GPCRs 3

Question 1

to use Ca2+ as an intracellular signal, the resting cytosolic Ca2+ levels are kept…

Answer 1

low

Question 2

what is the concentration of free Ca2+ in the cytosol?

Answer 2

less than or equal to 10^-7 M

extremely low

Question 3

what is the concentration in the extracellular fluid + ER?

Answer 3

10^-3 M

high

Question 4

what does the difference In concentrations allow?

Answer 4

a large gradient which drives Ca2+ into cytosol across plasma + ER membrane

Question 5

describe the events following a signal?

Answer 5

• calcium channels open

- calcium ions enter cytosol triggering calcium responsive proteins in cell

Question 6

for the signalling mechanism to work, what must the resting calcium ion concentration in the cytosol be kept?

Answer 6

low

Question 7

how can you keep the calcium ion conc low?

Answer 7

• keeping calcium ions out of cell

- keeping intracellular ca2+ low

Question 8

keeping calcium ions out of cell

Answer 8

• cells have ca2+-ATPase in plasma membrane

- uses ATP to pump ca2+ out of cell

Question 9

keeping intracellular ca2+ low

Answer 9

• ca2+ pump in ER helps keep cytosolic [ca2+] low
• if cell damaged, [ca2+] rises to high level (>10^-5 M) = dangerous
• low affinity, high-capacity ca2+ pump in inner mitochondrial membrane takes up ca2+

Question 10

give an example of a specific ca2+ binding protein?

Answer 10

calmodulin

Question 11

how many mechanisms are there altogether and how many are inside?

Answer 11

5 altogether

3 inside

Question 12

ca2+ as an intracellular messenger

Answer 12

intracellular injection of ca2+ caused skeletal muscle to contract

Question 13

ca2+ entry into cells

Answer 13

• depolarisation of PM -> influx of ca2+ via voltage-gated ca2+ channels
• binding of mol to cell-surface receptors -> ca2+ release from ER
• this = mediated via inositol trisphosphate (IP3)

Question 14

what does the smooth muscle not need?

Answer 14

extracellular calcium to function

Question 15

when was the role of the membrane first described?

Answer 15

1953

Question 16

what are the most important inositol phospholipids?

Answer 16

• PIP (phosphatidylinositol-phosphate)

- PIP2 - PI- bisphosphate

Question 17

hydrolysis of PIP2

Answer 17

PIP2 breakdown begins with binding of signalling mol to G protein linked receptor

• activated receptor stimulates trimeric G protein (Gq)
• activates enzyme phospholipase c-beta

Question 18

what is the best example of hydrolysis of PIP2?

Answer 18

m3 rat ileum receptor

Question 19

more than …. different cell-surface receptors use the hydrolysis of PIP2 pathway?

Answer 19

25

Question 20

when PLC-beta cleaves PIP2

what 2 products does it generate?

Answer 20

• IP3

- diacylglycerol (DAG)

Question 21

where does PLC cleave the bond?

Answer 21

cleaves between O of phosphate and CH2 of other mol

Question 22

IP3 diffuses through…

Answer 22

cytosol and releases Ca2+ from ER

Question 23

DAG remains in the membrane and …

Answer 23

activates protein kinase C

Question 24

what are the 3 types of phospholipase C?

Answer 24

beta, gamma and delta

beta class activated by G protein linked receptors

Question 25

what is the journey of IP3?

Answer 25

- leaves plasma membrane - releases ca2+ from ER by binding to IP3-gated ca2+ release channels in ER memb - channels regulated by POSITIVE feedback - released ca2+ binds back to channel to inc ca2+ release (CICR) - sensitive to ryanodine

Question 26

calcium + smooth muscle contraction

Answer 26

- smooth muscle cells store ca2+ in ER and SR - released when IP3 receptor activated - IP3 generated by Gq linked receptor activation

Question 27

what is the contraction of smooth muscle activated by?

