CHAPTER 15

Cell Signaling Pathways

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.1 | The Basic Elements of Cell Signaling Systems

Types of intercellular signaling: autocrine (left), paracrine (middle), and endocrine (right)

Extracellular messenger molecules transmit messages between cells.

In autocrine signaling, the cell has receptors on its surface that respond to
the messenger.

During paracrine signaling, messenger molecules travel short distances

through extracellular space.

During endocrine signaling, messenger molecules reach their target cells

through the bloodstream.
© 2017 John Wiley & Sons, Inc. All rights reserved.
 15.1 | The Basic Elements of Cell Signaling Systems

Receptors on or in target cells

receive the message.

Some cell surface receptors

generate an intracellular
second messenger through
an enzyme called an effector.

Second messengers are small

substances that activate (or
inactivate) specific proteins. An overview of the
major signaling
pathways by which
Other surface receptors recruit extracellular
proteins to their intracellular messenger
domains at the plasma molecules can
membrane. elicit intracellular
responses
© 2017 John Wiley & Sons, Inc. All rights reserved.
 15.1 | The Basic Elements of Cell Signaling Systems

Signaling pathways consist of a

series of proteins.

Each protein in a pathway alters the

conformation of the next protein.

Protein conformation is usually

altered by phosphorylation.

Kinases add phosphate groups

while phosphatases remove them.

Target proteins ultimately receive a

message to alter cell activity.
Signal transduction pathway consist
This overall process is called signal of kinases and phosphatases that
change the conformations and
transduction.
activities of target proteins.

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.1 | The Basic Elements of Cell Signaling Systems

Protein phosphorylation can change

protein behavior in different ways.

It can activate or inactivate an

enzyme.

It can increase or decrease protein-

protein interactions.

It can change the subcellular location

of the protein .

It can trigger protein degradation.

Phosphorylation patterns differ

between cell types (e.g. two different
types of breast cancer cells).

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.2 | A Survey of Extracellular Messengers and Their Receptors

Extracellular messengers include:

Small molecules such as amino acids and their derivatives (e.g.
acetylcholine, epinephrine, dopamine).
Gases such as NO and CO.
Steroids, derived from cholesterol, regulate sexual differentiation,
pregnancy, carbohydrate metabolism.
Eicosanoids, derived from arachidonic acid, regulate processes
including pain, inflammation, blood pressure, and blood clotting.
Various peptides and proteins, including transmembrane proteins
on the surface of an interacting cell, extracellular matrix interacting
proteins, and secreted proteins.

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.2 | A Survey of Extracellular Messengers and Their Receptors

Receptor types include:

G-protein coupled receptors (GPCRs), which contain seven
transmembrane α helices and activate GTP-binding proteins.

Receptor protein-tyrosine kinases (RTKs), which dimerize and

activate their cytoplasmic protein-kinase domain to phosphorylate
specific tyrosine residues of cytoplasmic substrate proteins.
Ligand gated channels, which conduct a flow of ions across the
plasma membrane to change its potential.
Steroid hormone receptors, which function as ligand-regulated
transcription factors.
Specific receptors such as B-and T-cell receptors which are
involved in the response to foreign antigens.

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.3 | Signal Transduction by G Protein-Coupled Receptors

Transducing
signals through
a heterotrimeric
G protein.

A model depicting
the activation of the
GPCR rhodopsin

G protein-coupled receptors (GPCRs) constitute the single largest

superfamily of proteins encoded by animal genomes.

GPCRs have seven a-helical transmembrane domains and interact with

G proteins.

Natural ligands: hormones (both plant and animal), neurotransmitters,

opium derivatives, chemoattractants (odorants, tastants, and photons).

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.3 | Signal Transduction by G Protein-Coupled Receptors

Ligands activate receptors that stimulate effectors to give rise to a

physiological response

G protein-coupled receptors normally have their amino-terminus present on

the outside of the cell, the seven a helices that traverse the plasma
membrane connected by loops of varying length, and the carboxyl-terminus
present on the inside of the cell.

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.3 | Signal Transduction by G Protein-Coupled Receptors
Receptors

Ligand binding to the receptor

extracellular domain changes the
conformation of its intracellular
domain.

The receptor’s affinity for G proteins

increases, and the receptor binds
the trimeric G protein.

A GDP is exchanged for GTP on

the Ga subunit, thus activating it
and promoting association with the
effector. Receptor-mediated
activation (or
One ligand-bound receptor can inhibition) of
activate many G proteins. effectors by means
of heterotrimeric G
proteins
© 2017 John Wiley & Sons, Inc. All rights reserved.
15.3 | Signal Transduction by G Protein-Coupled Receptors
Receptors

Termination of the response can

occur through multiple processes.

