Signal

transduc.on -2

Receptors in the Plasma Membrane

Most water-soluble signal molecules (eg. epinephrine, insulin, growth hormones)

bind to specific sites on receptor proteins in the plasma membrane

There are three main types of membrane receptors:

Ion channel-linked
receptors

Enzyme-linked
receptors

G-protein-linked
receptors
2

Ion Channel-linked Receptors

such as receptor for glutamate, serotonin and acetylcholine
act as a gate when the receptor change
shape
are involved in rapid synaptic signaling
between electrically excitable cells.
usually associated with a change in the
cells membrane potential.
This type of signaling is mediated by a
small number of neurotransmitters that
transiently open or close the ion channel
formed by the protein to which they bind.

Receptors with intrinsic (a) or associated (b)

enzymatic activity:

Receptor tyrosine kinases (RTKs) are

protein kinases that phosphorylates
tyrosine groups.
typically influence
cell proliferation and differentiation.

Receptors that Interact with

Cytoplasmic JAK Kinases: Just
Another
Kinase
such as cytokines receptor
Ligand molecule binds to the receptor
and causes a conformational change
within the receptor that leads to 4
activation of the JAK kinase.

Receptor tyrosine kinases (RTKs)

The most wide family of
RTKs: Ephineprins can act
simultaneously as ligands
and receptors

Extracellular region variable, with many different motifs

Usually cross membrane only once by a single transmembrane alpha-helix
Intracellular region contains conserved catalytic domains

General structure and ligand-induced activation of receptor tyrosine kinases

(RTKs)

Ligand promotes formation of RTK dimers, by different mechanisms:

Ligand itself is a dimer (PDGF)
One ligand binds both monomers (GH)
Dimerization allows trans-phosphorylation of catalytic domains, which
induces activation of catalytic (Y-kinase) activity
Activated TK domains phosphorylate each other and proteins nearby,
sometimes on multiple tyrosines
Y~P residues recruit other signaling proteins, generate multiple signals

How does dimerization activate RTKs?

Growth Factor Receptors (like many kinases) have sites in their T loops at which
phosphorylation activates

T-loop

Cat. loop

Y1162 occupies the

active site

Substrate Y
sits in active site

Y1162
flips out

Dimerization induces T-loop

phosphorylation in trans
Phosphorylation of Y (one or more)
in T-loop causes it to move out of
the way of the active site.
Once activated, each monomer can phosphorylate nearby Y residues in
the other, as well as in other proteins

Recruitment of signal-transduction proteins to the cell membrane by binding to

phosphotyrosine residues in activated receptors.

SH2 and PTB Domains Bind to Specific

Sequences Surrounding Phosphotyrosine
Residues

G ProteinCoupled Receptors (GPCRs)

Receptor is associated with a group

of heterotrimeric proteins known as
G proteins.
The G proteins can activate, or
inactivate, plasma membrane effector
proteins that function as ion channels
or enzymes
The heterotrimeric G proteins contains 3
subunits
The a subunit (Ga) binds GTP, and can
hydrolyze it to GDP + Pi.
The b subunit binds receptor, and forms a
stable complex with g subunit
a and g subunits have covalently attached lipid anchors that bind a G-protein to the9
plasma membrane cytosolic surface.

10

GPCR family

Schematic diagram of the general structure of G protein

coupled receptors.

11

Heterotrimeric G-protein

Figure 15-31 Molecular Biology of the Cell ( Garland Science 2008)

Switching mechanism for monomeric and trimeric G proteins.

13

14

Model of Ligand induced Activation of effector proteins associated with GPCRs

15

G ProteinCoupled Receptors (GPCR) family

Human genome encodes for
27 Ga
5 Gb
13 Gg

different Gbg display same function

Ga are divided in subclassis

16

GPCR family include

G ProteinCoupled Receptors
that activate or inhibit
Adenylyl Cyclase

G ProteinCoupled Receptors
that activate
cGMP phosphodiesterase

G ProteinCoupled Receptors
that activate
Phospholipase C
17

G ProteinCoupled Receptors That Activate or Inhibit Adenylyl Cyclase

Hormone-induced activation and inhibition of adenylyl cyclase by different GPCRs in adipose cells.
Ligand binding to Gs-coupled receptors causes activation of adenylyl cyclase, whereas ligand binding to Gicoupled receptors causes inhibitionof the enzyme. The Gbg subunit in both stimulatory and inhibitory G proteins is
identical; the Ga subunits and their corresponding receptors differ. Ligand-stimulated formation of active GaGTP
complexes occurs by the same mechanism in both Gs and Gi proteins. However, GsGTP and GiGTP interact
differently with adenylyl cyclase, so that one stimulates and the other inhibits its catalytic activity. [See A. G.
Gilman,1984, Cell 36:577.]
18

w Cholera toxin catalyzes covalent modification of Gsa in intestinal epithelial cells

ADP-ribose is transferred from NAD+ to an arginine residue at the GTPase

active site of Gsa.

ADP-ribosylation prevents GTP hydrolysis by Gsa .

The stimulatory G-protein is permanently activated.

w Pertussis toxin (whooping cough disease) catalyzes ADP-ribosylation at a

cysteine residue of the inhibitory Gia, making it incapable of exchanging GDP for
GTP in respiratory epithelial cells

The inhibitory pathway is blocked.

w ADP-ribosylation is a general mechanism by which activity of many proteins is

regulated, in eukaryotes (including mammals) as well as in prokaryotes.

