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Physiology Chapter III: Signal Transduction

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Mechanisms of cellular communication

o Some cells communicate with each other by direct contact through gap junctions
Neighboring cells communicate with each other by direct contact through gap junctions
formed between apposing cell membranes
Water-filled channels facilitate the passage of inorganic ions and small molecules such
as Ca2+ and cAMP
Gap junctions are also excellent pathways for the flow of electrical current, which plays a
critical role in coordinating the electrical activity in cardiac and smooth muscle
Gap junctions consists of two half channels that are arranged end to end in the
membranes of adjacent cells
Has six connexin molecules surrounding a hollow core
The permeability of gap junctions can be rapidly regulated by changes in cytosolic
concentrations of Ca2+, cAMP and H+
o Cells can communicate with one another via chemical signals
A substance that is produced in one tissue or organ and released into the blood
and carried to other organs (targets), where it acts to produce a specific
3 ways:
Endocrine: on distant tissues
Paracrine: on a neighboring cell in the same tissue
o Neuromuscular junctions
Autocrine: on the same cell that released the signaling molecule
o Chemical signals interact with target cells by binding to surface or intracellular receptors
Four types of chemicals can serve as extracellular signaling molecules
o Epinephrine
Peptides and proteins
o Angiotensin II
o Insulin
o Aldosterone
o Estrogens
o Retinoic acid
Small molecules
o Amino acids
o Nucleotides
o Ions (Ca+)
o Gases (nitric oxide)
For a molecule to act as a signal, it must bind to a receptor
A protein on the cell surface or within the cell that specifically binds a signaling
molecule (ligand)
The receptor is itself an ion channel, and ligand binding produces a change in
Receptors can be divided into four categories based on their associated
mechanisms of signal transduction
o Ligand-gated ion channels
Integral membrane proteins
Involved in signaling between electrically excitable cells

Physiology Chapter III: Signal Transduction

G-protein-coupled receptors
Work indirectly through an intermediary to activate or inactivate a
separate membrane-associated enzyme or channel
o Catalytic receptors
Enzymes themselves or part on an enzymatic complex
o Intracellular (or steroid-type) receptors
Proteins located in the cytosol or nucleus
Ligand activated transcription factors
Link extracellular signals to gene transcription
o Transmembrane proteins that release transcription factors
Signaling events initiated by plasma membrane receptors can generally be
divided into six steps:
o Step 1: Recognition of the signal by its receptor
Involves three types of noncovalent interactions
Ionic bonds
Van der Waals interactions
Hydrophobic interactions
o Step 2: Transduction of the extracellular message into an intracellular
signal or second messenger
Causes a conformational change in the receptor
Final consequences is the generation of a second messenger or
the activation of a catalytic cascade
o Step 3: Transmission of the second messengers signal to the
appropriate effector
Effectors include
Transport proteins
Contractile elements
Transcription factors
o Step 4: Modulation of the effector
Result in the activation of protein kinases and phosphatases
o Step 5: Response of the cell to the initial stimulus
o Step 6: Termination of the response by feedback mechanisms at any or
all levels of the signaling pathway
Second-messenger systems amplify signals and integrate responses among cell
o The involvement of second messengers in catalytic cascades provides
numerous opportunities to amplify a signal
o Second-messenger systems also allow for specificity and diversity
Regulated intramembrane proteolysis of certain transmembrane proteins
releases transcription factors
o Regulated intramembrane proteolysis (RIP)
Membrane proteins undergo proteolytic cleavage within the plane
of the membrane to liberate cytosolic fragments that enter the
nucleus to affect gene expression
Membrane proteins includes:
o Sterol regulatory element-binding proteins

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Serves an essential role in the control of cholesterol

Resident protein of the ER membrane
Cytoplasmic domain is liberated when cells are
depleted of sterols

o Plasma membrane receptor whose cytoplasmic
domain is released in response to Delta
A membrane-bound ligand that dictates cell
fate throughout development
Irel and ATF6
o The most recently recognized examples of RIP
o Transmembrane ER proteins that release their
cytoplasmic domains in response to the presence
of unfolded secretory proteins in the ER
o Amyloid precursor protein
o A protein of unknown function that is cleaved in the
membrane to produce the extracellular amyloid
o Implicated in Alzeimers disease
The intramembrane cleavage does not take place until a primary
cleavage event removes the bulk of the protein on the
extracytoplasmic face