Desensitization – by blocking active

receptors from turning on additional
G proteins.

G protein-coupled receptor kinase

(GRK) activates a GPCR via
phosphorylation.

Proteins called arrestins compete

with G proteins to bind GPCRs.
Receptor-mediated
Termination of the response is activation (or
accelerated by regulators of G inhibition) of
protein signaling (RGSs). effectors by means
of heterotrimeric G
proteins
© 2017 John Wiley & Sons, Inc. All rights reserved.
15.3 | Signal Transduction by G Protein-Coupled Receptors
G proteins

Ga subunits can turn themselves off by hydrolysis of GTP to GDP and Pi,
which causes a conformational change and a decreased affinity for the
effector and an increased affinity for the Gbg subunit.

Heterotrimeric G proteins function as molecular timers, and while active,

Ga subunits can turn on downstream effectors.

Heterotrimeric G proteins come in four flavors, Gs, Gq, Gi, and G12/13 based
on the Ga subunits and the effectors to which they couple.

Gs family members couple receptors to adenylyl cyclase; Gq family

members contain Ga subunits that activate PLCβ; Gi subunits function by
inhibiting adenylyl cyclase; and G12/13 members are less well characterized.

Following its dissociation from the Gα subunit, the Gbg complex also has a
signaling function and it can couple to a number of different types of
effectors, including PLCβ, K+ and Ca2+ ion channels, and adenylyl cyclase.

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.3 | Signal Transduction by G Protein-Coupled Receptors
Bacterial Toxins

Bacterial Toxins, such as cholera toxin and pertussis virulence factors, target
GPCRs and G proteins.

b Adrenergic Receptors stimulate Gas to activate adenylate cyclase. Gas is a

target of cholera toxin, as the GTPase activity is inhibited.

Pathway is locked in stimulatory state, as adenylyl cyclase molecules remain

in an activated mode, churning out cAMP, which causes the epithelial cells to
secrete large volumes of fluid into the intestinal lumen.

a Adrenergic Receptors stimulate Gai to inhibit adenylate cyclase. Gai is a

target of pertussis toxin.

The toxin also inactivates Ga subunits, thereby interfering with the signaling
pathway that leads the host to mount a defensive response against the
bacterial infection.

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.4 | Second Messengers

Localized formation of cAMP

in response to the addition of
an extracellular messenger
molecule. Photographs of a
sensory nerve cell (Aplysia)

Cyclic AMP is a second messenger that diffuses to other sites in the cell.

The synthesis of cyclic AMP follows the binding of a first messenger, a

hormone or other ligand, to a receptor at the outer surface of the cell.

Second messengers enable cells to mount a large-scale, coordinated

response following stimulation by a single extracellular ligand.

Other second messengers include Ca2+, phosphoinositides, inositol

trisphosphate, diacylglycerol, cGMP, and nitric oxide.

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.4 | Second Messengers
Phosphatidylinositol-Derived Second Messengers

Structure of a generalized Interaction: PLC enzyme Chemoattractant generates

phospholipid and a phosphoinositide PIP3 at the cell leading edge

Some phospholipids of cell membranes are converted into second

messengers by phospholipases (lipid-splitting), phospholipid kinases (lipid-
phosphorylating), and phospholipid phosphatases (lipid-dephosphorylating).

Phosphorylated phosphoinositides, derived from phosphatidylinositol, form

lipid-binding domains called PH domains.

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.4 | Second Messengers
Phosphatidylinositol-Derived Second Messengers

The generation of second

messengers as a result of
ligand-induced breakdown
of phosphoinositides (PI) in
the lipid bilayer.

Phosphatidylinositol-specific phospholipase C-b produces second messengers

derived from phosphatidylinositol-inositol triphosphate (IP3) and diacylglycerol
(DAG).

DAG activates protein kinase C, which phosphorylates serine and threonine

residues on target proteins.

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.6 | Regulation of Blood Glucose Levels

Different stimuli acting on the same

target cell may induce the same
response.

Glucagon and epinephrine bind to

different receptors on the same cell.

Both hormones stimulate glycogen

breakdown and inhibit its synthesis.

cAMP is activated by the G protein

of both hormone receptors.

Responses are amplified by signal

cascades. The reactions that lead to glucose
storage or mobilization.

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.6 | Regulation of Blood Glucose Levels
Glucose Mobilization: An Example of a Response Induced by cAMP

cAMP is synthesized by the

effector enzyme adenylyl cyclase.

