Download Signal Transduction

 Organisms can function normally only if:
 All the cells of their different organs communicating
effectively with their surroundings.
 Once a cell picks up a hormonal or sensory signal, it
must transmit this information from the surface to the
interior parts of the cell—
Example, to the nucleus.
 This occurs via signal transduction pathways that are:
very specific, both in their activation and in their
downstream actions.
 Thus, the various organs in the body respond in an
appropriate manner and only to relevant signals.
Signal Transduction
 The transmission of molecular
signals from a cell's exterior to its
interior.
Molecular signals transmitted
between cells by:
1. Hormones
2. Other chemical factors
3. Sensory signals received from
the environment, in form of:
Light, Taste, Sound, Smell, and
Touch.
Upon Environment changes:
Monad: responds directly
Multicellular organisms:
Respond with signal through a system
of intercellular or intracellular
communication，and consequently
regulate functions of organisms.
Signaling molecule
Receptor of target cell
Intracellular molecule
biological effect
Signal
transduction
Signaling molecules
• Released by signal-producing cells,
• Transfer biological signals to their
target cells to initiate specific cellular
responses.
Two types..
• Extracellular molecules
• Intracellular molecules
1. Extracellular molecules
protein & peptides: Hormone, cytokine
AA & its derivatives: Gly, Glu, adrenaline, thyroxine
Steroid: Sex Hormone, glucocorticosteroid
Fatty acid derivatives: prostaglandin
(1) Paracrine signaling
(local chemical mediators)
• Secreted by common cells.
• Reach neighboring target cells by
passive diffusion.
• Time of action is short.
• Such as GF
(2) Endocrine signal
•
•
•
•
Secreted by endocrine cells.
Reach target cells by blood circulation.
Time of action is long.
Such as insulin, thyroxine, adrenalin
(3) Synaptic signal
(neurotransmitters)
• Secreted by neuronal cells.
• Reach another neuron by synaptic
gap.
• Time of action is short.
• Such as Acetylcholine (Ach),
noradrenaline
(4) Gaseous signal
• Simple structure, half life is short
and active in chemistry .
• Such as NO, CO.
GAS MOLECULE
(5) Autocrine signal
• Act back to their own cells.
• Such as GF, cytokine, interferon,
interleukin.
2. Intracellular molecule
• Ca2+
ions
• DG, ceramide
• IP3
lipid derivatives
carbohydrate
derivatives
• cAMP cGMP
nucleotides
• Ras, JAK, Raf
proteins
Second messenger:
Small molecules synthesized in cells
in response to an external signal,
responsible for intracellular signal
transduction.
Such as Ca2+, DG, Cer, IP3, cAMP,
cGMP
Third messengers:
The molecules which transmit message
from outside to inside of nucleous or
from inside to outside of nucleous, also
called DNA binding protein.
Proteins and peptides:
Effect by
membrane
receptors
Hormones, cytokines
Amino acid derivatives:
Catecholamines
Fatty acid derivatives:
Extracellular
molecules
Prostaglandins
Effect by
intracellular
receptors
Signal
molecules
Intracellular
molecules
Steroid hormones,
Thyroxine, VD3
cAMP, cGMP, IP3, DG, Ca2+
2. Receptors
Receptors
Specific membrane proteins, able to
recognize and bind to corresponding
ligand molecules, become activated, and
transduce signal to next signaling
molecules.
Glycoprotein or Lipoprotein
ligand
A small molecule that binds
specifically to a larger one;
Example:
A hormone is the ligand for its specific
protein receptor.
• Membrane receptors
membrane
Glycoprotein
• Intracellular receptors
Cytosol or nuclei
DNA binding protein
1. membrane receptors
(1) Ligand-gate ion channels type (cyclic
receptor)
ligand→receptor→ion channel open or
close
Properties of binding of H and R
• highly specificity
• highly affinity
• saturation
• reversible binding
• special function model
Control of receptor activity
• Phosphorylation or
dephosphorylation of R
• Phospholipid of membrane
• Enzyme catalyzed hydrolysis
• G protein regulation
Function of receptor
(1) Recognize the special ligand
(2) Binding to special ligand
(3) Signal transduction
biological effect
Pathway of Signal Transduction
serine (Ser)
threonine (Thr)
H
H
H3N+
C
COO
H3N+
C
COO
CH2
CH OH
OH
CH3
Many enzymes are regulated by covalent
attachment of phosphate, in ester linkage, to the
side-chain hydroxyl group of a particular amino acid
residue (serine, threonine, or tyrosine).
