Cell Signaling Flashcards
Mechanism of Ras activation
Growth factor ligand binding to RTK triggers dimerization and autophosphorylation of the RTK; SH2 domain in Grb2 adaptor protein binds to phosphotyrosine residues on the RTK; Grb2 SH3 domains bind proline-containing peptide Sos, a Ras GEF; Sos activates Ras through proximity
Mechanism of TKI vs. Antibodies
Antibodies (i.e. Cetuximab) block the extracellular ligand binding site on EGFR, preventing dimerization
Targeted kinase inhibitors (TKIs) i.e. Gefinitib bind in the phosphorylation site of EGFR, blocking ATP from binding and preventing phosphorylation & downstream signaling
Tumor cell characteristics predictive of response to EGFR TKI
Patients whose tumor cells show gain-of-function mutations in EGFR are good candidates for TKIs, i.e. Gefitinib; EGFR amplification or overexpression as determined by FISH
3 Mechanisms for resistance to TKIs
- Acquired resistance via second mutations that arise under selective pressure from TKI therapy
- Pathological activation of other receptors, i.e. ErbB2
- Primary resistance - i.e. a tumor with a Ras mutation downstream of the receptor will be unaffected by TKIs
4 types of signaling receptors
Ligand-gated ion channels
GPCRs
Enzyme-linked receptors (including RTKs)
Nuclear receptors
5 examples of second messengers
Ca2+ cAMP IP3 DAG NO
5 mechanisms of signaling termination
Re-uptake, degradation, or diffusion of an extracellular signaling molecule
Phosphatases - de-phosphorylate kinase cascades
Phosphodiesterases - hydrolyze cAMP and cGMP; activity increased by allosteric binding of substrate and phosphorylation by c-NMP dependent kinases
Intrinsic GTPase activity
Constitutively active terminators, i.e. Ca2+ pumps
Signal pathway nodes
Nodes are points in a signaling network that receive multiple inputs and/or contribute to multiple outputs
i.e. Ca2+
4 types of cellular signaling
Paracrine - from signaling cell to target cell over a short distance
Contact-dependent - requires physical contact between a membrane-bound mediator on the signaling cell and a receptor on the target cell
Endocrine - from signaling cell to target cell over a long distance
Synaptic - mediated by release of neurotransmitter at a synaptic cleft
Characteristics of lipophilic signaling molecules
Lipophilic signaling molecules can penetrate the cell membrane and bind an intracellular receptor; they cannot be stored intracellularly and therefore they are regulated by synthesis only
Ex: Steroid hormones
Characteristics of hydrophilic signaling molecules
Hydrophilic signaling molecules cannot penetrate the cell membrane and therefore must bind receptors on the cell surface; they can be stored intracellularly within the signaling cell and therefore their release is regulated by vesicular release
Ex: Peptides
G-protein coupled receptor structure
7 transmembrane domain protein with N-terminus oriented toward the cytosol; 7 TM domains fold to form a barrel structure which contains an extracellular ligand-binding domain; intracellular loops and C-tail mediate conformational changes that activate G-protein
G protein signaling
Inactivated GCPRs are bound intracellularly to a heterotrimeric G-protein, composed of an alpha subunit(-GDP) and a beta/gamma subunit; agonist binding triggers a conformational change that favors the disassociation of GDP from the alpha subunit; GTP quickly binds to and activates G-alpha; G-alpha dissociates from beta-gamma and both subunits diffuse through the membrane to reach their effector proteins
Inactivation of G-protein signaling
G-alpha is a GTPase, which hydrolyzes GTP to GDP; hydrolysis triggers G-alpha-GDP to re-bind the beta-gamma subunit as well as the receptor;
GAPs accelerate the process of GTP hydrolysis, shortening the lifespan of the signaling pathway
Switch II
The switch II region of G-alpha is responsible for binding the beta-gamma subunit in its GDP-bound, inactive state; GTP-binding frees up the switch II region to dissociate from beta-gamma, activating the two subunits
Pertussin
PTX enzymatically puts an ADP-ribose onto G-alpha near the C-terminus of the alpha subunit to lock the G protein into it’s inactive state
Cholera
CTX ADP-ribosylates Gs-alpha near the ATP binding site, inhibiting ATPase activity, leading to constitituve activation of the G-protein
2 classes of GCPRs
- Adrenergic Receptors (Norepinephrine, sympathetic)
2. Muscarinic Receptors (Acetylcholine, parasympathetic)
Beta-1 adrenergic receptor
Binds norepinephrine to activate Gs protein; G-alpha subunit activates adenylyl cyclase, converting ATP to cAMP; cAMP activates PKA; PKA phosphorylates voltage-gated Ca2+ channels and Ryanodine Receptors in the SR, increasing intracellular concentration of calcium, leading to increased heart rate and contractility
Antagonists: propranolol, metoprolol (HTN)
Alpha-1 adrenergic receptor
Alpha-1 adrenergic receptor binds norepinephrine; activation of Gq protein activates PLC, which cleaves membrane lipid PIP2 into IP3 (cytosolic) and DAG (membrane-bound);
IP3 binds to IP3-receptor in ER and releases Ca2+
DAG-PKC stimulates Ca2+ through L-type, voltage gated Ca2+ channels
Increased intracellular Ca2+ stimulates smooth muscle contraction in peripheral vasculature, shifting blood flow from the skin to the viscera, increasing blood pressure
Antagonist: Prazosin (HTN)
m2-muscarinic cholinergic receptor signaling via alpha in the heart
M2 AchR binds agonist Ach; activated G-i protein antagonizes the effect of G-s on adenylyl cyclase; G-i can dominantly inhibit AC and the production of c-AMP; cAMP is degraded by PDE and so the sympathetic response is shut down, leading to decreased heart contraction
m2-muscarinic cholinergic receptor signaling via beta-gamma in the heart
Ach binds m2 AChR, activating G-i protein; activated beta-gamma subunit activates the GIRK membrane-bound K+ channel, which allows K+ efflux from the cell, causing cellular hyperpolarization and decreased excitability, leading to decreased heart rate contraction
Antagonist: Atropine, increases heart rate
Beta-2 adrenergic receptor signaling in the lungs
Epinephrine binds B2AR, activating Gs protein; Gs alpha subunit activates AC, which produces cAMP; cAMP activates PKA; PKA phosphorylation inhibits smooth muscle contraction, leading to bronchodilation
Agonist: Albuterol
m3-muscarinic cholinergic receptor signaling in lungs
ACh binds m3AChr, activating Gq protein; Gq-alpha activates PLC, which cleaves PIP2 into IP3 and DAG leading to Ca2+ release and increased bronchoconstriction
Antagonist: Ipratropium, asthma inhaler
