrinsic enzyme activity contained within the protein structure of the receptor

Question 19

Give an example of a membrane-bound receptor with integral enzyme activity and describe how it works

Answer 19

Tyrosine kinase-linked receptors

• Binding of hormone to extracellular binding sites activates protein kinase activity in the cytoplasmic domain of the receptor protein
• This autophosphorylates tyrosine residues on the cytoplasmic domain of the receptor
• These are recognised by transducing proteins or directly by enzymes containing phosphotyrosine recognition sites
• On association with receptor or transducing protein, effector enzymes become activated allosterically
• This transduces the message into an intracellular chemical event

Question 20

How do membrane-bound receptors with no integral enzyme or channel activity work?

Answer 20

• 7 transmembrane domain receptors couple to effectors molecules via a transducing molecule (a G-protein)
• This family of receptors are also know as the G-protein coupled receptor family)
• Effectors may be enzymes or ion channels
• A wide variety of extracellular signalling molecules utilise specific 7 transmembrane domain receptors
• Receptor binding results in a conformational change which activates GDP/GTP exchange in GTP-binding regulatory proteins
• This transduces the message onto an enzyme or channel in the membrane

Question 21

Give some examples of membrane-bound receptors with no integral enzyme or channel activity

Answer 21

• Muscarine ACh receptor
• Adrenoceptors
• Dopamine receptors

Question 22

How do intracellular receptors work?

Answer 22

• Hydrophobic ligands penetrate the plasma membrane
• They bind to monomeric receptors and are stabilised by association with heat shock or chaperone proteins
• The activated receptor dissociates from the chaperone protein and translocates to the nucleus
• It then binds to control regions in DNA defined by specific sequences, thereby regulating gene expression
• The effects of intracellular activation are slow in onset as transcription and translation are required

Question 23

How does phagocytosis occur?

Answer 23

In response to the binding of a particle to receptors in plasma membrane

• The cell extends pseudopods
• These permit further receptor interactions and membrane evagination and particle internalisations
• Internalised phagosomes fuse with lysosomes to form phagolysosomes in which the particulate material is degraded

Question 24

What is pinocytosis?

Answer 24

Invagination of the plasma membrane to form a vesicle

- Permits uptake of extracellular salutes and retrieval of plasma membrane

Question 25

What are the 2 forms of pinocytosis?

Answer 25

• Fluid-phase

- Receptor-mediated endocytosis

Question 26

What is receptor-mediated endocytosis?

Answer 26

Specific binding of molecules to cell surface receptors

Question 27

What is an example of receptor-mediated endocytosis and explain how it occurs?

Answer 27

Uptake of cholesterol

• Low density lipoproteins originate in the liver
• Animal cells that require cholesterol synthesise cell surface receptors that specifically recognise apoprotein B (found on LDL)
• LDL receptors are localised in clusters over coated pits
• Coated pits invaginate and pinch off from the plasma membrane to form coated vesicles
• Coated vesicles are quickly uncoated (driven by ATP)
• The uncoated vesicles then fuse with larger smooth muscle vesicles called endosomes
• The pH of the endosomes is maintained between approximately 5.5-6.0 by an ATP-dependent proton pump
• At this pH, the LDL receptor has low affinity for the LDL particle and the 2 dissociate
• The transmembranous receptors are sequestered to a domain within the endosome membrane which buds off as a vesicle (this recycles the LDL-receptor to the plasma membrane)
• The endosomes containing the LDL fuse with lysosomes such that the cholesterol can be hydrolysed from the esters and released into the cell
• Hence the receptor is recycled and the ligand is degraded

Question 28

How is the clathrin coat attached to the plasma membrane?

Answer 28

By a number of integral membrane adaptor proteins

- These form associations with both the clathrin and receptors

Question 29

What is another name for endosomes?

Answer 29

CURL

- Compartment for the uncoupling of receptor and ligand

Question 30

How does coated pit formation occur?

Answer 30

Spontaneously and non-specifically

Question 31

What mutations are there that can affect the LDL receptor?

Answer 31

• Receptor deficiency: mutations that prevent expression of LDL receptor
• Non-functional receptor: no binding of LDL, normal coated pits and internalisation
• Normal receptor binding but no internalisation: due to deletion in the C-terminal of the receptor that makes interaction with the coated pits. LDL receptors are found distributed over the whole call surface in these patients

Question 32

How are ferric (Fe3+) ions taken up?

Answer 32

• 2 Fe3+ ions bind to apotransferrin forming transferrin in the circulation
• Transferrin binds to the transferrin receptor at neutral pH and is internalised in the same way as LDL
• On reaching the acidic endosome, the Fe3+ ions are released from the transferrin
• Apotransferrin remains associated with the transferrin receptor
• The complex is sorted in the CURL for recycling back to the plasma membrane, where at pH 7.4, the apotransferrin dissociates from the transferrin receptor again
• The ligand and receptor are hence recycled

Question 33

How does the uptake of occupied insulin receptors occur?

Answer 33

• The insulin receptor only congregates over coated pits when the agonist is bound
• Insulin binding probably induces a conformational change in the insulin receptor, allowing it to be recognised by the coated pit
• In the endosome insulin remains bound to the receptor
• The complex is targeted to the lysosomes for degradation
• This mechanism allows down-regulation of insulin receptors on the membrane surface, desensitising the cell to a continued presence of high-circulating insulin concentrations
• The ligand has been degraded, as has the receptor

Question 34

What is transcytosis?

Answer 34

When ligands that remain bound to their receptors are transported across the cell

Question 35

How can membrane-enveloped viruses and toxins enter cells using endocytic pathways?

Answer 35

They exploit endocytic pathways to enter cells after adventitious binding to receptors in the plasma membrane

• Once in the endosome, where the acid pH is favourable, the viral membrane is able to fuse with the endosomal membrane
• This releases viral RNA into the cell
• It can then be translated and replicated to form new viral particles