Receptors that are ion channels

o Ligand-gated ion channels transducer a chemical signal into an electrical signal
The signaling molecule itself controls the opening and closing of an ion channel by
binding to a site on the receptor
These receptors are also called ionotropic receptors
Ionotropic receptors for Ach
- aminobutyric acid (GABA)
IP3 receptor
Ca2+ -release receptor (ryanodine receptor)
Most structural and functional information for ionotropic receptors come from the
nicotinic Ach receptor (AChR) present in skeletal muscle
Cation channel that consists of four membrane-spanning subunits
Stoichiometry of 2:1:1:1
Called nicotinic because nicotine contained in tobacco can activate or open the
channel and thereby alter Vm
Receptors linked to G proteins
o G-protein-lined receptors constitute the largest family of receptors on the cell surface
o Mediate cellular response of signaling molecules such as

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Vasoactive peptides
o Structures:
Consist of a single polypeptide chain with seven membrane-spanning helical segments
An extracellular N terminus that is glycosylated
A large cytoplasmic loop that is composed mainly of hydrophilic amino acids between
helices 5 and 6
Appears to be the major site of interaction with the G protein (3,4 also contribute
to binding in some cases)
A hydrophilic domain at the cytoplasmic C terminus
General properties of G proteins
o G proteins are heterotrimers that exist in many combinations of different , , and subunits
G proteins are members of a superfamily of GTP-binding proteins, as well as the socalled small GTP-binding proteins such as Ras
Heterotrimeric G proteins are composed of three subunits
16 subunits
o Binds and hydrolyzes GTP
o Also interacts with downstream effector proteins such as adenylyl cyclase
o Held to the membrane by either
Myristyl group
Palmitoyl group
5 subunits
11 subunits
o Held to the membrane by prenyl group
Adenylyl cyclase
Among the first effector found to be sensitive to G proteins
Gs: stimulates adenlylyl cyclase
Gi: responsible for the hormone-dependent inhibition of adenylyl cyclase
o G-protein activation follows a cycle
Receptor (R) consists of seven membrane-spanning segments
Ligand binds, receptor activates
Receptor interacts with the G protein to promote a conformational change and the
exchange of GDP for GTP
G protein dissociates from the receptor
-GTP and subunits dissociate
Both -GTP and can now interact with their appropriate effectors (E1, E2)
-catalyzed hydrolysis of GTP to GDP inactivates and promotes reassembly of the
members of the RGS family of G-protein regulators stimulate GTP hydrolysis with some
but not all subunits
o Activated subunits couple to a variety of downstream effectors, including enzymes, ion
channels, and membrane-trafficking machinery
Adenylyl cyclase
Is activated by G s
Catalyzes the production of cAMP from ATP
G i inhibits adenylyl cyclase and thus decrease cAMPi
G proteins can also activate enzymes that break down cyclic nucleotides
o Plays a key role in phototransduction

Physiology Chapter III: Signal Transduction

Activates the cyclic guanosine monophosphate phosphodiesterase

Which catalyzes the breakdown of cGMP to GMP
G proteins can also be coupled to phospholipases
These enzymes catabolize phospholipids
Can be grouped into phospholipases A2, C, or D based on the site at which the
enzyme cleaves the phospholipids
The G-protein q subunit activates phospholipase C (PLC), which breaks
phosphoinositol biphosphate (PIP2) into 2 intracellular messengers
o Membrane-associated DAG
Stimulates protein kinase C (PKC)
o Cytosolic IP3
Binds to a receptor on the ER membrane and triggers the release
of Ca2+ from intracellular stores
Some G proteins interact with ion channels
o The subunits of G proteins can also activate downstream effectors
complex interacts with a particular class of K+ channels
complex bind to a special protein kinase called the -adrenergic receptor kinase
(ARK) where it phosphorylates the ligand-receptor complex
o Small GTP-binding proteins are involved in a vast number of cellular processes
Ras proteins
21-kDa membrane-associated proteins that bind guanine nucleotides
Three isoforms of Ras (N, Ha, Ki)
o Relay signals from the plasma membrane to the nucleus via an elaborate
kinase cascade
2 classes of regulatory proteins modulate Ras activity
o GTPase-activating proteins GAP and neurofibromin
These proteins increases the rate at which Ras hydrolyzes bound
GTP and thus becomes inactive
o Guanine nucleotide exchange proteins such as son of sevenless (SOS)
Which promote the conversion of inactive Ras-GDP to active RasGTP
G-protein second messengers: cyclic nucleotides
o cAMP usually exerts its effect by increasing the activity of protein kinase A
Excess cAMP is also responsible for certain pathologic conditions
McCune-Albright syndrome
cAMP exerts many of its effects through cAMP-dependent protein kinase A (PKA)
This enzyme catalyzes transfer of the terminal phosphate of ATP to certain serine
or threonine residues within selected proteins
One important control mechanism is the use of regulatory subunits that constitutively
inhibit PKA
4 subunits
o Two regulatory subunits
o Two catalytic subunits
Binding of cAMP to the regulatory subunits induces a conformational change in these
proteins that diminishes their affinity for the catalytic subunits
The complex dissociates and the catalytic subunits are free to catalyze the
phosphorylation of protein substrates
Another mechanism that contributes to regulation of PKA is the targeting of the enzyme
to specific subcellular locations

