Biotech 5: Signal Transduction 1

cells need signals to

1. survive

2. grow + divide

3. differentiate

4. die

what are the different types of signaling?

• contact-dependent

• paracrine (local signaling)

• endocrine (signals through blood)

• synaptic (neurons)

• autocrine (self)

SEQ basic composition of transduction pathway

• extracellular signal molecule

• receptor protein

• intracellular signaling molecule

• effector protein

What are the different kinds of effector proteins?

What does each lead to?

1. metabolic enzyme —> altered metabolism

2. transcription regulatory protein —> altered gene expression

3. cytoskeleton protein —> altered cell shape or movement

what are the four major categories of receptors

1. gated channels (require ligand)

2. g-protein coupled receptors

3. enzyme linked receptors

4. Intracellular Receptor (ligand must get into cell)

What makes intracellular receptors different from cell surface receptors?

ligands may use carrier proteins to get to cell

can diffuse into cell

receptor may be inside cell OR on nucleas

do hydrophobic or hydrophilic ligands bind to intracellular receptors?

hydrophobic can cross membrane

hydrophilic bind to receptor (don’t come in)

growth factors, cytokines, hormones, and neurotransmitters are all examples of

ligands

what do kinase’s do?

add phosphate groups to substrates at their OH groups

What are the two main types of protein kinases?

1. Serine/ Threonine Kinase

2. Tyrosine Kinase

what does serine/threonine kinase do?

turn serine into phospho-serine

turn threonine into phospho-threonine

what do tyrosine kinases do

turn tyrosine into phospho-tyrosine

what can kinases add phosphates to?

• proteins

• lipids

• carbohydrates

what is P13 kinase?

lipid kinase important in cell signaling

phosphorylates membrane lipid phosphoinositide (Pl)

GTP binding proteins:

how are they activated? how are they deactivated?

activated when bind to GTP

deactivated when bind to GDP

what are the two major types of GTP binding proteins?

1. large HETEROTRImeric

2. small MONOmeric

__________ and ________ play important roles in regulating monomeric G protein activity

GAP and GEF

what is GAP? what does it do?

GAP: GTPase Activating Protein

activate GTPase which breaks apart GTP DEACTIVATING G-protein coupled receptor

what is GEF? what does it do?

Guanine Nucleotide Exchange Factor

promote the RELEASE of GDP, allowing new GTP to bind. ACTIVATE g-protein receptor

what binds to SH2 domains?

phosphotyrosine

what binds to SH3 domains?

proline rich domains

PTB domains bind to

phospho-tyrosine but with different specificity

PH domains bind to

phosphorylation sites on phosphoinositides

which cellular responses are slow?

which cellular responses are fast?

• altered protein function

• changes in gene expression

Cells can become desensitized to the signal over time. What are some different ways that cells can become desensitized to a stimulus?

1. negative feedback

2. delayed feed-forward

3. receptor inactivation

4. receptor sequestration (receptor inside endosome)

5. receptor destruction (receptor inside lysosome)

what pathways can GPCRs mediate

1. cell proliferation

2. cell differentiation

3. cell migration

4. ion regulation

5. sight

6. smell

7. taste

what ligands bind to GPCRs

• proteins

• peptides

• amino acid derivatives

• fatty acids

• photons of light

what are the three subunits of Trimeric G proteins?

• alpha

• beta

• gamma

the alpha G subunit can be classified into what 4 major groups

1. G12/13

2. Gi

3. Gs

4. Gq

what are the two different types of GPCR receptors? what are their subtypes?

1. a-adrenergic (a1 + a2)

2. b-adrenergic (b1,b2,b3)

What G-protein binds to the a-adrenergic receptor?

Gq and Gi

what G-protein binds to the b-adrenergic receptor

Gs

what does the a-adrenergic receptor (a-1) bind to?

what does it activate/ inhibit?

what happens upon activation/ inhibition?

• Gq

• activates phospholipase C (PLC)- P13-DAG

• increases calcium IN CELL causing vasoconstriction of vascular smooth muscle

what does the a-adrenergic receptor (a-2) bind to?

what does it activate/ inhibit?

what happens upon activation/ inhibition?