19

ADP
ribosylation
O

O
C

protein
NH2

NH

+
N

O P O CH2 O
H
H
H
H
OH
OH
NH2
O
N

O
NH
O P O CH2 O
H
H
H
H
OH
OH
NH2
O

N
O P O CH2 N
O
O
H
H
H
H
+
NAD
OH
OH

(nicotinamide
adenine
dinucleotide)

(CH2)3

protein
(CH2)3

NH

Arg
C
residue
NH2

N
O P O CH2

NH2+

+
N
H

H
O
C

NH2+

N
O

OH

N
H

H
OH

ADP-ribosylated
protein
NH2

nicotinamide

20

G ProteinCoupled Receptors that activate adenylyl cyclase

1st messenger binds to GPCR receptor

(a hormone such as epinephrine)
Receptor activates Gs protein

Gs protein activates adenylyl cyclase

Schematic diagram of
mammalian adenylyl cyclases.

3D crystal structure
of GsGTP
complexed with
two fragments
encompassing the
21 of
catalytic domain
adenylyl cyclase

Adenylyl cyclase, activated by GPCRs

in response to an extracellular signal,
converts a molecule of ATP into cyclic
AMP (cAMP), a second messenger.
cAMP then attaches to and activates
cAMP-dependent protein kinases that
can phosphorylate and activate enzymes
used in cellular responses.
The phosphodiesterase enzymes
terminate the second messenger
cAMP.
Caffeine and theophylline, the active
ingredients of coffee and tea
respectively, inhibit phosphodiesterase
activity
22

cAMP-induced activation of protein kinase A (PKA). [movie]

At low concentrations of cyclic AMP (cAMP), the PKA is an inactive tetramer. Binding of cAMP to the regulatory
(R) subunits causes a conformational change in these subunits that permits release of the active, monomeric
catalytic (C) subunits. (b) Cyclic AMP is a derivative of adenosine monophosphate. This intracellular signaling
molecule, whose concentration rises in response to various extracellular signals, can modulate the activity of
many proteins.
Figure 15-35 Molecular Biology of the Cell ( Garland Science 2008)

Molecular basis of the allosteric regulation of the regulative subunits of pKA.

pKA contains two regolative

subunits interconnected by a
dimerization domain
Each R subunit contains two sites
for cAMP (CNB-A and CNB-B).
Binding of cAMP to each R
subunit occurs in a cooperative
manner: binding of cAMP to CNBB decrease the kd of cAMP for
CNB-A.
When cAMP is bound to CNB-A
the R subunit undergoes to
conformational changes that
displaces the associated Csubunit

24

Figure 15-36 (part 1 of 2) Molecular Biology of the Cell ( Garland Science 2008)

Figure 15-36 (part 2 of 2) Molecular Biology of the Cell ( Garland Science 2008)

Figure 15-36 Molecular Biology of the Cell ( Garland Science 2008)

Amplification of an external signal downstream from a cell-surface receptor.

The cAMP system rapidly amplifies the response

capacity of cells: here, one first messenger led
to the formation of one million product molecules.

28

Cells can respond via the cAMP pathways using a diversity of cAMP-dependent
enzymes, channels, organelles, contractile filaments, ion pumps, and changes in
gene expression.
29

G ProteinCoupled Receptors that activate Phospholipase C

1st messenger binds to GPCR receptor

Receptor activates phospholipase C

Phospholipase C produces the secondary

messengers diacylglycerol (DAG), and
inositol triphosphate (IP3)

30

Synthesis of DAG and IP3 from membrane-bound phosphatidylinositol (PI).

31

Figure 15-39 Molecular Biology of the Cell ( Garland Science 2008)

Signal transduction pathway downstream G ProteinCoupled Receptors

Activating Phospholipase C

Inositol 1,4,5-Trisphosphate (IP3) Triggers Release of Ca2+ from the Endoplasmic Reticulum 33
Diacylglycerol activates PKC

Ca2+ as second messenger in signal trasduction

Figure 15-41b Molecular Biology of the Cell ( Garland Science 2008)

Oscillation of [Ca2+] in the citosol of a liver cell affects cell response

The frequency of
Ca2+ spikes reflects
the potency of a
signal
In the pituitary cells
to each Ca2+ spike
corresponds a fast
hormone secretion.
In other cells each
specific frequency of
Ca2+ spikes
induces a specific
pattern of genes
activation
Figure 15-42 Molecular Biology of the Cell ( Garland Science 2008)

Calmodulin, a Ca2+ binding protein that regulate the activity of several kinases and
other enzymes

Ca2+ can make calmodulin able

to bind to a target protein

Two or more Ca2+ have to bind calmodulin for its activation

Figure 15-43 Molecular Biology of the Cell ( Garland Science 2008)

Activation of the Tubby transcription

factor following ligand binding to
receptors coupled to Go or
Gq

Activation of gene expression

following ligand binding to Gs
proteincoupled
receptors

37

G ProteinCoupled Receptors that activate cGMP phosphodiesterase: Rhodopsin

Photon isomerize 11-cis-retinal molecule

Large Rhodopsin molecule change conformation and activate the associated
G-proteins, trasducins
Trasducins activate cGMP phosphodiesterases
cGMP phosphodiesterases hydrolyzes cGMPs
cGMPs reduction closes cGMP-dependent Na channel that prevent ionsdependent membrane hyperpolarization

38

Alteration of the membrane potential and reduction in neurotrasmitter release

39