Adenylyl cyclase is an integral

membrane protein whose catalytic
domain resides at the inner surface
of the plasma membrane.

cAMP evokes a reaction cascade

that leads to glucose mobilization.

cAMP breakdown is accomplished

by a phosphodiesterase. Formation of cyclic AMP from ATP
is catalyzed by adenylyl cyclase,
an integral membrane protein that
consists of two parts

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.6 | Regulation of Blood Glucose Levels
Glucose Mobilization: An Example of a Response Induced by cAMP

The first step in the

reaction cascade occurs
as the hormone binds to
its receptor, activating a
Gas subunit, which
activates adenylyl cyclase.

The activated enzyme

catalyzes the formation of
cAMP molecules, which
diffuse into the cytoplasm
where they bind a cAMP- The response by a
dependent protein kinase liver cell to glucagon
(protein kinase A, PKA). or epinephrine

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.6 | Regulation of Blood Glucose Levels
Signal Amplification

Binding of a single hormone molecule can activate a number of G

proteins, each of which can activate an adenylyl cyclase effector, each of
which can produce a large number of cAMP messengers quickly.

The production of a second messenger provides a mechanism to greatly

amplify the signal generated from the original message.

cAMP molecules activate PKA, which phosphorylates a large number of

phosphorylase kinase molecules, which in turn phosphorylate an even
larger number of glycogen phosphorylase molecules, which in turn can
catalyze the formation of a much larger number of glucose phosphates.

What begins as a barely perceptible stimulus at the cell surface is rapidly

transformed into a major mobilization of glucose within the cell.

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.8 | Protein-Tyrosine Phosphorylation as a Mechanism for
Signal Transduction

Protein-tyrosine kinases, which phosphorylate tyrosine residues on

target proteins, can be divided in two groups: receptor protein‐tyrosine
kinases (RTKs), integral membrane proteins with a single transmembrane
helix and an extracellular ligand binding domain, and non‐receptor (or
cytoplasmic) protein‐tyrosine kinases.

The human genome encodes nearly 60 RTKs and 32 non-receptor TKs.

RTKs are activated directly by extracellular growth and differentiation

factors such as epidermal growth factor (EGF) and platelet-derived
growth factor (PDGF) or by metabolic regulators such as insulin.

Non-receptor protein-tyrosine kinases are regulated indirectly by

extracellular signals, and they control processes as diverse as the
immune response, cell adhesion, and neuronal cell migration.

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.8 | Protein-Tyrosine Phosphorylation as a Mechanism for
Signal Transduction
Receptor Dimerization

Ligand binding results in dimerization of

the extracellular ligand-binding domains
of a pair of receptors.

Two mechanisms for receptor

dimerization: ligand-mediated
dimerization (e.g. PDGF) and receptor-
mediated dimerization (e.g., EGF).

For most RTKs, dimerization brings two

kinase domains in close contact for trans-
autophosphorylation; the protein kinase
activity of one receptor phosphorylates
tyrosine residues in the cytoplasmic
domain of the other receptor. Steps in the activation of a receptor
protein-tyrosine kinase (RTK)

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.8 | Protein-Tyrosine Phosphorylation as a Mechanism for
Signal Transduction
Protein Kinase Activation

Autophosphorylation sites can regulate the receptor’s kinase activity

or serve as binding sites for cytoplasmic signaling molecules.

Kinase activity is usually controlled by autophosphorylation on

tyrosine residues that are present in the activation loop of the kinase
domain.

Following its phosphorylation, the activation loop is stabilized in a

position away from the substrate-binding site, resulting in activation
of the kinase domain.

The receptor subunits then phosphorylate each other on tyrosine

residues that are present in regions adjacent to the kinase domain;
these sites act as binding sites for cellular signaling proteins.

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.8 | Protein-Tyrosine Phosphorylation as a Mechanism for
Signal Transduction
Phosphotyrosine-Dependent Protein-Protein Interactions

Phosphorylated tyrosines bind

effector proteins that have either a
Src-homology 2 (SH2) domain or a
phosphotyrosine-binding (PTB)
domain.

SH2 domains are composed of

roughly 100 amino acids and contain
a conserved binding-pocket that
accommodates a phosphorylated
tyrosine residue.

PTB domains can bind to The interaction between SH2

phosphorylated tyrosine residues domain and a peptide
and are usually present as part of an contain a phosphotyrosine
Asn-Pro-X-Tyr motif.

