GPCR and G-protein signaling

Question 1

WHAT ARE G-PROTEIN COUPLED RECEPTORS
(GPCR)

Answer 1

Cell surface receptors that bind G-proteins which use GTP to relay signals throughout the cell

Question 2

WHAT ARE G-PROTEINS

Answer 2

• Proteins composed of Alpha, Beta and Gamma.
• used to relay signals into the cell interior from GPCRs

Question 3

WHAT IS THE STRUCTURE OF G-PROTEINS

Answer 3

A single peptide chain in the form of a cylinder with a deep ligand binding site at the center

Question 4

HOW ARE G-PROTEINS AFFECTED AFTER GPCR ACTIVATION

Answer 4

• when GPCR is unstimulated GDP binds to alpha subunit and it is inactive
• when GPCR is activated, GDP is released and GTP now binds
• now G-alpha subunit is activated and it triggers a conformational change which releases the G-alpha subunit from the G-beta/gamma pair
• then alpha associates with and effector
• beta/gamma subunits bind to distinct effectors and effectors produce an effect

Question 5

WHAT IS cAMP

Answer 5

It is a second messenger (intercellular) that activate GPSRs

Question 6

WHAT IS ADENYLYL CYCLASE

Answer 6

• It is an enzyme that synthesizes cyclic AMP from ATP
• when it is activated cAMP production is enhanced
• when it is inhibited cAMP production is reduced

Question 7

WHICH ENZYME DEGRADES CYCLIC AMP

Answer 7

Phosphodiesterase

Question 8

WHAT IS PKA

Answer 8

It is a kinase that phosphorylates Serine and Theronine

Question 9

WHAT IS CHOLERA TOXIN AND HOW DOES IT STIMULATE G-PROTEIN AND WHAT IS THE RESULT

Answer 9

• it is an enzyme that catalyzes the transfer of ADP ribose from intercellular NAD+ to alpha subunit of the Gs (simulatory G-proteins) so that the alpha subunit can no longer hydrolize its GTP
• as a result the drastic increase in cAMP by the toxin allows a large efflux of Cl- and H2O into the gut and causes diarrhea accosiated with the cholera toxin

Question 10

HOW cAMP AND PKA INTERACT

Answer 10

• when cAMP binds to PKA it alters the PKA conformation causing PKA to release its catalytic subunit
• these subunits can now phosphorylate specific targeted proteins
• when cAMP concentration is high-PKA activated
  -when cAMP concentrations are low-PKA inactive

Question 11

HOW DOES A RISE IN INTRACELLULAR cAMP ALTER GENE EXPRESSION

Answer 11

• a rise in intracellular cAMP concentration allows the activation of PKA
• when PKA is activated it enters nucleus and phoshphorylate an inactive CREB (cyclic AMP response element binding protein) and activates it
• activated CREB binds CBP (CREB binding protein) and this complex can now bind the cis-regulatory sequence (CRE)
• one these bind the gene transcription is activated

Question 12

WHAT IS PHOSPHOLIPASE-C

Answer 12

it is a PM bound enzyme that cleaves PI(4,5)P2 into two products: IP3 (inositol 1,4,5-trisphosphate) and diacylglycerol

Question 13

HOW DOES THE PHOSPHOLIPASE C PATHWAY WORK

Answer 13

• Gq activates the phospholipase C-beta which cleaves PI(4,5)P2 to generate IP3 and diacylglycerol
• when IP3 is released, it quickly diffuse through the cytosol because of its water soluble characteristics and reaches the ER membrane
• once in the ER, it binds IP3-gated Ca2+ release channels and Ca2+is increased in the cytosol
• the released Ca2+ also activates the PKC
• on the other hand, diacylglycerol remains in the PM and activates PKC (Ca2+ dependent protein kinase C)

Question 14

WHAT ARE IP3 AND DIACYLGLYCEROL

Answer 14

They are the secondary messengers produced in the phospholipase-C signaling pathway
- IP3 leaves the membrane and goes to the ER to activate the release of Ca2+ in the ER
- diacylglycerol remains membrane bound and activates PKC.

Question 15

WHY IS CALCIUM SUCH A GOOD SECOND MESSENGER

Answer 15

It self regulates its release by exhibiting negative and positive feedback to activate and inhibit its own release based on the concentration
- Ca2+ is also a great messenger because its concentration in the cytosol is very low whereas its concentration in the extracellular fluid and in the lumen of the ER is high

Question 16

HOW DOES NEGATIVE AND POSITIVE FEEDBACK PRODUCE CALCIUM WAVES AND OSCILLATIONS

Answer 16

• positive feedback involves the activation of IP3 or ryanodine receptors by Ca2+ binding to these receptors
• activation of these receptors triggers the activation of other adjacent lateral receptors
• this results in a huge efflux of Ca2+
• high Ca2+ concentration inhibits the receptors (negative feedback) and Ca2+ is no longer leaving the PM
• the inactivation of the receptors lasts for a certain period of time (refractory period) until another Ca2+ binds and then the process starts all over to produce waves

Question 17

HOW ARE SIGNALS AMPLIFIED WITH SECOND MESSENGERS

Answer 17

• second messengers act as enzymes that catalyze the productions or relay of certain pathways or signals
• the result is more activity within the cell due to the activation done by the second messenger

Question 18

HOW ARE GPCRs DESENSITIZED

Answer 18

• GPCRs are desensitized in three ways:
  -Receptor sequestration; the receptor is internalized or moved to the interior of the cell so that it can no longer access the ligand
  -Receptor down-regulation; destroyed by lysosomes after internalization
  -Receptor inactivation; they are altered so that they can no longer interact with G-proteins

Question 19

WHAT ARE GRKs AND HOW DOES IT CONTRIBUTE TO THE DESENTIZATION

Answer 19

• GRKs are GPCR kinases which include rhodopsin-specific kinase
  -thay phosphorylate serines and threonines on a GPCR
• It desensitizes GPCR by phosphorylating it and allowing this phosphorylation to be recognized by arrestin which prevents the receptor from binding to its G protein.