Second Messengers 1 Flashcards
cAMP: a secondary messenger
cyclic Adenosine MonoPhosphate (cAMP) is derived from ATP
- adenylyl cyclase cleaves a pyroshosphate (2x Pi) from ATP
- Acts through a protein kinase to stimulate glucose mobilisation, fight/flight and triglyceride breakdown
cAMP phosphodiesterase alters conformation to produce AMP, a useful intracellular metabolite
Phosphodiesterases (PDEs)
Same complexity as adenylyl cyclase
- made of a catalytic core and regulatory/targeting features
Target cAMPs and cGMPs
- inhibit cAMP (hydrolysed to AMP)
Adenylyl cyclase
ATP –> cAMP (turnover of 1000 per minute)
- found in the sarcolemma
- differential regulation by CaM (calcium binding protein) as well as the alpha and beta-gamma subunits of the heterotrimeric G-protein
GPCR activating cAMP
1) ligand (adrenaline/noradrenaline) binding to Beta-adrenergic receptor causing conformational change to bind GTP intracellularly (activation)
2) alpha subunit dissociates and binds adenylyl cyclase to activate it
all compartments in close proximity speeds up response
Role of cAMP in fight/flight response
Sudden stress triggers increased adrenaline levels which in turn increases beta-adrenergic receptor activation and cAMP production
- PDEs contain the cAMP to a specific area
- cAMP binds specific kinases to increase heart rate and force of muscle contraction (increase Ca2+ flux levels)
Importance of control of cAMP
cAMP activation must be limited to short pulses and must not be constitutive.
Pathogens disrupting cAMP regulation
1) Cholera toxin
- covalently modifies G-alpha with an ADP ribose group
- prevents GTPase activity, extending activation
- constant cAMP disturbs ion flux thus disrupting water potentials leading to cholera symptoms
2) Bordettella pertusis (Whooping cough)
- injects soluble adenylyl cyclase into host, preventing any host regulation
Control points to limit cAMP production
1) Keeping cAMP concentration low:
- destructive enzyme (PDEs) constitutively active
- Production enzymes (a cyclase) require ligand activation
2) Receptor optimised for inactivity
- can ONLY become active when ligand binds
3) Receptor internalised (endocytosed) upon binding
- reduces exposure time to curb activation levels
4) Heterotrimer G-protein activates GTPase to inactivate the alpha subunit
cAMP (cyclic 3’:5’ AMP) mode of signalling
Activate the cAMP-dependent Protein Kinase A (PKA)
- it’s autoinhibitory domain occupies kinase activity as a substrate look-alike (pseudosubstrate)
- ATP binding N-terminal lobe allows binding to protein in C-terminus (seq consensus RRxSxxxx)
Protein Kinase A downstream signalling pathways
1) Binding phospholamban (on Ser16) accelerates Ca2+ uptake into SR
2) Binding RYR2 (on Ser2808) increases Po of channel
3) Binding TnI (on Ser23) reduces sensitivity of contractile proteins
Containing reactions/pathways using Binding Partners
cAMP signalling is very localised to maximise efficiency. A-Kinase Anchoring Proteins (AKAPs) located on sub-cellular structures bind PKA along with other signalling components
- Localise signalling machinery in close proximity to substrate proteins (creating local circuits and speeding reaction response)
- Associate with E-C coupling proteins
Altered signalling outcome at receptor level:
Beta-1 and Beta-2 Adrenergic receptors
B1-AR and B2-AR both found in the heart trigger cAMP pathways but give different responses:
1) B1-AR (fight/flight)
- Substantial (400%) enhancement in contractility
- Cardiotoxicity when chronically active
2) B2-AR (very moderate)
- Modest (25%) enhancement in contractility
- Cardioprotective
Chronic cAMP affects other pathways (non-PKA) to cause disease as well
- Active Epac = hypertrophy
- Active Src = hypertrophy
Activating CaMKII
- short term = increased contractility
- long term = apoptosis
- a major target for treating heart disease due to its role in so many signalling pathways
