Signal Transduction

Question 1

What are the two main types of kinases

Answer 1

• tyrosine kinase

- serine/threonine kinase

Question 2

what are the two types of GTP-binding proteins

Answer 2

• trimeric G proteins

- monomeric GTPases

Question 3

how many kinases and phosphates does the human genome code for

Answer 3

520 kinases and 150 phosphates

Question 4

What are G-protein coupled receptors

Answer 4

Ligand bindning activates associated G protein which in turn activates or inhibits a downstream protein/enzyme. often this is the enzyme that generates a specific intracellular SECOND MESSENGER

Question 5

Examples of G protein coupled receptors

Answer 5

epinephrine, glucagon, serotonin

Question 6

tell me about the structure of a G protein coupled receptor

Answer 6

they have 7 membrane spanning regions (helices) composed of hydrophobic amino acids

• amino termini on the extracellular surface and also interacts with heterotrimeric G protein complex
• carboxy termini on the cytoplasmic face of the plasma membrane

Question 7

what does trimeric mean

Answer 7

composed of three different subunits (a, b and g)

Question 8

many cell-surface receptors are coupled to

Answer 8

trimeric signal-transducing G proteins that bind either GTP or GDP.

Question 9

what is the mechanism of action in regards to coupling of G protein receptors and trimeric signal transfusing G proteins

Answer 9

• the binding of a ligand changes the confirmation of the receptor
• causing it to bind Galpha protein in such a way that GDP is displaced and GTP is bound
• this triggers Gbetagamma dissociation activating downstream pathways
• activation is short lived, as GTP bound to Galpha hydrolyses to GDP in seconds leading to the re-association of Galpha and Gbetagamma and inactivation of adenylate cyclase

Question 10

What does the system need to stimulate these types of reaction

Answer 10

1) A receptor
2) A transducer(G-protein)and
3) An amplifier (adenylate
cyclase) that generates large amounts of a second messenger.

Question 11

what are phospholipase C isoforms

Answer 11

Proteins which possess distinct domain structures but catalyse the same reaction (liberation of IP3 and DAG from PIP2)

Question 12

examples of common domains

Answer 12

(catalytic, membrane localisation)

Question 13

examples of unique domains

Answer 13

regulatory

Question 14

what different kinds of pathway can activate these domains

Answer 14

GPCR activation (Gαq) 
Receptor tyrosine kinases 
Kinases (possibly ↑Ca2+) 
Small GTPases Fertilisation
↑Ca2+, Gβγ, other mechanisms

Question 15

what is resting calcium concentration

Answer 15

~100 nM

Question 16

what is activated calcium concentration

Answer 16

0.5-1 μM (500-1000 nM)

Question 17

how does calcium help fertilisation of an egg by a sperm

Answer 17

• initial spark by PLC-ζ (zeta) triggers opening of surface calcium channels
• Calcium wave triggers start of embryonic development and prevents other sperm from entering the cell

Question 18

What are Protein Kinase C (PKC)

Answer 18

Most are present as catalytically inactive, soluble proteins in the cytoplasm

Question 19

what is the action of PKC

Answer 19

• Rise in cytosolic calcium levels causes PKC to bind to the cytosolic leaflet of the plasma membrane, where it can be activated by the membrane-associated DAG and/or Ca2+
• PKC then phosphorylates a wide variety of substrate proteins on serine and threonine residues.

Question 20

PKC an gene expression

Answer 20

can function indirectly

Question 21

Binding of adrenalin to the b2 adrenergic receptor

Answer 21

Mediates the body’s response to stress/fear (fight or
flight!!)
 release of glucose and fatty acids from liver/fat cells
 increased contraction of cardiac muscle
Binding of adrenalin to b2 adrenergic receptor increases the intracellular concentration of cAMP (cyclic AMP) as receptor couples to Gas
cAMP is synthesized within cells from ATP by the enzyme adenylate cyclase
cAMP is degraded by the enzyme cAMP phosphodiesterase

Question 22

How does increased cAMP activate the cAMP-dependent protein kinase, PKA?

Answer 22

Signal Transduction results in an AMPLIFICATION of a signal:

A small change at the top can have a large change at the bottom.

Question 23

Role of PKA in the nucleus

Answer 23

• PKA allows the phosphorylates CREB, CREB then binds to CRES which is the CAMP response element region which switches on gene transcription

Question 24

Cholera Toxin

Answer 24

Oligomeric complex which after cleavage becomes active and enters intestinal epithelial cells to stimulate Gαs
The overstimulation of cAMP production results in a release of water and ions including Na+, K+, Cl- and HCO3- into the lumen of the small intestine
This leads to rapid fluid loss and dehydration
Pertussis toxin acts in the reverse manner inhibiting Gαi to increase cAMP production in lung epithelia

Question 25

Receptor Tyrosine Kinases (RTKs)

Answer 25

enzyme linked receptors e.g. insulin-like growth factors activate RTKs to control cell proliferation

Question 26

Receptor Tyrosine Kinases activate Ras

Answer 26

Small GTPases
Ras superfamily contains over 100 members: Rho, Rap, Rab, Arf
Regulate cellular processes: proliferation, cytoskeletal dynamics, membrane trafficking/vesicular transport

Question 27

How do GTPases relate to disease

Answer 27

Several small GTPases of the Rac/Rho subfamily are direct targets for clostridial cytotoxins.
Further, Ras proteins are mutated to a constitutively- active (GTP-bound) form in approximately 20% of human cancers

Question 28

What pathway does Ras activate

Answer 28

MAPK (mitogen-activated protein kinase) pathway

Question 29

EGFR Receptor Pathway and Cancer

Answer 29

EGFR (epidermal growth factor receptor) is a RTK activated by TGFα (Transforming growth factor alpha)
The receptor activates Ras via Grb2/SOS proteins
Ras activates Raf (kinase), which in turn stimulates gene transcription via other kinases (MEK and ERK).
Mutations in EGFR, Ras and Raf are associated with tumorigenesis
These cause overexpression and/or hyperactivation of the respective proteins