Chapter 15

Cellular Signal Transduction

The biochemistry and molecular biology department of CMU

When environment changes:

Monad——responds directly.

Multicellular organisms——signal through elaborate system of intercellular or intracellular communication ， and consequently regulate functions of organisms.

Signaling molecule

Receptor of target cell

Intracellular molecule

biological effect

Signaltransduction

§1 Signaling Molecules

Signaling molecules

• Signaling molecules, which are released by signal-producing cells, reach and transfer biological signals to their target cells to initiate specific cellular responses.

• 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, PG

(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 lipid derivatives

• IP3 carbohydrate

derivatives

• cAMP cGMP nucleotides

• Ras, JAK, Raf proteins

Second messenger:

Small molecules synthesized in cells in response to an external signal are the second messengers, which are responsible for intracellular signal transduction.

Such as Ca2+, DG, Cer, IP3, cAMP, cGMP

Third messengers:

Third messengers are the molecules which transmit message from outside to inside of nucleous or from inside to outside of nucleous, also called DNA binding protein.

Effect by membrane receptors

Effect by intracellular receptors

Intracellular molecules

Extracellular molecules

Signal molecules

cAMP, cGMP, IP3, DG, Ca2+

Proteins and peptides:

Hormones, cytokines

Amino acid derivatives:

Catecholamines

Fatty acid derivatives:

Prostaglandins

Steroid hormones, Thyroxine, VD3

§2 Receptor

Receptor

Receptors are specific membrane proteins, which are 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; for example, a hormone is the ligand for

its specific protein receptor.

• Membrane receptors

membrane

Glycoprotein

• Intracellular receptors

Cytosol or nuclei

DNA binding protein

(1) Ligand-gate ion channels type

(cyclic receptor)

ligand→receptor→ion channel open or close

1. membrane receptors

1) 7-helices transmembrane receptor

(2) G Protein-Coupled Receptors (serpentine R)

Cytosolicside

Oligosaccharideunit

• G protein refers to any protein which binds to GDP or GTP and act as signal transduction.

• G proteins consist of three different subunits (, , -subunit). • -subunit carries GTPase activity, binding and hydrolysis of GTP.

2) G protein (Guanylate binding protein)

Gs→ s→AC→cAMP↑

　 Gi→ i→AC→cAMP↓

　 Gq→ q →PI-PLC→IP3+DAG

　 Go→ o→ion channel

Gt→ t →cGMP PDE→cGMP→

Rhodopsin

3) Classes of G protein

Glucagon

-adrenaline →s →AC↑

ACTH

-adrenaline

angiotensin Ⅱ

acetylcholine(M2 M4)

GF release inhibitory factor

→i→AC↓

　

Ribosylation of Arg of G

G s -ATP Gs -ADP

AC

cAMP Cl- + H2O Cavity of intestinediarrheaHCO3

-

Cholera toxin

ATPase

Pertussis toxin

i -ADP-ribosylation

Gi

AC↑

allergy of histamine

cAMP ↑

AC

cGMP PDE (phosphodiesterase ）

PLC

PLA2

Channel protein

4) Effect proteins of G protein

cAMP

ATP

5) Pathway of G protein linked receptor

H R G protein Es

secondary messeger

Protein kinase

Phophorylation of Es or functional protein

Biological effect

(3) Single transmembrane α-helix receptor

• Tyrosine protein kinase Receptor

(catalytic receptor)

IGF － R, EGF － R

• Non tyrosine protein kinase Receptor

Growth Hormone R, interferon R

Tyrosine protein kinase Receptoror receptor tyrosine kinase (RTK)

Cys-richdomain

Immunoglobulin-like

domain

EGFR IGF-1R PDGFR FGFR

Intracellularinsulin effectsCytosol

Insulin

Non-receptor tyrosine kinase (NRTK)

Domain of Downstream molecules of TPKR

SH2 domain (Scr homology 2 domain)

SH3 domain

PH domain (pleckstrin homology domain)

Membrane receptor –ANP

Soluble receptor – NO, CO

(4) Guanylate cyclase (GC) receptor

2. Intracellular receptor (transcription regulated

receptor) Intracellular R is trans-acting

elememt cis-acting element gene expression

Localized in the cytosol and/or in the nucleus.

ligand: Steroid H, VD3, Thyroxine

3. Properties of binding of H and R

• highly specificity

• highly affinity

• saturation

• reversible binding

• special function model

4. Control of receptor activity

• Phosphorylation or dephosphorylation of R

• Phospholipid of membrane

• Enzyme catalyzed hydrolysis

• G protein regulation

(1) Recognize the special ligand

(2) Binding to special ligand

(3) Signal transduction biological effect

5. Function of receptor

§3 Pathway of Signal

Transduction

Signal transduction mediated by membrane receptor

• cAMP dependent-protein kinase A pathway

• cGMP dependent PKG pathway 　• Ca2+ dependent PK pathway 　• Tyrosine protein Kinase pathway

• NF-κB pathway 　• TGF- βpathway

1. cAMP dependent-protein kinase A pathway

H R

G protein

Phosphorylation of Es or functional proteins

Biological effects

PKA

cAMPAC

(1) cAMP metabolism

ATPMg2+

PPi

cAMP

H2OMg2+ 5'-AMP

AC PDE

AC ： Adenylate cyclase

PDE ： Phosphodiesterase

PDE

H2O

N

N N

N

NH2

O

H

OHOH

CH2

H

H H

OPO

O

O

PO

O

O

P

O

O

O ATP

ACPPi

N

N N

N

NH2

O

H

OHO

CH2

H

H H

O

PO

O

cAMP

N

N N

N

NH2

O

H

OHOH

CH2

H

H H

OPO

O

O

5'-AMP

(2) Mechanism of cAMP effect

Activate cAMP-dependent protein kinase (PKA).

