11.2 Signaling Transduction II

Question 1

Which receptors are the largest group of receptors and involved in almost eery type of signaling process?

Answer 1

G protein coupled receptors aka GPCRs

Question 2

What is the structure of G protein coupled receptors?

Answer 2

They all have an extracellular ligand binding domain and seven transmembrane binding domains.

All of them work by activating a G protein complex.

they are all membrane associated trimers which are made up of an alpha, beta, and gamma subunit. The alpha subunit also has a binding site which may hold either GTP or GDP

Question 3

G proteins are very diverse and have specific activities, what is similar among all G proteins? What is the structure of G proteins?

Answer 3

they are all membrane associated trimers which are made up of an alpha, beta, and gamma subunit. The alpha subunit also has a binding site which may hold either GTP or GDP

Question 4

What is G Protein associated with in its inactive state?

Answer 4

in the inactivated state GDP is associated with the alpha subunit of a G protein.

Question 5

What does an activated G protein coupled receptor d (GPCR)

Answer 5

it acts as a guanine nucleotide exchange factor (GEF), and stimulates the replacement of GDP with GTP.

Question 6

What does the association of GTP with the alpha subunit of the G protein cause?

Answer 6

it activates the G protein complex, which often dissociates into two pieces; an active alpha subunit and an active beta subunit.

Question 7

What happens when hydrolysis of GTP to GDP occurs on the G protein?

Answer 7

it turns off the alpha subunit and causes it to re-associate with the beta gamma subunit.

Question 8

The alpha subunit of the G protein complex binds to…?

Answer 8

GTP or GDP

Question 9

What are the four G proteins discussed in this lecture?

Answer 9

Gs
Gi
Golf
G12/13

Question 10

What is the function of Gs proteins?

Answer 10

stimulatory. activates calcium channels and adendylyl cyclase

Question 11

What is the function of Gi proteins?

Answer 11

inhibitory. Inhibits K+ channels.

Question 12

What is the function of Golf?

Answer 12

a g protein found in olfactory neurons. It activates adendylyl cyclase

Question 13

What is the function of G12/13 proteins?

Answer 13

activates Rho family to regulate the actin cytoskeleton

Question 14

What is PKA?

Answer 14

protein kinase A, a master signaling kinase.

Question 15

What are the steps of the G protein linked receptor signaling pathway using activation of PKA?

Answer 15

1) G protein coupled receptor activates the trimeric G protein (alpha, beta, gamma) binding site and this leads to the activation of a membrane associated enzyme called adenylyl cyclase.
2) activated adenylyl cyclase catalyzes the formation of cyclic AMP (cAMP) from ATP. cAMP is a second messenger
3) Production of cyclic AMP is a second message in the chain of signals
4) Cyclic AMP binds to a negative regulator subunit attached to protein kinase A (PKA) and causes the release of active PKA.
5) PKA then phosphorylates many downstream effector targets like CREB

Question 16

What is the role of adenylyl cyclase in the typical g protein linked receptor signaling system pathway?

Answer 16

this enzyme catalyzes the formation of cyclic AMP (cAMP) from ATP

Question 17

What enzyme deactivates cyclic AMP (cAMP)?

Answer 17

cyclic AMP phosphodiesterase.

it converts cAMP to adenosine monophosphate of AMP.

Question 18

What would the result be of inhibiting the mechanism of cAMP deactivation?

Answer 18

it would lead to an overall stimulation in the signaling pathways regulated by G proteins.

Question 19

What are the steps in the G protein Signaling via Phospholipase C?

