Transcript G-protein-mediated pathway

Signal Transduction and
the Related Disorders
Department of Pathophysiology
Shanghai Jiao-Tong University School of Medicine
CHAPTER 1
General Introduction of Cell Signal
Transduction
Concept of Cell Signaling
The process in which cells sense the extracellular stimuli
through membranous or intracellular receptors, transduce the
signals via intracellular molecules , and thus regulate the
biological function of the cells
Signal molecules
Physical signals
Light, electronic, mechanic, UV, heat, volume or osmotic, etc
Chemical signals
Hormones, neurotransmitters, Growthe factors, cytokines,
odor molecules, ATP, active oxygen, drugs, toxins, etc
Modes for the function of endogenous signals
Endocrine: Act on a far away organ via blood circulation
Paracrine: Act on a nearby target
Autocrine: Act on itself after secreted
Synaptic: Presynaptic to postsynaptic,
Endocrine
Autocrine
Paracrine
Synaptic
The primary pathways of cell signalling
G-protein-mediated pathway
Adenylate cyclase mediated pathway
Phospholipase mediated pathway
Small G-protein-mediated pathway
Non-G-protein-mediated pathway
Receptor tyrosine kinase mediated pathway
Receptor serine/threonine kinase mediated pathway
Receptor guanilate cyclase mediated pathway
Intracellular (unclear) receptor mediated pathway
G-protein-mediated pathway
G-proteins, coupled with members of the seven transmembrane
domain of the receptor superfamily, are regulatory proteins that
act as molecular switches. They control a wide range of
biological processes
Classification of G-protein
High moleular weight G-protein
(trimeric GTP-binding regulatory protein）
Low moleular weight G-protein
Ras
Regulation of G-Protein Activity
G protein-coupled receptors exhibit a common structural motif consisting of
seven membrane spanning regions. Receptor occupation promotes interaction
between the receptor and the G protein on the interior surface of the
membrane. This induces an exchange of GDP for GTP on the G protein 
subunit and dissociation of the  subunit from the  heterodimer. Depending
on its isoform, the GTP- subunit complex mediates intracellular signaling
either indirectly by acting on effector molecules such as adenylyl cyclase (AC)
or phospholipase C (PLC), or directly by regulating ion channel or kinase
function.
Regulation of G-Protein Activity
Regulation of G-Protein Activity
Regulation of G-Protein Activity
Regulation of G-Protein Activity
G-protein-mediated pathway
Adenylate cyclase mediated pathway
Phospholipase mediated pathway
Small G-protein-mediated pathway
G-protein-mediated
pathway
Adenylate cyclase mediated pathway
cAMP can activate protein kinase A(PKA), which can phosphorylate CREB（
binding protein of cAMP-respones element）and initiate gene transcription.
CRE is cAMP response element in DNA.
G-protein-mediated pathway
Phospholipase C mediated pathway
Non-G-protein-mediated pathway
Receptor tyrosine kinase mediated pathway
Receptor serine/threonine kinase mediated pathway
Receptor guanilate cyclase mediated pathway
Intracellular (unclear) receptor mediated pathway
Receptor tyrosine kinase mediated pathway
Receptor tyrosine kinases transmit signals across the plasma membrane,
from the cell exterior to the cytoplasm.
The interaction of the external domain of a receptor tyrosine kinase with
the ligand, often a growth factor, up-regulates the enzymatic activity of the
intracellular catalytic domain, which causes tyrosine phosphorylation of
cytoplasmic signaling molecules.
Receptor tyrosine kinase mediated pathway
Receptor tyrosine kinases transmit signals across the plasma membrane,
from the cell exterior to the cytoplasm.
The interaction of the external domain of a receptor tyrosine kinase with
the ligand, often a growth factor, up-regulates the enzymatic activity of the
intracellular catalytic domain, which causes tyrosine phosphorylation of
cytoplasmic signaling molecules.
Mechanism of Tyrosine Kinase Receptors
When hormone binds to the extracellular domain the receptors aggregate
Mechanism of Tyrosine Kinase Receptors
When the receptors aggregate, the tyrosine kinase domains phosphorylate
the C terminal tyrosine residues
Mechanism of Tyrosine Kinase Receptors
This phosphorylation produces binding sites for proteins with SH2 domains.
GRB2 is one of these proteins. GRB2, with SOS bound to it, then binds to the
receptor complex. This causes the activation of SOS.
