L10 Intracellular Signalling

Question 1

What cannot cross the lipid bilayer?

Answer 1

Proteins, peptides and charged molecules

Question 2

What are integral membrane proteins?

Answer 2

Transmit signal into the cell and span the membrane

Ligand may interact directly or indirectly

Question 3

How are signalling molecules controlled?

Answer 3

Post Translational Modification - phosphorylation
Regulating G protein’s GDP or GTP
Provision of activators - calcium ions

Question 4

What is kinase?

Answer 4

An enzyme that phosphorylates proteins

Question 5

What is phosphatase?

Answer 5

An enzyme that dephosphorylates proteins

Question 6

What do protein kinases and phosphates act on?

Answer 6

OH group of serine, threonine and tyrosine (free hydroxyl groups)

Question 7

Serine/Threonine kinases examples

Answer 7

Ca2+/calmodulin-dependent protein kinases  or CaM kinases 
 Protein Kinase A (PKA)
 Protein Kinase B (PKB)
 Protein Kinase C (PKC)
Mitogen-activated protein kinases (MAPK)

Question 8

Tyrosine kinases examples

Answer 8

Non receptor tyrosine kinases e.g. Src family kinases -

Receptor tyrosine kinases (RTKs) –
e.g. Insulin receptor
Epidermal growth factor receptor (EGFR

Question 9

What are GTP binding proteins?

Answer 9

Many GTP binding proteins are individual proteins (small GTPases)

Used by heterotrimeric G proteins

Hydrolyse GTP-GDP by intrinsic GTPase activity (act as molecular switch)

Question 10

What do ion channels do? [4]

Answer 10

Transport Na+, K+, Ca2+, Cl- Ions along an electrochemical gradient

Specificity of the channel is defined by the amino acids lining the channel

Fast regulated opening/closing mechanism (mSec)

Channels are formed of protein subunits

Question 11

What are voltage gated ion channels?

Answer 11

Voltage gated ion channels open in response to a change in voltage

The alpha-subunit of the Na+ channel contains the pore and has 4 homologous domains, each with 6 TM regions

Region 4 of the alpha-subunit contains amino acids with positive R-groups which sense the voltage across the membrane causing movement of region 4 which opens the channel

Question 12

What are ligand gated ion channels?

Answer 12

Transmembrane proteins consisting of a receptor part and a channel which traverses the membrane
Open in response to binding of a ligand
Receptors are often classified based on which agonists they bind

Question 13

What events are regulated by calcium ions? [4]

Answer 13

Secretion of enzymes, hormones and neurotransmitters
Changes in some transcription factor activities
Skeletal muscle contraction
Development of an egg upon fertilisation with sperm

Question 14

What mediates calcium ion effects?

Answer 14

Calmodulin which binds Ca2+ and activates other proteins

CAM kinases calmodulin-dependent protein kinases

Calcineurin Ca2+-dependent protein phosphatase

Question 15

How many transmembrane domains do GPCRs usually have?

Answer 15

7

Question 16

GPCR initiation of signalling

Answer 16

Action of the receptor causes GDP attached to alpha subunit to be replaced by GTP

G-protein splits up in beta-gamma complex an α-subunit

Both complexes then initiate further cell signalling

GTP is then hydrolyzed to GDP and beta-gamma recombines with alpha ready to associate with another GPCR

Question 17

Adenylate cyclase and cAMP

Answer 17

Adrenaline + GPCR
Activated adenylyl cyclase
cAMP activates PKA

Gs

Question 18

Phospholipase C Activation

Answer 18

Phosphorykates inositol phospholipid tp form inosition 1,4,5-triphosphate and diacylglycerol -> PKC

Gq

Question 19

GPCRs and K+

Answer 19

Muscarinic ACh receptor in heart

ACh bind GPCR and activate beta-gamma unit, which binds potassium channel = open

Hyperpolerisation

Question 20

Cholera

Answer 20

Cholera toxin
Inhibits the GTPase activity of the
subunit Gαs (stimulates adenylyl cyclase)
Prolonged signal causes water and Cl- to
move out of the cells lining the intestine
Results in diarrhoea, dehydration and death

Question 21

Whooping Cough

Answer 21

Pertussis toxin
Renders Gαi inactive (inhibits adenylyl cyclase)
Prevents Gαi from associating with GPCRs
Prolonged signal that stimulates coughing

Question 22

Enzyme receptors

Answer 22

The cytoplasmic enzymes that induce signalling are normally protein tyrosine kinases
These receptors usually have a single membrane spanning domain

Question 23

Receptor tyrosine kinases

Answer 23

Ligand binding activates an enzyme activity associated with the cytoplasmic domain

The response usually requires receptor dimerisation

Pairs of tyrosine residues in the intracellular domains are
auto-phosphorylated in response to the signal

Question 24

JAK-STAT

Answer 24

In the JAK-STAT pathways
a ligand binding to its 
receptor activates an 
associated protein tyrosine
kinase, which directly
phosphorylates and thus
activates a STAT
transcription factor