Answer 19

1) GPCR Activates G protein trimer (alpha, beta, gamma) and this activates membrane associated enzyme phospholipase C
2) phospholipase C cleaves phosphoinositides (made of diacylglycerol and IP3)
3) this results in the release of triphosphorylated inositol (IP3) into the cytoplasm and diacylglycerol (lipid part of triphosphorylated inositol) stays in the membrane.
4) IP2 bind to calcium channels on the endoplasmic reticulum and this releases calcium from the ER into the cytoplasm
5) Release of calcium (calcium is a second messsenger) activates many different enzymes including second messenger protein kinase C

**diacylglycerol and calcium also activate protein kinase c, which then phosphorylates a variety of downstream targets.

Question 20

Which pathways involves the release of calcium from internal cell stores as a second messenger? The release of calcium leads to what?

Answer 20

-G protein signaling via phospholipase C

release of calcium leads to activation of master kinase called protein kinase C

Question 21

What are the mechanisms involved for reducing the amount of free calcium in the cytoplasm?

Answer 21

1) Ca2+ pump
2) Sodium/Calcium antiport
3) ATP pumps in the ER membrane can sequester Ca2+ into the ER
4) Ca2+ can be sequestered into mitochondria using the energy of the pH gradient by the electron transport chain
5) Calcium sequestering proteins are present in cytoplasm and can bind up free calcium

Question 22

What can create oscillations in signaling events?

Answer 22

positive and negative feedback and calcium levels.

Question 23

What is the most common mechanism for converting a calcium increase into the activation of enzymes?

Answer 23

the binding of calcium to calmodulin protein

Binding of calcium to calmodulin causes the protein to fold in the middle and the folded protein can wrap itself around regulatory domains of other proteins

Question 24

What is the structure of calmodulin?

Answer 24

four calcium binding domains.

Two calcium binding sites are present on each side of the protein and the two bindings ends of the molecules are separated by a flexible hinge region.

Question 25

How can frequency of calcium oscillations be converted into a signal?

Answer 25

if only a small number of calcium camoduline
kinase 2 proteins are activated by a single brief calcium spike. Each CAM kinase will eventually become dephosphorylated and turned off, so if the calcium spike are infrequent, the resulting downstream activation of signals will be small and will fade

On the other hand,
if the calcium signals are being generated at high frequency, the next spike in a chain of spikes may occur before the effects of the previous spike have
completely worn off,

Question 26

How can some G proteins regulate ion channel activity without using a signaling pathway?

Answer 26

In the heart, the acetylcholine receptor activates an inhibitory G protein labeled Gi.

The Gi trimer binds to potassium channels in the muscle membrane and opens them.The opening of these channels makes it harder to depolarize the cell and thereby diminishes cardiac contraction.

The activation of Golf, a G protein
involved in regulating the sense of smell by olfactory neurons.

In this instance, the activation of adenylyl cyclase results in the production ofcAMP, which then directly binds to and opens gated sodium channels, which then trigger an action potential in the nerve

Question 27

How are G protein mediated signals down regulated?

Answer 27

Inactivation occurs by arrestin

G protein coupled receptors are phosphorylated by one of the downstream activated kinases such as PKA, PKC, or some other kinase.

This in turn causes the recruitment of arrestin to the G protein coupled receptor, which blocks further interaction of the receptor with the G protein trimer complex

this effectively prevents additional signaling to occur, even in the presence of a bound ligand.

Question 28

What is the role of arrestin protein?

Answer 28

to inactivate the G protein coupled receptor (GPCR) by preventing its interaction with G proteins. GPCR has to be phosphorylated so that arrestin can bind and deactivate

Arrestins can also act as adaptor proteins and recruit phosphorylated receptors to clathrin coated pits from where the receptors are endocytosed and afterwards they can be either degraded in lysosomes or activate new signaling pathways.

Question 29

What is the structure of PKA? Attachment of cAMP causes which subunits to activate?

Answer 29

2 regulatory subunits and 2 catalytic subunits

Catalytic subunits activate and detach from regulatory subunits. The catalytic subunits enter the nucleus and phosphorylate a transcription factor called CREB

Phosphorylated CREB recruits a creb binding protein CREB + CREB binding protein activates transcription factors