Signalling at Cell Surface 2 April 2007. Receptors.

Signalling at Cell Surface

2 April 2007

Receptors

Classification of receptors • Intracellular receptors (for lipid soluble

messengers)• function in the nucleus as transcription factors to

alter the rate of transcription of particular genes.

• Plasma membrane receptors (for lipid insoluble messengers)

• Receptors function as ion channels

• receptors function as enzymes or are closely associated with cytoplasmic enzymes

• receptors that activate G proteins which in turn act upon effector proteins, either ion channels or enzymes, in the plasma membrane.

Cell Surface Receptors

• May work both fast and slow

• Always use “second messengers”

Table 20-1. Characteristic Properties of Principal Types of Mammalian

Hormones

Property Steroids Thyroxine Peptides and

Proteins

Catecholamines

Feedback

regulation of

synthesis

Yes Yes Yes Yes

Storage of

preformed

hormone

Very little Several weeks One day Several days, in

adrenal medulla

Mechanism

of secretion

Diffusion through

plasma membrane

Proteolysis of

thyroglobulin

Exocytosis of

storage vesicles

Exocytosis of storage

vesicles

Binding to

plasma

proteins

Yes Yes Rarely No

Lifetime in

blood

plasma

Hours Days Minutes Seconds

Time course

of action

Hours to days Days Minutes to hours Seconds or less

Receptors Cytosolic or nuclear Nuclear Plasma

membrane

Plasma membrane

Mechanism

of action

Receptor-hormone

complex controls

transcription and

stability of mRNAs

Receptor-hormone

complex controls

transcription and

stability of mRNAs

Hormone binding

triggers synthesis

of cystolic

second

messengers or

protein kinase

activity

Hormone binding

causes change in

membrane potential or

triggers synthesis of

cystolic second

messengers

Cell-Surface Receptors Belong to Four Major Classes

• G protein coupled receptors : epinephrine, serotonin, and glucagon.

• Ion-channel receptors: acetylcholine receptor at the nerve-muscle junction.

• Tyrosine kinase linked receptors: cytokines, interferons, and human growth factor.

• Receptors with intrinsic enzymatic activity

Four classes of ligand-triggered cell-surface receptors

RECEPTOR ION CHANNELS

• multi-subunit, transmembrane protein complexes

• complex is both the receptor and ion channel

• stimuli: chemical, stretch or voltage

• stimulus induces conformational change to open or close ion channel

• two types:

1) ligand-gated ion channel

2) voltage-gated ion channel

LIGAND-GATED ION CHANNELS

• chemical stimuli bind to receptor and open or close ion channel

• stimuli can be extracellular or intracellular

EXTRACELLULAR STIMULI: (neurotransmitters)– e.g. acetylcholine, dopamine, GABA,

glutamate

INTRACELLULAR STIMULI: (second messengers)

– e.g. IP3, cAMP, cGMP, Ca2+

LIGAND-GATED ION CHANNEL AT THE SYNAPSE

• occurs at gap (synaspe) between nerve and target cell

• acetylcholine (ACh) released into synapse

• ACh binds to ion channel on target cell, opens channel, influx of Na+

• enzyme acetylcholinesterase released into synapse to

breakdown ACh

Na+ Na+

Na+

ACETYLCHOLINE ANTAGONISTS

• very potent neurotoxins

• bind to receptor and prevent opening of Na+ channel– e.g. cobratoxin from Indian cobra– atropine from deadly nightshade– S. American arrow poison (curare) - very

fast acting so shot animals don’t run too far

VOLTAGE GATED ION CHANNELS

• ion channel undergoes conformational change folllowing electrical stimulus

• this “depolarization” opens the channel– leads to flow of Na+ into cell– constitutes an “action potential”

• channel recloses

Signaling pathways downstream from G protein coupled receptors (GPCRs) and receptor tyrosine

kinases (RTKs)

Structural formulas of four common

intracellular second messengers.

