ERS06 Mechanism Of Action Of Peptide Hormones Flashcards
Peptide hormones
- made up of a.a.
- small - medium size
- different charges (ionisation) / polarity (polar / non-polar side chains)
- soluble alone / with carrier proteins (e.g. albumin)
Function:
- enable communication between organs / tissues
- coordinate organ activities
- form a hierarchical functional network (do not function alone)
Example:
- Calcitonin
- Glucagon (GPCR)
- Oxytocin (GPCR)
- Endorphins
- ACTH
- ADH (GPCR)
How do peptide hormones regulate cellular activities?
Peptide hormone (with different surface charges / polarity)
—> cannot penetrate plasma membrane
—> interact with specific receptor (extracellular + transmembrane + cytosolic domain)
—> change in conformation of extracellular domain
—> binding events transmitted across whole receptor
—> change in conformation of cytosolic domain
—> ***intracellular signal (mediate effect of peptide hormones)
—> cellular response
Extracellular domain: hormone specific recognition
Transmembrane domain: anchoring receptor
Cytosolic domain: generate intracellular signals
How intracellular signals act?
Qualitative model: Signalling molecule binds to inactive protein —> Active protein —> downstream events —> Cellular response
Quantitative model (how intensity can be controlled):
- More signalling molecules produced —> Greater the cellular response
- Uncontrolled production of signalling molecules —> Over / Under response
Core machinery for the controlled production of an intracellular signal
Hormone-receptor complex
—> conformational change in Intracellular domain
—> activation of **Effector protein (usually an enzyme) (e.g. Adenyl cyclase)
—> synthesise / generate **Intracellular signalling molecule (e.g. cAMP)
—> hormonal response
Coupling protein (e.g. G protein): - produce ***Quantitative effect
G protein:
- α, β, γ, subunit (trimeric protein) + GDP (bound to α)
- α subunit bound to inner plasma membrane via a fatty acid (lipid anchor)
—> activation of G-protein
—> β, γ, subunit dissociate + **GDP swapped with GTP
—> **α subunit activated by binding to GTP
—> interact spontaneously with **Adenyl cyclase (Effector protein) (only activated by GTP-bound α subunit)
—> convert ATP to **cAMP (Intracellular signalling molecule)
***G protein-based mechanism
Utilised by 7-transmembrane receptors
- Cytoplasmic domain: interact with G protein
- Extracellular domain: recognition and binding to hormone
- Transmembrane domain: transmit signal from extracellular to cytoplasmic domain
- Intracellular + Extracellular loops
- **Components of G-protein based signalling mechanism:
- Hormone
- Receptor
- Coupling protein (G protein)
- Effector enzyme (Adenyl cyclase)
- Output / Intracellular 2nd messenger (cAMP)
***Quantitative model of intracellular signalling
As long as Adenyl cyclase is bound to α subunit/GTP
—> continuously produce cAMP
—> signal increases continuously with time
—> ***over-production of signal
—> require control of activity
On-off cycle of G protein activity:
α subunit (***intrinsic GTPase activity)
—> able to catalyse GTP to GDP
—> Adenyl cyclase dissociate from α subunit/GDP (inactive Adenyl cyclase)
—> α subunit reassociate with β, γ subunit
—> inactive G protein
—> Hormone-receptor complex
—> starts another cycle of activity
—> just slow down cAMP production, still not very helpful in controlling over-production of signal
Some possible sites of control/regulation of GPCR signalling
1. Hormone-receptor complex act as **GEF (guanine nucleotide exchange factor)
—> control **rate of exchange of GDP to GTP on α subunit
-
**GAP (GTPase accelerating protein) / RGS (Regulator of G-protein signalling)
—> **promote intrinsic GTPase activity of α subunit
GEF + GAP work together —> determine how long α subunit remain in active state —> rate of cAMP synthesis
(3. β, γ subunit stabilise association of GDP with α subunit)
4. Degradation of cAMP by ***Phosphodiesterase
Summary:
Control is achieved by careful balance between:
1. Synthesis (active α subunit)
2. Degradation of signalling molecules (Phosphodisesterase degrading cAMP)
—> cAMP oscillates with time
—> Time average level of cAMP depends on frequency of oscillation
Frequency of oscillation: combined effect of synthesis and degradation
- GTP-GDP cycle on α subunit of G protein (synthesis)
- Phosphodiesterase (degradation)
Receptor-induced cAMP changes (Effector of one hormone) can also be modulated by ***another Hormone/Receptor (i.e. Inhibitory receptor):
—> also work via G protein mechanism
—> however end result is: Inhibition of Adenyl cyclase + Activation of Phosphodiesterase
—> Antagonise receptors that elevate cAMP
***Main theme of peptide hormone signal transduction
- Cell surface receptor
- 7-transmembrane receptor - Coupling protein
- G-protein - Effector protein / enzymes
