Amanda

Question 1

P2X

Answer 1

LGIC
Activated by ATP
Non-selective cation-permeable channel
Mediates excitatory postsynaptic response
Third major family of ionotropic receptors

Question 2

Structure of P2X

Answer 2

2 transmembrane domains - the second TMD lines the pore and is responsible for channel gating
N- and C-teminals intracellular
Extracellular domain binds ATP
Exists as a trimer

Question 3

P2X6

Answer 3

Cannot form a functional homomeric receptor

Question 4

P2X7

Answer 4

Cannot form a functional heteromeric receptor

Question 5

Expression of P2X2/P2X3

Answer 5

Commonly found co-expressed in sensory neurones - do they co-assemble into functional P2X2/3 heteromers?

Question 6

P2X2/3 heteromer

Answer 6

1 P2X2, 2 P2X3

Non-desensitising currents (like P2X2) and are sensitive to alpha,beta-meATP (like P2X3)

Question 7

P2X2

Answer 7

Non-desensitising currents in the presence of ATP

Insensitive to alpha,beta-meATP

Question 8

P2X3

Answer 8

Can be activated by alpha,beta-meATP

Desensitise very quickly upon activation

Question 9

What is the evidence that P2X2 and P2X3 form a functional heteromer as a result of their overlapping expression in sensory neurones?

Answer 9

The endogenous receptors have a pharmacological and biophysical phenotype that differs from that of the homomeric receptors of P2X2 or P2X3

Question 10

Why are novel heteromers difficult to identify?

Answer 10

Lack of subunit-specific drugs
Presence of homomeric receptors
Lack of clearly distinct phenotype - indistinguishable from the current produced by a homomer

Question 11

Why are mutant receptor subunits now used to identify heteromeric P2X channels?

Answer 11

Due to the lack of subunit-specific pharmacological tools for P2X receptors

Question 12

What are the advantages of using mutated subunits?

Answer 12

Can knock out the function of one population of homomeric channels
Using P2X2/3 as a model, it is likely that a heteromer containing 2 mutated subunits is also non-functional

Question 13

Disadvantages of heterologous expression systems

Answer 13

Tend to result in over-expression of the protein of interest - could force a heteromeric interaction/subunit composition not seen in native environments
The systems may lack key regulatory/interacting proteins that may affect the function of the protein of interest e.g. HEK cells have few other ion channels (so P2X can be studied in relative isolation)
Expression of the protein of interest may change the properties of the heterologous cell

Question 14

Disadvantages of mutated subunits

Answer 14

Mutations may alter the function of any observed heteromer

Mutations can change receptor pharmacology/trafficking/kinetics/the ability of the subunits to form receptors