Intracellular proteins

• Two groups of evolutionary conserved proteins function in signal transduction

• 1. GTPase switch proteins– Conversion from GDP bound inactive state to

GTP-bound active state is mediated by guanine nucleotide exchange factors (GEFs)

– Intrinsic GTPase activity hydrolyzes bound GTP to GDP + Pi

• GTP hydrolysis is accelerated by GTPase accelerating protein (GAPs)

• Two classes of GTPase switch proteins:– Trimeric (large) G proteins

• Directly bind to receptors

– Monomeric (small) G proteins• Linked to receptors via adapter proteins and GEFs

Common intracellular signaling proteins

• 2. Protein kinases and phosphatases– Human genome encodes 500 PKs and 100

PPs– Two types of PK

• Those that P* OH group on Tyr residue• Those that P* OH group on Ser or Thr residues

– PK is activated• By other kinases• By direct binding to other proteins• By second messengers

Regulation of signaling

• External signal decreases– Degradation of second mesenger

• Desensitization to prolonged signaling– Receptor endocytosis

• Modulation of receptor activity– Phosphorylation– Binding to other proteins

G Protein-Coupled Receptors

• A very large family of receptors coupled to trimeric G proteins

• Activate or inhibit adenylyl cyclase• All have seven membrane spanning region• Ligands include:

– Hormones, neurotransmitters, light activated receptors (rhodopsins), thousands of odorant receptors

GPCRs and G proteins are involved in the regulation of many important

physiological functions

GPCRs and G proteins are involved in the regulation of many important

physiological functions

• Signal transducing G protein has 3 subunits– G, Gß and G

• G is the GTPase switch protein and modulates the activity of an effector protein

• Effector proteins are either membrane bound ion channels or enzymes generating second messengers

• GPCR-mediated dissociation of trimeric G proteins has been demonstarted in fluorescence energy transfer experiments

GDP

GTP

+

GTP

GDP

Inactiveeffector

Activeeffector

Pi

GTP GDP

Agonist-receptor complex

Activeeffector

The activation/deactivation cycle of G proteins

The activation/deactivation cycle of G proteins

G proteins can be linked to:

• adenylate cyclase – produces cyclic AMP (cAMP)

• guanyl cyclase – produces cyclic GMP (cGMP)

• phospholipase C – produces inositol trisphosphate (IP3)

and diacyl glycerol (DAG)

• ion channels

G-Protein-Activated Enzymes

OUT

ING protein

adenylatecyclase

cAMP

adrenalin

MEM BRANE

adrenalinreceptor

firstmessenger

receptor

transducer

amplifier

secondmessenger

Activity of subunits

• Activation of K+ Channels

G-Protein-Activated Enzymes • Generate new molecules - “second messengers

G proteins and cAMP

cAMP vs PKA

cAMP and gene transcription

Epinephrine case

• Mediates body’s response to stress, when all tissues need glucose and fatty acids to produce ATP

• ß-adrenergic receptors – Heart muscle: contraction

– Smooth muscle cells of intestine: relax

2-adrenergic receptors– Smooth muscle cells of endothelium, skin, kidney and

intestine: constrict

• ß1 and ß2 adrenergic receptors are coupled to stimulatory G protein (Gs)– Actvates adenylyl cyclase

1 adrenergic receptor is coupled to inhibitory G protein (Gi)– Inhibits adenylyl cyclase

2 adrenergic receptor is coupled to Gq that activates another effector enzyme

• Bacterial toxins– Vibrio cholera

• Catalyzes chemical modification of Gs that prevents hydrolysis of GTP to GDP

– Active state

– Bordetella pertussis• Catalyzes chemical modification of Gi that

prevents release of GDP– Inactive state

• Critical domain of GPCR resides in C3 loop according to chimeric receptor expression experiments

• Differential modulation of adenylyl cyclase

• Different hormone-receptor complexes modulate the activity of the same effector molecule– In liver glucagon and epinephrine bind to

different receptors but activate the same Gs: same metabolic responses