O
Protein Kinase
OH + ATP
Protein
Protein
O
P
O + ADP
O
Pi
H2O
Protein Phosphatase
 A protein kinase transfers the terminal phosphate of
ATP to a hydroxyl group on a protein.
 A protein phosphatase catalyzes removal of the Pi by
hydrolysis.
Phosphorylation may directly alter activity of an
enzyme, e.g., by promoting a conformational change.
Alternatively, altered activity may result from binding
another protein that specifically recognizes a
phosphorylated domain.
 E.g., 14-3-3 proteins bind to domains that include
phosphorylated Ser or Thr in the sequence
RXXX[pS/pT]XP, where X can be different amino acids.
 Binding to 14-3-3 is a mechanism by which some
proteins (e.g., transcription factors) may be retained
in the cytosol, & prevented from entering the
nucleus.
O
Protein Kinase
OH + ATP
Protein
Protein
O
P
O + ADP
O
Pi
H2O
Protein Phosphatase
Protein kinases and phosphatases are themselves
regulated by complex signal cascades. For example:
 Some protein kinases are activated by Ca++calmodulin.
 Protein Kinase A is activated by cyclic-AMP (cAMP).
Adenylate Cyclase (Adenylyl
Cyclase) catalyzes:
ATP  cAMP + PPi
Binding of certain hormones
(e.g., epinephrine) to the outer
surface of a cell activates
Adenylate Cyclase to form cAMP
within the cell.
Cyclic AMP is thus considered to
be a second messenger.
NH2
cAMP
N
N
N
N
H2
5' C 4'
O
O
O
H
H 3'
O
P
O-
H
1'
2' H
OH
Phosphodiesterase enzymes
catalyze:
cAMP + H2O  AMP
N
N
The phosphodiesterase that
cleaves cAMP is activated by
phosphorylation catalyzed by
Protein Kinase A.
N
N
H2
5' C 4'
O
Thus cAMP stimulates its own
degradation, leading to rapid
turnoff of a cAMP signal.
NH2
cAMP
O
O
H
H 3'
P
O
O-
H
1'
2' H
OH
Protein Kinase A (cAMP-Dependent Protein Kinase)
transfers Pi from ATP to OH of a Ser or Thr in a
particular 5-amino acid sequence.
Protein Kinase A in the resting state is a complex of:
• 2 catalytic subunits (C)
• 2 regulatory subunits (R).
R2C2
R2C2
Each regulatory subunit (R) of Protein Kinase A
contains a pseudosubstrate sequence, like the
substrate domain of a target protein but with Ala
substituting for the Ser/Thr.
The pseudosubstrate domain of (R), which lacks a
hydroxyl that can be phosphorylated, binds to the
active site of (C), blocking its activity.
R2C2 + 4 cAMP  R2cAMP4 + 2 C
When each (R) binds 2 cAMP, a conformational
change causes (R) to release (C).
The catalytic subunits can then catalyze
phosphorylation of Ser or Thr on target proteins.
PKIs, Protein Kinase Inhibitors, modulate activity of
the catalytic subunits (C).
Pathway of G protein linked receptor
H
R
G protein
Es
secondary messeger
Protein kinase
Phophorylation of Es or functional protein
Biological effect
G Protein Signal Cascade
Most signal molecules targeted to a cell bind at the
cell surface to receptors embedded in the plasma
membrane.
Only signal molecules able to
cross the plasma membrane
(e.g., steroid hormones) interact
with intracellular receptors.
A large family of cell surface
receptors have a common
structural motif, 7
transmembrane a-helices.
Rhodopsin was the first of these to
have its 7-helix structure confirmed by Rhodopsin
X-ray crystallography.
PDB 1F88
 Rhodopsin is unique.
Lysozyme
insert
It senses light, via a bound
chromophore, retinal.
 Most 7-helix receptors have
domains facing the
extracellular side of the
plasma membrane that
recognize & bind signal
molecules (ligands).
E.g., the b-adrenergic
receptor
is activated by
epinephrine & norepinephrine.
ligand

b-Adrenergic
Receptor
PDB 2RH1
Crystallization of this receptor was aided by genetically engineering
insertion of the soluble enzyme lysozyme into a cytosolic loop between
transmembrane a-helices.
The signal is usually
passed from a 7-helix
receptor to an
intracellular G-protein.
 Seven-helix
receptors are thus
called GPCR, or
G-Protein-Coupled
Receptors.
 Approx. 800 different
GPCRs are encoded
in the human
genome.
G-protein-Coupled Receptors may dimerize or
form oligomeric complexes within the membrane.
Ligand binding may promote oligomerization, which
may in turn affect activity of the receptor.
