Pharm - Lecutre 3 - signal transduction Flashcards
What does cortisol bind to once inside the target cell?
GR = glucocorticoid receptor
What carrier protein does cortisol bind to?
CBG
How does cortisol get into through the plasma membrane?
diffusion
What occurs after GR is bound by cortisol in the cytoplasm?
1) Cortisol causes a conformation change on the GR receptor,
2) This releases inhibitory proteins HSP70. The GR receptor is now free to diffuse into the nucleus
What occurs once once the active GR receptor diffuses into the nucleus?
The active receptor binds to a hormone response element (GRE = glucocorticoid response element)
What are the subunits of the Nicotinic acetylcholine receptor?
a1, a2, B, y, delta
What type of receptor are Nicotinic acetylcholine receptors?
Is this excitatory or inhibitory?
Ion channel-linked receptor
Excitatory
Where is the Nicotinic acetylcholine receptor located? (3)
1) Nueromuscular junction
2) peripheral autonomic nervous system
3) central nervous system
What type of receptor is the GABA receptor?
gated Cl- channel
What type of response is elicited from GABA receptors?
inhibitory - GABA is the main post-synaptic inhibitory transmitter in CNS
What affect does the activated GABA receptor have on the cell electrical state?
It allows Cl- and so hyperpolorizes the cell
How many times does the GCPR pass through the membrane?
7
What are the three subunits of G proteins and what are their function?
Ga: binds GT, binds effectors, acts as GTPase
Gbeta and Ggamma form the beta/gamma dimer and anchors to the membrane
What state of the G protein elicits an active response? What turns off this response?
GTP bound to the Galpha subunit occurs when activated
GTPase activity of the alpha subunit converts GTP to GDP and turns activity off
What are three types of Galpha proteins and what do they do?
1) Gas (stimulates adenylyl cyclase)
2) Gai (inhibits adenylyl cylcase)
3) Gaq (stimulates phospholipase C)
What are two possible structures of Tyrosine Kinase-Linked receptors?
1) single protein with one transmembrane domain
2) tetramer composed of 2 EC subunits and 2 transmembrane subunits
Where does the ligand bind a Tyrosine Kinase-Linked receptor?
EC domain
What occurs upon ligand binding to the Tyrosine Kinase-Linked receptor? (3)
1) dimerization
2) activation of cross-phosphorylation (the 2 cytoplasmic subunits phosphorylate each other
3) binding of intracellular signaling molecules
What are examples of serine/threoinine specific protein kinases? (4)
PKA
PKC
Ca2+/calmodulin dependent protein kinase
MAP kinases
What are examples of tyrosine specific protein kinases?
EGF receptor (and other tyrosine kinase-linked receptors)
Src,
Abl
What is an example of dual specificity protein kinase? What do these proteins phosphorylate?
Phosphorylate both threonine and tyrosine
MAP kinase kinase (MKK1)
What are examples of second messengers?
cAMP
DAG
IP3
Calcium
How is cAMP generated?
generated by adenyl cyclase when it is activated by Gas
How is DAG and IP3 generated?
Generated when PLC cleaves PIP2 into DAG + IP3
What is the signaling pathway for cAMP regulated by GPCRs?
1) ligand binds receptor
2) Gas binds GTP and dissociates from By subunits
3) Gas-GTP activates adenylyl cyclase
4) adenylyl cyclase converts ATP to cAMP
5) cAMP activates PKA
6) PKA phosphorylates and activates or inactivates an enzyme
7) enzyme makes (or stops making) products
What is the structure of PKA?