• Gi

• Inhibits adenylate cyclase

• decrease cAMP inhibiting neurotransmitter release from pre synaptic to post synaptic

what does the b-adrenergic receptor (b-1) bind to?

what does it activate/ inhibit?

what happens upon activation/ inhibition?

• Gs

• activates adenylyl cylase + cAMP

• increase heart rate and contraction force

what does the b-adrenergic receptor (b-2) bind to?

what does it activate/ inhibit?

what happens upon activation/ inhibition?

• Gs

• activate adenylate cyclase + cAMP

• smooth muscle relaxation and bronchodilation

what does the b-adrenergic receptor (b-3) bind to?

what does it activate/ inhibit?

what happens upon activation/ inhibition?

• Gs

• activate adenylate cyclase + cyclic cAMP

• regulation of thermogenesis in adipose tissue

what are the two different types of signaling pathways for G-protein coupled receptors?

1. adenylate cyclase

2. phospholipase-C

what are the components of the Adenylate Cyclase pathway?

• cAMP

• PKA

• CREB

what are the components of the Phospholipase-C (PLC) pathway?

• PLC

• DAG

• IP3

• Calcium

• PKA

which g-protein subunit and receptor ACTIVATES the adenylate cyclase pathway?

which g-protein subunit and receptor INHIBITS the adenylate cyclase pathway?

• activate = Gs (b-adrenergic receptor)

• deactivate = Gi (a2 adrenergic receptor)

SEQ Adenylate Cyclase G-Protein Pathway

1. ligand binds to b-adrenergic receptor

2. Gs protein binds to b-adrenergic receptor

3. GDP removed from Gs

4. GTP bind to Gs

5. G protein released from GPCR

6. Gs unbinds from Beta Y unit

7. Both Gs and GBY interact with targets (enzymes and ion channels)

8. Adenylyl cyclase converts ATP to cAMP

9. Protein Kinase (PKA) adds phosphates to CREB (transcription factor)

10. by regulating CREB, PKA can regulate target genes that have CREB binding sites in their promoters

what is most often the ligand in the adenylate cyclase pathway? where do they bind to? what is the typical response?

norepinephrine and epinephrine

b-adrenergic receptors

fight or flight

which tissues can the Gs pathway impact?

1. cardiac tissue

2. liver

3. adipose tissue

4. central nervous system

5. bronchial smooth muscle

How does the Gs pathway impact cardiac tissues?

• Increased Heart Rate: PKA phosphorylates two channel proteins leading to faster action potentials

• Increased Contractility: PKA phosphorylates proteins involved in contraction (higher blood pressure)

How does the Gs pathway impact the liver ?

increased glucose in bloodstream

• PKA phosphorylation of CREBS leads to expression of genes involved in the regulation and synthesis of glucose

• PKA also phosphorylates proteins important in releasing glucose from glycogen stores

How does the Gs pathway impact adipose tissue?

Breakdown of Stored Fat

• PKA phosphorylation of CREB leads to increased expression of gene that encodes an enzyme that breaks down triglycerides

• PKA also directly phosphorylates enzymes that break down triglycerides

How does the Gs pathway impact the central nervous system?

epinephrine acts as ligand

• PKA phosphorylates CREB leading to expression of genes that are associated with synaptic plasticity, neurotransmitter release, and neuronal excitability

• PKA also phosphorylates ion channels leading to enhanced neurotransmission

How does the Gs pathway impact the bronchial smooth muscle?

Muscle relaxation and bronchodilation

• PKA phosphorylation of CREB regulates expression of genes involved in contractility

• PKA phosphorylates mysosin light chain kinase INHIBITING myosin light chain phosphorylation leading to muscle relaxation

Which three drugs BLOCK the action of beta 1 adrenergic receptors? Which pathway does this block?

• beta blockers (dipines)

• inderal (propanol)

• tenormin (atenolol)

blocking beta 1 adrenergic receptors blocks the Gs (adyenylate cyclase) g-protein coupled response

blocking beta adrenegic receptors can relieve

• hypertension

• angima

• arrythmias

is ventolin (albuterol) a beta adrenergic 2 agonist or antagonist? what does it do?

beta adrenergic 2 AGONIST

mimics epinephrine by binding to Beta-adrenergic 2 receptor

leads to bronchodilation

which g-protein subunit and receptor mediates the phospholipase- C (PLC) pathway?