Various GPCR-interacting proteins (GIPs)
modulate receptor function. Effects of GIPs may
include:
 altered ligand affinity
 receptor dimerization or oligomerization
 control of receptor localization, including transfer to or
removal from the plasma membrane
 promoting close association with other signal proteins
 G-proteins are heterotrimeric, with 3 subunits a,
b, g.
 A G-protein that activates cyclic-AMP formation
within a cell is called a stimulatory G-protein,
designated Gs with alpha subunit Gsa.
 Gs is activated, e.g., by receptors for the
hormones epinephrine and glucagon.
The b-adrenergic receptor is the GPCR for
epinephrine.
hormone
signal
outside
GPCR
The a subunit of
a G-protein (Ga)
binds GTP, &
can hydrolyze it
to GDP + Pi.
plasma
membrane
agga
AC
GDP bbGTP
GTP
GDP
cytosol
ATP cAMP + PPi
a & g subunits have covalently attached lipid anchors
that bind a G-protein to the plasma membrane
cytosolic surface.
Adenylate Cyclase (AC) is a transmembrane protein,
with cytosolic domains forming the catalytic site.
hormone
signal
outside
GPCR
plasma
membrane
agga
AC
GDP bbGTP
GTP
GDP
cytosol
ATP cAMP + PPi
The sequence of events by which a hormone activates
cAMP signaling:
1. Initially Ga has bound GDP, and a,b, & g subunits
are complexed together.
Gb,g, the complex of b & g subunits, inhibits Ga.
hormone
signal
outside
GPCR
plasma
membrane
agga
AC
GDP bbGTP
GTP
GDP
cytosol
ATP cAMP + PPi
2. Hormone binding, usually to an extracellular domain
of a 7-helix receptor (GPCR), causes a conformational
change in the receptor that is transmitted to a G-protein
on the cytosolic side of the membrane.
The nucleotide-binding site on Ga becomes more accessible
to the cytosol, where [GTP] > [GDP].
Ga releases GDP & binds GTP (GDP-GTP exchange).
hormone
signal
outside
GPCR
plasma
membrane
agga
AC
GDP bbGTP
GTP
GDP
cytosol
ATP cAMP + PPi
3. Substitution of GTP for GDP causes another
conformational change in Ga.
Ga-GTP dissociates from the inhibitory bg complex & can
now bind to and activate Adenylate Cyclase.
hormone
signal
outside
GPCR
plasma
membrane
agga
AC
GDP bbGTP
GTP
GDP
cytosol
ATP cAMP + PPi
4. Adenylate Cyclase, activated by the stimulatory
Ga-GTP, catalyzes synthesis of cAMP.
5. Protein Kinase A (cAMP Dependent Protein Kinase)
catalyzes transfer of phosphate from ATP to serine or
threonine residues of various cellular proteins, altering
their activity.
Turn off of the signal:
1. Ga hydrolyzes GTP to GDP + Pi. (GTPase).
The presence of GDP on Ga causes it to rebind to
the inhibitory bg complex.
Adenylate Cyclase is no longer activated.
2. Phosphodiesterases catalyze hydrolysis of
cAMP  AMP.
3. Protein Phosphatase catalyzes removal by
hydrolysis
of phosphates that were attached to
proteins via Protein
Kinase A.
Signal amplification is an important feature of signal
cascades:
 One hormone molecule can lead to formation of
many cAMP molecules.
 Each catalytic subunit of Protein Kinase A catalyzes
phosphorylation of many proteins during the lifetime of the cAMP.
 Different isoforms of Ga have different signal roles. E.g.:
• The stimulatory Gsa, when it binds GTP, activates
Adenylate cyclase.
• An inhibitory Gia, when it binds GTP, inhibits
Adenylate cyclase.
Different effectors & their receptors induce Gia to
exchange GDP for GTP than those that activate Gsa.
 The complex of Gb,g that is released when Ga binds GTP is
itself an effector that binds to and activates or inhibits
several other proteins.
E.g., Gb,g inhibits one of several isoforms of Adenylate
Cyclase, contributing to rapid signal turnoff in cells that
express that enzyme.
 Cholera toxin catalyzes covalent modification of Gsa.
• 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.
 Pertussis toxin (whooping cough disease) catalyzes ADPribosylation at a cysteine residue of the inhibitory Gia,
making it incapable of exchanging GDP for GTP.
• The inhibitory pathway is blocked.
 ADP-ribosylation is a general mechanism by which activity
of many proteins is regulated, in eukaryotes (including
mammals) as well as in prokaryotes.