Principles of Signal Transduction/ G-Protein Coupled Receptors/ Receptor protein-tyrosine kinase- Lecture 5-8 Flashcards
What are the ligands used in G-protein coupled receptor pathways?
neurotransmitters (epinephrine, serotonin, dopamine); histidine; sensory stimuli (light/ odorants), many prescription drugs
What are the receptors used in G-protein coupled receptor pathways?
proteins with 7 transmembrane domains that interact with ligands (binding extracellularly) to initiate conformational change in the receptor and allow for intracellular domain of the receptor to interact with heterotrimeric G proteins
What are the subunits of the heterotrimeric G proteins in the G-protein coupled receptor pathway?
alpha, beta, and gamma (beta and gamma are normally found together but alpha is bound to GTP when alone and GDP when in complex with the other two)
What is the activation pathway of G-protein coupled receptors?
upon ligand binding, receptor adopts a conformation allowing for interaction with the alpha subunit of a Heterotrimeric G-protein (with alpha subunit bound to GDP and the other two subunits)
alpha subunit releases GDP and binds GTP
alpha subunit dissociates from beta-gamma complex, now both active and able to initiate downstream effects
What is the process of inactivation for a G-protein in the G-protein coupled receptor pathway?
alpha subunit hydrolyzes GTP to GDP (with the help of a membrane bound RGS protein because its a crappy GTPase on its own) after activating its target protein
alpha reassociates with beta-gamma
Describe the downstream effects of the Gs pathway.
Gs (the alpha subunit at least) activates adenylyl cyclase which converts ATP to cAMP
four cAMP bind to the the two regulatory subunits of PKA (inactive it is two regulatory subunits and two catalytic subunits) and activates it (now regulatory subunit-cAMP complex and two separate active catalytic subunits)
which can phosphorylate targets
What are examples of targets of active catalytic subunits of PKA?
glycogen phosphorylase (breaks down glycogen) glycogen synthase (inactivated by PKA phosphorylation ensuring glycogen is not synthesized at the same time as breakdown)
What is the downstream effect of the Gi pathway?
inhibition of adenylyl cyclase
What is the downstream effect of the Gq pathway?
activates phospholipase Cbeta
phospholipase Cbeta breaks down the membrane phospholipid PIP2 into IP3 and DAG which each have different downstream targets
What is the downstream effect of IP3?
activates release of calcium inside the cell, activating PKC
among other things
What is the downstream effect of DAG?
directly activates PKC
What is the downstream effect of the Gt pathway?
activated by rhodopsin receptor, this activates cGMP phosphodiesterase which breaks down cGMP
What is the mechanism of cholera toxin?
adds an ADP-ribose molecule to the alpha subunit of Gs, resulting in its inability to catalyze the hydrolysis of GTP to GDP (it remains active permanently) keeping adenylyl cyclase activity prolonged, resulting in rises in cAMP which activates PKA
prolonged PKA sctivation (in the intestinal epithelium) causes efflux of Cl- and water into the gut –> watery diarrhea
What is the mechanism of pertussis toxin?
addition of ADP-ribose molecule to alpha subunit of Gi which prevents biding to GPCRs and thus prevents action
What are the ligands used in the Receptor Tyrosine Kinase pathway?
growth factors (platelet-derived growth factor, epidermal growth factor, fibroblast growth factor, insulin, etc)
What are the general steps of signal transduction for the Receptor Tyrosine Kinase pathway?
binding of ligand to single pass transmembrane protein (RTK) causes receptor dimerization
dimerization causes auto/trans phosphoryolation at the active site and receptors become active
phosphorylation leads to receptor;s catalytic function and creates site for target protein recruitment
What is the pathway for signal transduction of a MAP Kinase cascade in the Receptor Tyrosine Kinase pathway?
RTK is phosphorylated after ligand binding
Grb2 binds via SH2 domain (phospho-tyrosine binding)
Grb2 is constituatively bound to SOS via Grb2’s SH3 domains binding proline-rich regions in SOS
SOS activates Ras
Ras activates Raf
Raf phosphorylates MEK (a MAPKK)
MEK phosphorylates ERK (a MAPK)
What is Ras?
a small G-protein which can bind GTP (inactive) and GDP (active) and has intrinsic GTPase activity
SOS is a _____ for Ras.
GEF (guanine nucleotide exchange factor)
What is the pathway for signal transduction of the Phosphatidylinosital-3 kinase (PI-3K) pathway of the Receptor Tyrosine Kinase pathway?
PI-3 (complex of regulatory subunit and catalytic subunit) binds to RTK via the SH2 domain of the regulatory subunit (p85), allowing the catalytic subunit (p110) to access PIP and PIP2 in the membrane and phosphorylate them on the 3 position of the inositol ring
phosphoinositides phosphorylated on the 3 position activate (directly and indirectly) Akt (alternatively PKB)
this phosphorylates substrates that promote cell survival and inhibit apoptosis
PTEN removes phosphate from 3 position of phosphoinositides (tumor supressor)
What mutations in the RTK pathway most often result in cancers?
RTKs, Ras, or BRaf lead to activation in the absence of growth factors and constitutive activation of the PI-3K pathway
What are the ligands for the Cytokine Receptor pathway?
prolactin, growth hormone, interferons, erythropoietin, etc.
What are the steps of signal transduction for the Cytokine Receptor pathway?
cytokine receptors (each bound to a JAK1 and JAK2 on the intracellular portion) are dimerized by ligand, activating the JAK subunits that act as cross-phosphorylate and activate each other these phosphorylate teh intracellular position of the cytokine receptor so it can bind SH2-containing proteins (SATs) which are transcription factors that dimerize once active and travel into the nucleus to bind DNA and activate expression of specific genes
What are the ligands for the TGFbeta/BMP pathway?
teh TGFbeta/BMP family (heterodimers of type I and type II receptors)