Mechanism of Tyrosine Kinase Receptors
SOS is a guanyl nucleotide-release protein (GNRP). When this is
activated, it causes certain G proteins to release GDP and exchange it for
GTP. Ras is one of these proteins. When ras has GTP bound to it, it
becomes active.
Mechanism of Tyrosine Kinase Receptors
Activated ras then causes the activation of a cellular kinase called raf-1
Mechanism of Tyrosine Kinase Receptors
Raf-1 kinase then phosphorylates another cellular kinase called
MEK. This cause the activation of MEK
Mechanism of Tyrosine Kinase Receptors
Activated MEK then phosphorylates another protein kinase called MAPK
causing its activation. This series of phosphylating activations is called a
kinase cascade. It results in amplification of the signal
Mechanism of Tyrosine Kinase Receptors
Among the final targets of the kinase cascade are transcriptions factors (fos
and jun showed here). Phosphorylation of these proteins causes them to
become active and bind to the DNA, causing changes in gene transcription
Signal Transduction Through Receptor Tyrosine Kinases
Receptor serine/threonine kinase mediated pathway
(1) Type I and type II
receptors for TGF(beta) in
a cell prior to binding of
the growth factor.
(2) Binding of growth factor
results in clustering of type
I and type II receptors, and
phosphorylation of type I
receptors by type II
receptors.
(3) The activated type I
receptors then
phosphorylate particular
receptor-mediated Smads.
(4) These Smads then bind to
other Smads (co-Smads),
and together they enter the
nucleus.
Intracellular (nuclear) receptor mediated pathway
Basic Structure of nuclear receptor
Hormone-bindind
domain
DNA-binding
domain
Transcriptionactivating domain
nuclear receptor mediated pathway
The signal pathway by steroid hormones
The signal pathway by steroid hormones
Networks of Signal Transduction
600 G protein-coupled receptors
+
Multiple gene families and combinations
of G protein subunits
20G isoforms
6 G isoforms
12 G isoforms
+
Multiple gene families for selected effector proteins
Adenylyl cyclases
Phospholipases
Ion channels
Cascade structure of cellular signal pathways
The magnitude of amplification within this cellular cascade structure often
exceeds 10+4. That is, the binding of one molecule of ligand to a cell-surface
receptor leads a change of 10,000-fold in the intracellular concentration of a
metabolic product.
CHAPTER 2
Dysfunction of cellular signal
transduction in diseases
Aberrant Signal in cell signaling
Aberrant Receptor in cell signaling
Aberrant G-protein in cell signaling
Aberrant Intracellular Signaling
Multiple Abnormalities in cell signaling
Aberrant Signal in cell signaling
ischemia, epilepsy, neurodegenerative diseases
extracellular glutamate/aspartic acid
NMDAR activation
Ca2+ influx
[Ca2+]i , activation of enzymes
excitatory intoxication
Aberrant Receptor in cell signaling
Receptor-based diseases
Alterations in number, structure or function of receptors
will lead to disorder in cellular signal transdution
Up-regulation/hypersensitivity
Down-regulation/desensitization
Receptor Gene Mutation
Myasthenia Gravis
Myasthenia Gravis is an autoimmune receptor disorder in
which antibodies form against acetylcholine(Ach) nicotinic
postsynaptic receptors at the neuromuscular junction
mechanism
anti n-AchR
AchR
Ach
The Neuromuscular
Junction
Contraction of
muscle fiber
influx of Na
manifestations
Drooping of the eyelids
Double vision
Difficulty smiling, speaking, swallowing
Difficulty raising the arms
Difficulty walking
Difficulty breathing if chest muscle are affected
Autoimmune Thyroid
Diseases
hyperthyroidism (Grave's disease)
hypothyroidism (Hashimoto's thyroiditis)
mechanism
Stimulatory Ab
Blocking Ab
TSH-R
TSH-R
Gs
Gq
295~302
385~395
AA residues
PLC
AC
IP3
DG
Ca2+
PKC
Binding of TSH to R↓
cAMP
Thyroid proliferation & secretion 
hyperthyroidism
hypothyroidism
manifestations
Grave's disease
Stimulatory antibodies mimic the function of TSH
Stimulating thyroid hormone synthesis, secretion,
and thyroid growth
female:male incidence -- 5:1 to 10:1
diffusely enlarged goiter
manifestations
Hashimoto's thyroiditis
Inhibitory antibodies antaonize the function of TSH
Inhibiting thyroid hormone synthesis, secretion, and
thyroid growth
Thyroid gland is gradually destroyed
Myxedema
Receptor Gene Mutation-Genetic insulin-resistant diabetes
Diabetes Mellitus Type 1
Diabetes Mellitus Type 2
Non-Insulin dependent diabetes mellitus, NIDDM
NIDDM is a chronic metabolic syndrome defined by resistance to
the hormone insulin. This leads to inappropriate hyperglycaemia
(increased blood sugar levels) and deranged metabolism of
carbohydrate, fats and proteins.
mechanism
The cause of Diabetes Mellitus Type 2 is not known, but it may involve a defect
or change in the insulin receptor (IR).