(3) PKA effect

Phosphorylate specifically

Ser/Thr residues in several proteins

（ 1 ） Regulation of metabolism

（ 2 ） Regulation of gene

expression

hormons: glucagon, epinephrine

inactive AC active AC

ATP cAMP

inactive PKA active PKA

phosphorylase b kinase

phosphorylase b kinase

P

ATP

ADP

H2O

Pi

phosphorylase b

P

P

ATP ADP

Pi

H2OATP ADP

glycogen synthase

glycogen synthase

P

H2OPi protein phosphatase-1

(active) (inactive)

inhibitor-1 (active)

inhibitor-1 (inactive)

phosphorylase a

ATP

CRE : cAMP response element

(TGACGTCA)

CREB: CRE binding protein

DNACRE

CREB

PKA

Ｐ

Transcription

mRNA

CREB

PKA

Ｐ

CREBＰ

CR

EBＰ

CR

EB

Ｐ

CREBＰ

CR

EB

Ｐ

CR

EB

Ｐ

2. Ca2+ dependent PK pathway

(1) Ca2+ -DAG -dependent PKC pathway

H R

PIP2

G protein PLC

IP3 DG

ERPKC

Ca2+ PS

Biological effects

Phosphorylation of Es or functional proteins

[Ca 2+]i 0.01-1 mol/L （ 10-7 mol/L ）

[Ca 2+]o 2.5mmol/L （ 10-3 mol/L ）

5000~10000×

1 ） Function of DG and IP3

IP3 + R→open of Ca2 ＋　channel →[Ca2 ＋ ]↑

PS, Ca2 ＋DG PKC ↑

2 ） Function of PKC

• regulation of metabolism

PKC →Ser/Thr-P of R, enzyme,

Protein of Mb.

• Gene expression 　

Late response:

Trans-acting factor-P

Third messengerImmediate early genes

Third messenger-P

Activate genes

Cell proliferation

PKC

PKC

Early response:

(2) Ca2 ＋ -CaM dependent protein kinase pathway

H R G protein PLC

IP3

Ca2+ CaM

CaMK

Biological effects

Phosphorylation of Es or functional proteins

Calmodulin (CaM ）： Ca2 ＋　binding protein

4 Ca2 ＋ + CaM → Ca2 ＋ - CaM

↓ CaM kinase↑

↓　 　 Ser/Thr - P ↓

Ca2 ＋　pump, AC ↑ GC ↑

Es (glycogen synthase, phosphorylase kinase)

3. cGMP-dependent PKG pathway

Biological effects

Phosphorylation of Es or functional proteins

ANPNO, CO

Soluble GC

PKG

cGMPReceptor-linked GC

(1) cGMP

(2) Function of PKG

Ser/Thr- P of protein and E

Mg2+

PPi H2O

Mg2+GTP cGMP 5'-GMP

GC PDE

GC: Guanylate cyclase

ANP （ atrial natriuretic peptides ） ↓

GC NO

↓

cGMP

↓

PKG

↓

Vascular dilatation

cis-acting elementnucleus

expression

cell memberane

phosphorylation of enzymes or proteins effects

CaM-PK

Ca2+-CaMCa2+

IP3DGcAMP cGMP

PKA PKG PKC

ATP GTP PIP2(PC)

G AC GC G PL

peptide hormonesneurotransmitters AFP

neurotransmittershypothalamic pituitrin

Ptrans-acting factor

4. Tyrosine-protein kinase pathway (TPK)

• TPK receptor is related to proliferation, differentiation, dissociation, carcinomatous change.

• TPK ：receptor TPK ： Mb.

non receptor TPK ： cytosol

(1) Receptor TPK － Ras －MAPK pathway

• GRB2, SOS, Ras, Raf

• Small G protein: Ras

• MAPK (mitogen-activated protein kinase) ：

MAPK 、 MAPKK 、 MAPKKK

ÊÜÌå ÐÍTPKϸ°ûĤ

EGF, PDGF

receptor TPK Ras － GTPSOS － PGRB2 － P

Raf － P

MAPKK － P

MAPK － Ptrans-acting factor

expression

nucleus

－ P

(2) JAKs － STAT pathway

ligand

non TPK receptor

JAKs STAT

gene expression

• JAKs

• STAT: Signal transductors and

activator of transcription

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JAK

11391

84

48

113

P

91P 84 P

48

113

91 84P P

P

interferons

inactive STAT

Plasma membrane

STAT complex

Nuclear membrane

geneInterferons response element

transcription

Dimerization of Receptor

Interferon

Autophosphorylation of JAK

Phosphorylation of STAT

Nuclear translocation of STAT

Interferon response element

Expression of gene

5. Nuclear factor-κB pathway

6. TGF-βpathway

Intracellular receptor (DNA transcription regulated

receptor)

• Steroid H, VD3, Thyroxine

• Cytosolic R: glycocorticosteroid H

• Nuclear R: thyroxine, estrogen, androgen, progesterone

Serum binding proteinWith bound bormone