- Adenyl cyclase
- Guanyl cyclase
- Phospholipase C - Intracellular signal / 2nd messenger
- cAMP
- IP3 (from phosphotidylinositol 4,5-bisphosphate: Glycerol phospholipid) —> Intracellular Ca release
- Diacylglycerol (from phosphotidylinositol 4,5-bisphosphate: Glycerol phospholipid) —> Protein Kinase C
- Fatty acids (from phosphotidylinositol 4,5-bisphosphate: Glycerol phospholipid)
- cGMP (from GTP)
Production of IP3 + DAG
Phosphotidylinositol 4,5-bisphosphate —(***Phospholipase C)—> IP3 + Diacylglycerol
- Signalling module I (**G-protein based):
Receptor activation (7-transmembrane receptor)
—> **G protein activation
—> α subunit + β, γ subunit
—> Phospholipase C activation (by either α subunit / β, γ subunit)
—> IP3 + DAG production
2. Signalling module II (***Receptor-kinase based, Trimeric G protein NOT involved): Receptor activation —> ***Receptor phosphorylation —> Phospholipase C activation —> IP3 + DAG production
Function of IP3 (Inositol 1,4,5-trisphosphate) and Diacylglycerol as signalling molecules
IP3: release ***intracellular Ca from ER (inactive: cytosolic [Ca] is low) —> activate different protein within cell
Diacylglycerol: activate ***Protein Kinase C
Receptor-kinase based signalling
- Extracellular domain
- ***ONLY 1 Transmembrane domain
- Cytoplasmic domain:
- ***Tyrosine specific protein kinase —> phosphorylation of Tyrosine residues of itself + other proteins
- ***Serine/Threonine specific protein kinase —> phosphorylation of Serine/Threonine residues of itself + other proteins
- All 3 types protein kinase —> capable of phosphorylation of all 3 a.a. residues
Tyrosine specific protein kinase example:
- ***Insulin receptor (oligomeric receptor —> when dimerise —> Tetrameric receptor)
- ***IGF-1 receptor
- NGF receptor
- VEGF receptor
Mechanism of action of EGF receptor (Tyrosine kinase receptor)
EGF bind to extracellular domain
—> 2 receptors come together to form complex (dimerisation)
—> **Cross phosphorylation of tyrosine residues on each other’s cytosolic domain by protein kinase (self-phosphorylation)
—> Protein kinase becomes more active (self-activation)
—> Tyrosine phosphate recruit + phosphorylate **Adaptor protein (多左呢樣)
—> Adaptor protein recruit + phosphorylate **Effector protein (e.g. Phospholipase C, Protein kinase, Protein phosphatase, Monomeric G-protein)
—> Production of **intracellular signals (e.g. IP3, DAG)
Overall:
Changes in protein activities in cytosol (Cytoplasmic response) + Expression of new genes in nucleus (Nuclear response)
—> Changes in cellular activities
Summary:
- **Protein phosphorylation is for **assembly of functional receptor signalling complexes
Control of Receptor-kinase based signalling mechanism
- Dephosphorylation of phosphoprotein
- Degradation of IP3, DAG
(Vs G-protein based:
- ↓ Synthesis of cAMP by GEF, GAP
- Inhibition of Adenyl cyclase
- Activation of PDE)
Regulation of Receptor abundance at cell surface
- Receptor desensitisation (**↓ Affinity)
**cAPK (cAMP dependent protein kinase)
—> Activated when **too much cAMP produced
—> Phosphorylate cytosolic domains of **all types of receptors
—> ***↓ affinity of receptors for hormone (i.e. receptor desensitised) (+ Internalisation of receptors) - Receptor endocytosis (***↓ Receptor number)
Hormone-receptor complex comes together as a patch
—> internalisation
—> Receptosome (intracellular membrane vesicle)
—> return receptors to surface later / combine with lysosome for degradation
—> avoid overstimulation of cell
GRK-arrestin pathway:
βAPK (G-protein coupled receptor kinase / GRK)
—> Activated (Phosphorylated) by **βγ subunits
—> Phosphorylate cytosolic domains (Serine/Threonine residues) of **β-adrenergic receptors
—> Recruit ***β-arrestin to bind to phosphorylated cytosolic domain of β-adrenergic receptors
—> Internalisation into endosome (Clathrin-coated vesicle)
—> receptors dephosphorylated and returned to surface later / combine with lysosome for degradation
Regulation of Hormone abundance
Production + Secretion
- Secretion control by Receptor (via Ca, Protein Kinase C)
GnRH binds to receptor
—> G protein activation
—> Phospholipase C activation
—> IP3 —> release intracellular Ca from ER —> facilitate release of FSH, LH via exocytosis
—> DAG —> activate Protein Kinase C —> facilitate release of FSH, LH via exocytosis - Production control of hormone:
- Gene transcription
- mRNA translation
Every step can be regulated: Insulin gene —> mRNA —> nascent polypeptide —> post-translational modification —> packaging into secretory vesicles —> secretion
Overall summary
Quantitative response (Magnitude + Duration) determined by:
- Level of hormone (control of synthesis and secretion)
- Intracellular signals (G-protein mechanism, Receptor-kinase based mechanism, 2nd messenger metabolism, Receptor abundance)
- Modulation by other hormones