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PKA targeting is achieved by the association of a PKA regulatory subunit with an

A kinase anchoring protein (AKAP)
Over 35 AKAPs are known
o Epinephrine stimulates glycogen breakdown and inhibits glycogen synthesis via cAMP
A glucose polymer stored primarily in liver and muscle cells
Major storage form of carbohydrate in the body
Binding of epinephrine to its adrenergic receptor results in activation of PKA, which
phosphorylates three enzymes
PKA phosphorylates the regulatory and subunits of inactive glycogen
phosphorylase kinase (PK)
o Subunit structure: ()4
o is also regulatory
o only the subunits are involved in catalysis
PKA phosphorylates the active form of glycogen synthase (GS) and renders it
PKA also inactivates phosphoprotein phosphatase-1 (PP1)
o Protein phosphatases reverse the action of kinases
Another way to terminate the action of cAMP is to dephosphorylate effector proteins
mediated by enzymes called serine/threonine phosphoprotein phosphatases
Four groups of serine/threonine phosphoprotein phosphatases:1, 2a, 2b, 2c
These enzymes are regulated by phosphorylation at their serine, threonine, and
tyrosine residues
The Ca2+-dependent PP2b
Also known as calcineurin
Most prevalent in the brain
The target of the immunosuppressive drugs FK-506 and cyclosporine
The first phosphotyrosine phosphatase (PTP) to be characterized was the cytosolic
enzyme PTP1B from human placenta
o During phototransduction, a decrease in [cGMP]i inhibits a nonselective cation channel
G-protein second messengers: products of phosphoinositide breakdown
o Many messengers bind to receptors that activate phosphoinositide breakdown
Phosphatidylinositols (PIs) can be phosphorylated to yield the two major
phohshoinositides that are involved in signal transduction
Phosphatidylinositol biphosphate (PI4,5P2)
Extracellular signals that activate receptors linked to certain G proteins stimulate
phospholipase C (PLC) which breaks down PIP2 into
o Travels through the cytosol to stimulate Ca2+ release from intracellular
o Remains in the plane of the membrane to activate PKC which migrates
from the cytosol and binds to DAG in the membrane
PLCs are classified into three families (, , )
Differ in their
o catalytic properties
o Cell type specific expression
o Modes of activation
Phosphatidylcholine (PC)

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Abundant phospholipids in the cell membrane