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.8 | Protein-Tyrosine Phosphorylation as a Mechanism for
Signal Transduction
Activation of Downstream Signaling Pathways

SH2 and PTB domain proteins

include:

1) Adaptor proteins that bind A diversity of

other proteins. signaling
proteins. Cells
2) Docking proteins that supply contain numerous
receptors with other tyrosine proteins with SH2
phosphorylation sites. or PTB domains
that bind to
3) Signaling enzymes (kinases) phosphorylated
that lead to changes in cell. tyrosine residues

4) Transcription factors

© 2017 John Wiley & Sons, Inc. All rights reserved.

15.8 | Protein-Tyrosine Phosphorylation as a Mechanism for
Signal Transduction
Ending the Response

Signal transduction by RTKs is usually terminated by internalization

of the receptor, primarily through clathrin-mediated endocytosis.

Some RTKs may bind to the clathrin adaptor protein AP-2, or may be
targeted by ubiquitination by ubiquitin ligases through SH2 domains
or adaptor proteins.

Internalized RTKs can have several alternate fates; they can be

degraded in lysosomes, returned to the plasma membrane, or
become part of endosomal signaling complexes and engage in
continued intracellular signaling.

Two important downstream signaling pathways: 1)the Ras-MAP

kinase pathway is probably the best characterized signaling cascade,
and 2) the insulin receptor-mediate cascade.

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.15 | Apoptosis (Programmed Cell Death)

EM: normal (left) and apoptotic Morphological differences apoptosis

cv
(right) T-cell hybridoma
cv
(upper right) and necroptosis (center)

Apoptosis is an ordered process involving cell shrinkage, loss of adhesion to

other cells, dissection of chromatin, and engulfment by phagocytosis.

Apoptotic changes are activated by proteolytic enzymes, caspases, which

target:
Protein kinases, some of which cause detachment of cells.
Lamins, which line the nuclear envelope.
Proteins of the cytoskeleton
Caspase activated DNase (CAD)
© 2017 John Wiley & Sons, Inc. All rights reserved.
 15.15 | Apoptosis (Programmed Cell Death)

Apoptosis carves out the

structure of the
mammalian
cv digits:
tracking cyan fluorescent
macrophages

Apoptosis is needed during embryonic development to form structure, organs

and tissues (e.g. spaces between digits, pruning unneeded neurons).

Apoptosis is also active in the adult, where about 1010-1011 cells die per day.

Reduced or elevated apoptosis is linked to several human diseases:

Cancer; Parkinson’s, Alzheimer’s, and Huntington’s diseases; Diabetes type I

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.15 | Apoptosis (Programmed Cell Death)
The Extrinsic Pathway of Apoptosis

The extrinsic pathway of apoptosis is

initiated by external stimuli.

As an example, the death ligand tumor

necrosis factor (TNF) is detected by a
TNF cell surface receptor.

Bound TNF receptors recruit

“procaspases” to the intracellular
domain of the receptor.

Procaspases convert other

procaspases to caspases.

Caspases activate executioner

caspases, leading to apoptosis. The extrinsic (receptor-mediated)
pathway ofcvapoptosis

© 2017 John Wiley & Sons, Inc. All rights reserved.

 15.15 | Apoptosis (Programmed Cell Death)
The Intrinsic Pathway of Apoptosis

The intrinsic pathway of apoptosis is

initiated by intracellular stimuli, e.g. DNA
damage.

Proapoptotic proteins stimulate

mitochondria to leak proteins, mostly
cytochrome c.

Once in the cytosol, cytochrome c forms

part of a multiprotein complex called the
apoptosome, that also includes several
molecules of procaspase-9.

Release of apoptotic mitochondrial

proteins irreversibly commits the cell to
apoptosis. The intrinsic (mitochondria-mediated)
pathway of apoptosis
© 2017 John Wiley & Sons, Inc. All rights reserved.
 15.15 | Apoptosis (Programmed Cell Death)
The Intrinsic Pathway of Apoptosis

Antiapoptotic proteins promote survival.

Cell fate depends on the balance
between pro- and anti-apoptotic signals.

Apoptotic cell death occurs without

spilling cellular contents to prevent
inflammation, compared to necroptosis.

During apoptosis, a phospholipid

“scramblase” moves phosphatidylserine
to the outer leaflet of the plasma
membrane, and is recognized as an “eat
me” signal by specialized macrophages.
Clearance of apoptotic cells is
accompanied cv
by phagocytosis
Apoptotic cells are subsequently cleared
by phagocytosis.

© 2017 John Wiley & Sons, Inc. All rights reserved.