Genetic insulin-resistant diabetes
IR gene mutations
synthesis
transfer to the membrane
Disturbances in
affinity to insulin
RPTK activation
proteolysis
Diabetes Mellitus Type 2
G protein-based disease
GHRH
pituitary tumor
Pituitary
GHRH Receptor
Gsα
(+
)
cAMP
(-)
GH secretion
Gi
GHRH--Growth-hormone-releasing hormone
somatostatin
GH--Growth-hormone
mechanism
Gs gene mutation
GTPase activity
Persistent activation of Gs
Persistent activation of AC
cAMP
Pituitary proliferation and secretion
Acromegaly or Gigantism
manifestations
G-protein modification——
cholera
lumen of intestine
Na+
H2O Cl-
cAMP ↑ ↑ ↑
CT
CT--Cholera toxin
Gs
AC
Gs ribosylation at Arg201
manifestations
Diarrhea
Dehydration
Circulation failure
Aberrant Intracellular Signaling
The intracellular signaling involves various messengers,
transducers and transcription factors. Disorders can occur in
any of these settings.
WNT sinal pathway
Cancer
Wnt-1 was found as an oncogene activated by the Mouse Mammary Tumor Virus in murine breast cancer.
APC was first isolated as a tumor suppressor gene in human colon cancer. After establishing that APC and
beta-catenin bind to each other activating mutations in the human beta-catenin gene were found in human
colon cancer and melanomas .These mutations alter specific beta-catenin residues important for GSK3
phosphorylation and stability .The role for Frat/GBP in cancer is illustrated by its activation by proviral
insertion in mouse lymphomas. Interestingly, mutations in the human AXIN1 gene were reported in human
hepatocellular carcinomas. TCF1 can also act as a tumor suppressor gene , as Tcf1 mutant mice develop
adenomas in the gut and mammary glands
Multiple Abnormalities in Signaling Pathway
In the development of diseases, the aberrant cellular signal
transduction usually involves multiple molecules or pathways.
Such diseases include type-2 diabetes mellitus, cancers,
hypertension, and so on
Multifactor Aberrancies and Cancer
(Enhancement of proliferating signals)
Ligands (GFs)
Receptors (overexpression, activation of TPK)
Intracellular transducers：
Ras mutation Ras-GTPase
MEK
ERK
Proliferation
Ras activation
Cancer
Raf
Multifactor Aberrancies and Cancer
(Deficits in proliferation-inhibiting signal)
（TGF-β）2
Cell memberane
GS
Ⅱ
Ⅰ
Ⅱ
Ⅰ
Smad2
SARA
（—）
Smad6,7
Smad2 -P
Smad4
Smad4
Cytosal
Smad2 -P
Nuclear membrane
P300
P300
Smad4 Smad2 -P
Fast2
Fast2
P15、P21
Principles for Treatment
To regulate the level of extracellular molecules
To regulate the structure and the function of receptors
To regulate the level and modifications of intracellular
messenger molecules and transducers
To regulate the level of nuclear transcription factors
Target Therapy
The Philadelphia Chromosome:
t(9;22) Translocation
Chr.9+
Chr.9
Chr.22
Ph
bcr
abl
bcr-abl
FUSION PROTEIN
WITH TYROSINE
KINASE ACTIVITY
Structure of BCR-ABL Fusion Proteins
p210Bcr-Abl Fusion Protein Tyrosine Kinase
Extracellular
space
Cytoplasm
Y177
BAP-1
SH3
SH2
SH1
GRB2
CBL SHC CRKL
Gleevec®-Tyrosine Kinase Inhibitor
Bcr-Abl
Bcr-Abl
Substrate
Substrate
P
P
P
ATP
P
STI571
Y = Tyrosine
P = Phosphate
Goldman JM. Lancet. 2000;355:1031-1032.