Also a source of DAG
The cell can produce DAG from PC by either 2 mechanisms
o PLC can directly convert PC to phosphocholine and DAG
o Phspholipase D, by cleaving the phosphoester bond on the other side of
the phosphate, converts PC to choline and phosphatidic acid (PA)
PA can then be converted to DAG via PA-phosphohydrolase
Production of DAG from PC produces the slow wave of increasing cytosolic DAG
Factors that stimulate the production of DAG from PC
o Tumor necrosis factor- (TNF- )
o Interleukin-1 (IL-1)
o Interleukin-3 (IL-3)
o Interferon- (IFN- )
o Colony-stimulating factor
DAG can be further cleaved by DAG lipase to arachidonic acid
Can be metabolized to other signaling molecules, the eicosanoids
Inositol triphosphate liberates Ca2+ from intracellular stores
Receptor for IP3 is a ligand-gated Ca2+ channel located in the membrane of the ER
The receptor is a substrate for phosphorylation by PKA, PKC, and
Ca2+/calmodulin (CAM)-dependent protein kinases
Release of Ca2+ from intracellular stores is terminated by dephosphorylation of IP3
An ATP-fueled Ca2+ pump (SERCA) then moves the Ca2+ back into the ER
Some of the IP3 is further phosphorylated to IP4
In addition to IP3, cyclic ADP ribose (cADPR) can mobilize from intracellular Ca2+ stores
For Ca2+ to function as a second messenger, it is critical that [Ca2+]i be normally
maintained at relatively low levels (at or below ~100 nM)
Calcium activates calmodulin-dependent protein kinases
The effects of changes in [Ca2+]i are mediated by Ca2+-binding proteins, the most
important of which is calmodulin (CaM)
Each molecule of CaM cooperatively binds four calcium ions
Many of the effects of CaM are mediated by protein phosphorylation that is catalyzed by
a family of Ca2+-CaM-dependent kinases (CaM kinases): narrow substrate specificity,
can also phosphorylate itself thus remain active
Myosin light chain kinase
o An important CaM kinase in smooth muscle cells
Glycogen PK
o Subunit structure ()4
o and (subtrates for PKA)
o subunits are in fact CaM
o Thus an increase in [Ca2+]i can also activate PK
CaM kinase II
Broad substrate specificity
One action is to phosphorylate and thereby activate the rate-limiting enzyme in
the synthesis of catecholamine neurotransmitters
Diacylglycerols and Ca2+activate protein Kinase C
The most important function of DAG is to activate protein kinase C (PKC), a Ca2+dependent serine/threonine kinase
G-protein second messengers: arachidonic acid metabolites
Other hydrolysis products of membrane phospholipids can also act as a signalling

Physiology Chapter III: Signal Transduction

Hydrolysis product as in arachidonic acid

Which is found esterified at the second carbon of the glycerol backbone of
membrane phospholipids
Is converted to a biologically active metabolites called eicosanoids
3 pathways can convert AA to eicosanoids
o Cyclooxygenase enzymes produce thromboxanes, prostaglandins, and
o 5-lipoxygenase enzymes produce leukotrienes and some
hydroxyeicosatetraenoic acid (HETE) compounds
o Epoxygenase enzymes, which are members of the cytochrome P-450
class, produce other HETE compounds as well as cis-epoxyeicosatrienoic acid (EET) compounds
Eicosanoids have powerful activites including
o Allergic and inflammatory processes
o Platelet aggregation
o Vascular smooth muscle
o Gastric acid secretion
Phospholipase A2 is the primary enzyme responsible for releasing arachidonic acid
The G-protein dimer may stimulate PLA2 either directly or via mitogen-activated
protein (MAP) kinase which phosphorylates PLA2 at a serine residue
Results is rapid hydrolysis of phospholipids that contain AA
Agonists acting on other receptors may promote AA release indirectly
First, A ligand may bind to a receptor coupled to PLC, which would lead to the
release of DAG
o DAG lipase can cleave DAG to yield AA and a monoacylglycerol (MAG)
Second, any agonist that raises [Ca2+]i can promote AA formation because
Ca2+can stimulate some cytosolic forms of PLA2
Third, any signal transduction pathway that activates MAP kinase can also
enhance AA release because MAP kinase phosphorylates PLA2 on a serine
Cyclooxenase, lipoxygenases, and epoxygenases mediate the formation of biologically active
First pathway, Cyclooxygenase catalyze the stepwise conversion of AA into the
intermediates prostaglandin G2 (PPG2) and prostaglandin G2 (PGH)2
COX exists into 2 forms
o COX-1
o COX-2
May participate in the inflammatory response
Inhibited by aspirin
(PGH)2 is the precursor of the other
o Prostaglandins
o Prostacyclins
o Thromboxanes
Second pathway, 5-hydroxygenase initiates the conversion of AA into biologically active
LTE4, D4, and E4 are the cysteinyl leukotrienes
o Slow reacting substance of anaphylaxis
o Participate in allergic and inflammatory responses
Third pathway, begins with the formation of AA by epoxygenase
This pathway converts AA into two major products