G protein = Gq

receptor = a1 adrenergic

what ligands typically bind to the a-adrenergic receptor of the PLC pathway?

norepinephrine + epinephrine

SEQ phospholipase- C (PLC) Pathway

1. ligand binds to a-adrenergic receptor

2. Gq alpha subunit attaches to reAceptor

3. Activation of phospholipitase C (PLC)

4. phospholipitase cleaves PIP2 into IP3 and DAG

5. DAG is membrane-bound, IP3 released to cytoplasm

6. IP3 binds to ligand-gated CALCIUM channels on ER membranes leading to calcium release

7. Calcium + DAG regulate PKC which phosphorylates MANY substrates

is phospholipase an enzyme? is it membrane bound?

yes it is a membrane bound enzyme

Phospholipase C (PLC) pathway ultimately leads to increased calcium levels by IP3

What can calcium control?

1. contraction

2. secretion

3. activation

of signaling pathways

Calcium cannot work on its own. Calcium can use _________________ (which holds 4 of them) to do its function.

calmodulin

One calmodulin is bound to 4 carbons, what can it interact with? what can then happen?

CaM - Kinase (calcium calmodulin dependent protein kinase)

CaM Kinase can phosphorylate CREB transcription factor

how does the G1-PLC-DAG-IP3-Ca2+(PLC) pathway effect smooth muscle contraction?

Calcium-calmodulin complex activates myosin light chain kinase (MLCK) —> muscle contraction

how does the G1-PLC-DAG-IP3-Ca2+ (PLC) pathway effect platelet aggregation?

increase

Ca2+ activates several Ca2+ dependent enzymes such as the ones involved in platelet aggregation

how does the G1-PLC-DAG-IP3-Ca2+ (PLC) pathway effect neurotransmission?

Ca2+ influences neurotransmitter release, synaptic transmission, and modulation of neuronal excitability

how does the G1-PLC-DAG-IP3-Ca2+ (PLC) pathway affect immune responses?

Ca2+ contributes to immune cell activation, proliferation, and cytokine production

how does the G1-PLC-DAG-IP3-Ca2+ (PLC) pathway affect hormone secretion?

ca2+ stimulate the release of hormones from endocrine cells

how does the G1-PLC-DAG-IP3-Ca2+ (PLC) pathway affect vision?

Ca2+ modulate photoreceptors sensitivity to light

How does asprin impact the PLC pathway?

impacts Gq indirectly

• blocks COX-1 which inhibits production of thromboxane receptor which Gq binds to

reduces platelet aggression (inhibit clotting) —prevent strokes and heart attacks

Prolonged exposure to a stimulus can lead to ____________ of the GCFR

desensitization

GRK (GPCR kinase) phosphorylates which parts of the GPCR?

What happens one GPCR is phosphorylated?

Serines and threonines

GPCR binds to Arrestin (Arr)

what does Arrestin do?

binds to GPCRs that are phosphorylated at their serine and theronines and PREVENTS GPCR from interacting with G PROTEIN

also deals with endocytosis of GPCR

how does Tryvio (aprocitentan) treat high blood pressure? Tryvio was the first of its kinda after statins to be able to accomplish this!

• acts as antagonist of GPCRs on smooth muscle (ET-A and ET-B) and

endothelin binds to ET-A and ET-B receptors which can lead to over-activation and high blood pressure

does VELSIPITY (etrasimod) inhibit or enhance GPCRs? what does it do?

inhibits S1PR (sphingosine-1-phosphate receptor)

S1PR regulates movement of immune cells from the lymph nodes to the intestine

relieves inflammation in the digestive system

Does Zavzepret inhibit or enhance GPCR?

What does it do?

inhbits CALCRL on neurons

CGRP (CALCRL ligand) causes migraines

CGRP needs RAMP1 to bind to CALCRL

Zavzepret binds to RAMP1—CALCRL complex, not allowing CGRP to bind to the complex

PREVENTS VASODILATION WHICH CAUSES MIGRANES

Is Ohtuvayre (ensifentrine) inhibitory or stimulating?

inhibits two phosphodiesterases so ATP is not converted to cyclic ATP