Tetramer - 2 catalytic and 2 regulatory subunits
How does PKA become active and what happens? (3 steps)
1) The R subunits each bind a cAMP molecule
2) The R and C subunits dissociate
3) C subunits become autophosphorylated and activated
What happens: C subunit phosphorylates proteins on Threonine and serine residues activating or inactivating them
What is the difference between Gas activity and Gai? (2)
1) Gas stimulates adenylyl cyclase, Gai inhibits it
2) they are activated by different GPCR’s
What inositol-lipid pathway? (5 steps)
1) ligand binds to and activates GPCR
2) Gaq binds GTP and dissociates from By subunits
3) Gaq-GTP activates phospholipase C (PLC)
4) PLC cleaves PIP2 to make DAG and IP3
5a) IP3 diffuses into cytoplasm and releases calcium from ER
5b) DAG remains attached to membrane and together with calcium, activates PKC
How are Monomeric G proteins activated?
Monomeric G proteins are recruited to receptor linked-tyrosine kinase and turned on by GEF
How are monomeric G proteins turned off?
Monomeric G proteins are recruited to receptor linked-tyrosine kinase and turned off by GAP
What type of molecule is RAS?
monomeric G protein
What type of molecule is SOS?
GEF
How is RAS-GTP activated?
1) a ligand binds the tyrosine kinase receptor causing autophosphorylation
2) the receptor binds an adapter protein called GRB2
3) GRB2 activates Sos
4) Sos (a GEF) activates Ras (a monomeric G protein)
5) The GEF (sos in this case) causes Ras-GDP to release the GDP and bind GTP thus activating it
How is Ras-GTP inactivated?
When Ras-GTP interacts with GAP, the GAP induces Ras-GTP’s inherent GTPase activity and GTP is hydrolyzed to GDP
What activates Raf to begin the MAP Kinase signaling cascade?
Ras-GTP activates Raf
What type of molecule is Raf?
a MAP kinase kinase kinase
What are the steps for the MAP kinase signaling cascade starting from an activated Ras-GTP?
1) Ras-GTP activates Raf
2) Raf phosphorylates MKK1
3) MKK1 phosphorylates ERK
4) ERK phosphorylates many different transcription factors
What type of molecule is MKK1?
MAP Kinase Kinase
What type of molecule is ERK?
MAP kinase
What type of receptor is the EGF receptor?
Tyrosine kinase receptor
What would mutations in EGF receptor cause?
Abnormally high activity of the MAP kinase signaling cascade - promotes tumor growth
What would mutations in Ras cause?
Abnormally high activity of the MAP kinase signaling cascade - promotes tumor growth
What is the mechanism of gefitinib?
Inhibits signaling of the EGF receptor
What is the mechanism of erlotinib?
Inhibits signaling of the EGF receptor
What two drugs inhibit signaling of the EGF receptor?
1) gefitinib
2) erlotinib
What determines the efficacy of gefitinib and erlotinib regarding tumor supression?
Efficacy is high if the tumor expresses a MUTATED EGF receptor. They don’t work as well (-10%) if the EGF receptor mutations are absent
What are mechanisms of adaption? (2)
Receptor mediated endocytosis
Receptor desensitization
What is adaption?
Modulation of signals in response to intensity and frequency of stimulation
How is receptor mediated endocytosis carried about?
1) upon binding the ligand, receptors migrate to coated pits
2) the receptor is internalized
3) some receptors are recycled, some are degraded by lysosomes
What is the role of clathrin?
clathrin is the structural protein that forms the electron dense cage that forms the vesicle that receptors are endocytosed in
What is CURL?
the tubular-reticular structure that forms the compartment for receptor-ligand uncoupling
What is the p.H. in CURL and why?
4.5 - 5 to favor dissociation of ligand and receptor
What is retroendocytosis?
The recycling of endocytosed receptors
What is homologous desensitization? What is the process? (2 steps)
The desensitization of the stimulated receptor
Process:
1) GPCR is phosphorylated by betaARK
2) beta arrestin binds to phosphorylated GPCR and inactivates it
What is Heterologous desensitization? Mechanism?
Both the stimulated and unstimulated receptors are blocked
Mechanism:
1) phosphorylation of receptors
2) competition for shared second messengers