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o EETs
o Prostaglandins, Prostacyclin, and Thromboxanes (COX products) are vasoactive, regulate
platelet action, and modulate ion transport
o Platelet aggregation
o Constriction of the airways
o Rennin release and inflammation
o Vasoactive
o Regulation of renal blood flow
Thromboxane A2
o Short-lived compound that can aggregate platelets
o Bring about the platelet release reaction
o Constrict small blood vessels
Prostacyclin I2
o Also known as PGI2
o Inhibits platelet aggregation and dilates blood vessels
Eicosanoids may participate in regulation of the Na-K pump
o The leukotrienes (5-Lipoxygenase products) play a major role in inflammatory responses
o The HETEs and EETs (Epoxy)genase products) tend to enhance Ca2+ release from
intracellular stores and to enhance cell proliferation
In smooth muscle, increase proliferation and migration
Involved in the formation of atherosclerotic plaque
In blood vessels, can be potent vasoconstrictors
Generally to enhance the release of calcium from intracellular stores
Na-H exchange
Cell proliferation
Cause vasodilation and angiogenesis
o Degradation of the eicosanoids terminates their activity
o Platelet-activating factor is a lipid mediator unrelated to arachidonic acid
PAF is an important lipid signaling molecule
A potent inducer of platelet aggregation and stimulates the chemotaxis and
degranulation of neutrophils thereby facilitating the release of /LTB4 and 5-HETE
Involved in allergic reactions
Enhances vascular permeability and the adhesion of neutrophil and platelets to
endothelial cells
Exerts its effects by binding to a specific receptor on the plasma membrane
Receptors that are catalytic
o Five classes of catalytic receptors
Receptor guanylyl cyclases
Catalyze the generation of cGMP from GTP
Receptor serine/threonine kinases
Phosphorylate serine and/or threonine residues on cellular proteins
Receptor tyrosine kinases (RTKs)
Phosphorylate tyrosine residues on themselves and other proteins
Tyrosine kinase-associated receptors
Interact with cytosolic tyrosine kinases
Receptor tyrosine phosphatases

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Cleave phosphate groups from tyrosine groups of cellular proteins

The receptor guanylyl cyclase transduces the activity of atrial natriuretic peptide, whereas a
soluble guanylyl cyclase transduces the activity of nitric oxide
Receptor guanylyl cyclase
Receptors for atrial natriuretic peptide (ANP)
ANP has two major effects
o Acts on vascular smooth muscle to dilate blood vessels
o It enhances Na+ excretion into urine which is termed natriuresis
The receptor for ANP is a membrane protein with a single membrane spanning
Binding of ANP to its receptor causes the conversion of GTP to cGMP and raises
intracellular levels of cGMP
Soluble guanylyl cyclase
Receptor for nitric oxide
A soluble guanylyl cyclase that contains a heme moiety that binds NO
NO plays an important role in the control of blood flow and blood pressure
NO also appears to play an important role in the destruction of invading
organisms by macrophages and neutrophils
NO also serves as a neurotransmitter and may play a role in learning and
NO synthase
o Used by vascular endothelial cells
Cleave arginine to citrulline plus NO in response to stimuli such
as Ach, Bradykynin, Substance P, Thrombin, Adenine nucleotides,
o Activaiton of NOS also requires the cofactors tetrahydrobiopterin and
NO stimulates its receptor, soluble guanylyl cyclase, which then converts GTP to
NO had long been exploited unwittingly to treat angina pectoris
Some catalytic receptors are serine/ threonine kinases
CaM-dependent protein kinases
Growth factor- (TGF- )
o Includes large group of cytokines such as
TGF- s
Anti-Mullerian hormone (AMH)
Bone morphogenic proteins
o Glycoproteins with a single membrane-spanning segment and intrinsic
serine/threonine-kinase activity
o Receptor I
Required for ligand binding and catalytic activity
Does not bind TGF-
It recognizes that the ligand has bound to the type II receptor and
forms a very stable ternary ligand/type I receptor/type II receptor

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Recruitment of complex results in phoosphorylation of the type I

receptor and activates the kinase and propagates the signal to
downstream effectors
o Receoptor II
Does not transmit the signal downstream
Receptor tyrosine kinases produce phosphotyrosine motifs recognized by SH domains of
downstream effectors
All receptor tyrosine kinases phosphorylate themselves in addition to other cellular
Ligands attached to RTK
Insulin and insulin-related growth factor type I (IGF-1)
o Tetrameric
contains a cysteine-rich region
functions in ligand binding
single-pass transmembrane protein with a cytoplasmic
tyrosine kinase domain
o and subunits are held together by disulfide bonds forming a

Most RTKs are single-pass transmembrane proteins that contain a single intracellular
kinase domain