signal transduction: signaling of inositol phospholipids

what is the phosphatidylinositol pathway?

• e.g. of receptors that use this pathway: ⍺1-adrenergic, muscarinic M1, serotonin 5-HT2, angiotensin AT1

• involves G⍺q subunit

general scheme:

• extracellular binding of signal to GPCR

• activation of phospholipase C (PLC)

what are phospholipids?

• glycerol backbone with ester bonds that can be hydrolyzed by phospholipases
• breakdown products yield intracellular second messengers

what is phospholipase C?

hydrolyzes membrane-bound phosphatidylinositol 4,5-biphosphate (PIP2) into 2 second messengers: inositol 1,4,5-triphosphate (IP3), diacylglycerol (DAG)

what is inositol 1,4,5-triphosphate (IP3)?

• water-soluble
• binds to a receptor on the membrane of the smooth ER and mitochondria
• opens the Ca2+ channel

what is diacylglycerol (DAG)?

• remains membrane-bound
• activates protein kinase C (PKC): phosphorylates other proteins, changing their activity

what are the mechanisms of phospholipase C activation?

• G proteins: ⍺ subunit of Gq/11 activates phospholipase C through an interaction at its C-terminus
• tyrosine kinase: domains of phospholipase C interact with tyrosine kinase and become phosphorylated, which turns on activity

what is the process of the phosphatidylinositol pathway?

1. ligand binds to G-protein linked receptor

2. activation of Gq⍺ subunit

• exchange GDP to GTP

3. activation of PLC

4. PIP2 is cleaved into IP3 and DAG

5. IP3 can go into the cytoplasm and bind ER, allowing for the release of Ca2+

6. DAG recruits activated protein kinase C

7. Ca2+ also activates protein kinase C

8. PKC can now phosphorylate other proteins

what is protein kinase C (PKC)?

• inactive form: synthesized as a non-phosphorylated precursor that is inactive
• active form: associated with the plasma membrane by interacting with DAG
• regulation linked to PLC activity
• examples pathways of PKC signaling: glucose/glycogen homeostasis, bronchoconstriction, constriction, ejaculation, secretion

how is PKC regulated?

• phosphorylation by phosphoinositide-dependent kinase-1 (PDK1) partially activates the enzyme
• autophosphorylation fully activates PKC
• also regulated by Ca2+

what are the mechanisms to turn off PKC?

• phosphorylation of DAG
• dephosphorylation of PKC
• degradation of PKC protein

what are adrenergic receptors?

• ligands: epinephrine, norephinephrine
• found in adrenal medulla and CNS (synaptic vesicles) and throughout body
• response: fight-or-flight; sweat, piloerection, pupil dilation, dilate bronchi, increase rate and force of heart contraction

what are the types of adrenergic receptors?

• ⍺1: couples to G⍺q; increases IP3, DAG
• ⍺2: couples to G⍺i; decreases cAMP
• β1: couples G⍺s; increases cAMP
• β2: couples to G⍺s or G⍺i; can increase/decrease cAMP

what is ⍺-receptor signaling?

• ⍺1 activation leads to PLC activation through G⍺q

• ⍺2 inhibits adenylyl cyclase and increases PLC

• regulator of G-protein signaling (RGS): shortens signal length by increasing GTP hydrolysis

what is β-receptor signaling?

• both adrenergic receptor β-subtypes can activate adenylyl cyclase: β2 receptors also have a G⍺i component
• β-arrestin and RGS can decrease signaling

what is epinephrine?

• therapeutically used in anaphylaxis and acute asthma exacerbations

concentration-dependent receptor activation:

• low concentrations: predominantly β1 and β2 receptor activation

• high concentrations: ⍺1 receptor effects become predominant

what are the physiologic effects of β-adrenergic receptor activation?

β1 receptors:

• (+) inotrophy: increased cardiac contractility

• (+) chronotropy: increased heart rate

β2 receptors:

• vasodilation leads to decreased peripheral vascular resistance and diastolic BP

• increased skeletal muscle blood flow

• bronchial smooth muscle relaxation

• increased metabolic activity (catabolism)

what is norepinephrine?

• agonism ⍺1, ⍺2, and β1 adrenergic receptors

• no effect on β2

what are the effects of norepinephrine?

• increased systolic pressure
• increase diastolic pressure and total peripheral resistance
• chronotropic effects countered by vagal response: increased stroke volume but no change in CO

what are the types of dopamine receptors?

all GPCRs:

• D1, D5: increased cAMP
• D2: decreased cAMP
• D3, D4: decreased cAMP

what is dopamine?

• precursor of norepinephrine
• systemically administered dopamine has poor CNS entry
• used for treatment of shock

what are the low-dose effects of dopamine?

D1 receptor in renal, mesenteric, coronary vasculature:

• response: vasodilation

what are the high-dose effects of dopamine?

has adrenergic β1 stimulatory activity:

• response: positive inotrope in heart

higher doses have adrenergic ⍺1 agonist activity:

• response: vasoconstriction

what are the types of ACh receptors?

• nicotinic: ion channel
• M1, M3: GPCR; increases IP3, DAG
• M2: GPCR; decreases cAMP

what are the mechanisms of GPCR regulation?

• GTPase activity: RGS hydrolyzes GTP into GDP by increasing GTPase activity
• desensitization: phosphorylate the receptor, allowing binding of beta-arrestin, which prevents binding to trimeric G protein
• receptor degredation/recycling: internalization of the receptor in the endosome which can fuse with a lysosome for degradation or be returned to the plasma membrane

what is the process of desensitization?

1. G-protein receptor kinase (GRK) phosphorylates the GPCR on particular amino acids

• increase in beta-arrestin binding

2. decreased signaling

• beta-arrestin prevents the association with the G protein

3. over the long term, beta-arrestin binding can target the receptor for endocytosis

• leads to a decreased number of receptors on the cell surface (receptor degradation)

how do beta-adrenergic receptors activate competing second messenger systems?

• agonism of the β1AR leads to the activation of G⍺s and GRK

• in chronic stimulation, there is excess GPCR signaling

GRK is cardioprotective:

• decreased signaling response to agonism

• decreased beta-arrestin-mediated receptor number

• transcriptional control and transactivation

how are beta-arrestins involved with desensitization?

• beta-arrestin leads to GPCR incorporation into clathrin-coated pits

• dynamin mediates internalization of the GPCR and plasma membrane

• release of beta-arrestin allows for recycling of receptor