Answer 27

MLC phosphorylation catalysed by myosin light chain kinases (MLCK) activated when bound to ca2+ - calmodulin

Question 28

what do myosin phosphatase reverse?

Answer 28

phosphorylation + cause relaxation

Question 29

no regulatory proteins are involved in ...

Answer 29

smooth muscle contraction

Question 30

function of ca2+ calmodulin complex

Answer 30

- calmodulin binds to 4 ca2+ ions - calmodulin changes conformation, resulting in active complex - 2 globular hands of complex wrap around binding site on target protein

Question 31

role of ca2+ in cells

Answer 31

- activate membrane ion channels - regulate enzyme activity - modulate protein function - regulate gene expression - kill cells! (necrotic + apoptotic death) - life and death messenger

Question 32

what is calcium the only link between?

Answer 32

electrical activity of neurone and end response of cell

Question 33

what triggers muscle contraction?

Answer 33

ca2+ liberation from SR

Question 34

what is different about the ca2+ unlike the other ions?

Answer 34

it is chemical signal carried that is important not the charge

Question 35

what are the sources of ca2+ to cells?

Answer 35

- extracellular fluid (2mM) - plasma membrane channels - ligand gated channels (glutamate NMDA) - store operated channels (open when less ER calcium stores)

Question 36

what are the 2 mechanisms that operate to terminate the ca2+ response?

Answer 36

- IP3 dephosphorylated by enzymes | - ca2+ that enters cytosol rapidly pumped out

Question 37

what happens to IP3 if it isn't dephosphorylated?

Answer 37

phosphorylated to make IP4

Question 38

the enzyme that catalyses the production of IP4 is activated by...

Answer 38

increase in cytosolic (ca2+) induced by IP3

Question 39

why is the resynthesis of phospholipids important in regulation?

Answer 39

cell would run out of substrate

Question 40

at least how many reactions are involved in the metabolism of inositol lipids?

Answer 40

13

Question 41

the regulation reactions can be divided into 2 cycles what are they?

Answer 41

- inositol lipid cycle - occurs in membrane | - inositol phosphate cycle - occurs in cytosol

Question 42

what is PKC?

Answer 42

ca2+ dependent enzyme

Question 43

what does DAG increase the affinity of?

Answer 43

PKC for ca2+ -> activating enzyme

Question 44

what does activated PKC do?

Answer 44

phosphorylated specific target proteins that vary depending on cell type

Question 45

what are the several physiological functions of PKC?

Answer 45

- secretion - modulation of ion conductance - control of growth + differentiation

Question 46

where are the highest concentrations of PKC found?

Answer 46

in brain phosphorylates ion channels there

Question 47

G protein linked receptors can desensitise in 3 ways what are they?

Answer 47

- become altered so longer interact with G proteins (inactivation) - temp moved to interior of cell -> no longer have access to ligand (sequestration) - destroyed in lysosomes after internalisation (receptor down regulation)

Question 48

what does desensitisation depend on?

Answer 48

phosphorylation of receptor by PKA, PKC or G protein linked receptor kinases

Question 49

what do GRKs phosphorylate?

Answer 49

serine + threonines

Question 50

once receptor has been phosphorylated, what does it bind to?

Answer 50

member of arrestin family of proteins

Question 51

what can receptor desensitisation be classified into?

Answer 51

- homologous | - heterologous

Question 52

what are the enzymes involved in receptor phosphorylation?

Answer 52

- beta adrenergic receptor kinase (BARK) - cAMP- dependent protein kinase - protein kinase C (PKC)

Question 53

homologous desensitisation

Answer 53

- BARK most imp | - capable of phosphorylating active agonist

Question 54

desensitisation of GPCR?

Answer 54

1. inhibitory structural alteration of receptor 2. receptor internalisation 3. receptor degration arrestin takes to Cathrin coated pits + degradation

Question 55

give examples of receptor up + down regulation?

Answer 55

- cholesterol UP by statins - opiate UP by antagonist naloxone - beta adrenoreceptors UP by antagonist