SIGNAL TRANSDUCTION

Marigold D.R. Majarucon-Ferrolino, M.D.

SIGNAL TRANSDUCTION

The process of converting extracellular signals into cellular responses.

extracellular signaling molecules (ligands) - substances synthesized and released by signaling cells and produce a specific response only in target cells that have receptors for the signaling molecules.

SIGNAL TRANSDUCTION

SIGNAL TRANSDUCTION

Receptor – a specific protein that specifically binds a signaling molecule to initiate a response in a target cellCell responses :

changes in gene expressioncell morphology

cell movements

SIGNAL TRANSDUCTION

Communication by extracellular signals usually involves six steps:

1) synthesis and 2) release of the signaling molecule by the signaling cell 3) transport of the signal to the target cell 4) detection of the signal by a specific receptor protein 5) a change in cellular metabolism, function, or

development triggered by the receptor-signal complex ;and

6) removal of the signal, which often terminates the cellular response.

TWO GENERAL KINDS OF CELL RECEPTORS

CELL SURFACE RECEPTORS

LIGAND – hydrophilic signaling molecules

INTRACELLULAR RECEPTORS

LIGAND – hydrophobic signaling molecules

CHARACTERISTICS OF RECEPTOR PROTEINS

LIGAND-BINDING SPECIFICITY

-a ligand binds to a specific receptor

EFFECTOR SPECIFICITY- receptor-ligand complex mediates a

specific cellular response

TYPES OF SIGNALING

ENDOCRINE SIGNALING - signaling molecules (hormones) act on target cells distant from their site of synthesis by cells of endocrine organs. PARACRINE SIGNALING - the signaling molecules (neurotransmitters) released by a cell only affect target cells in close proximity to it. AUTOCRINE SIGNALING - cells respond to substances (growth factors) which they themselves release.

CLASSIFICATION OF HORMONES BASED ON SOLUBILITY AND

RECEPTOR LOCATION

SMALL LIPOPHILIC MOLECULES that diffuse across the plasma membrane and interact with intracellular receptors.

Examples: steroids, thyroxine and retinoic acidWATER-SOLUBLE HORMONES with cell-surface receptors

Examples: Peptide hormones (insulin, growth factors, glucagons)

CLASSIFICATION OF HORMONES

-SMALL CHARGED MOLECULES ( epinephrine, histamine)

LIPOPHILIC HORMONES with cell surface receptors

Examples: Prostaglandins

( prostacyclins, thromboxanes,

leukotrienes )

MAJOR CLASSES OF CELL SURFACE RECEPTORS

1)     G-protein coupled receptorsExamples: epinephrine,serotonin, glucagon receptors

2)     Ion channel receptors Example : Acetylcholine receptor 3)     Tyrosine kinase-linked receptors Examples: receptors for cytokines, interferons, and

growth factors 4)  Receptors with intrinsic enzymatic activity Examples: receptors for insulin and growth factors

SECOND MESSENGERS

- Intracellular signaling molecules 3’,5’ cyclic AMP (cAMP) 3’,5’ cyclic GMP (cGMP) 1,2 diacylglycerol (DAG) inositol 1,4,5 triphosphate (IP3)

inositol phospholipids (phosphoinositides) Ca++.

OTHER INTRACELLULAR SIGNALING PROTEINS IN SIGNAL

TRANSDUCTION1) GTPase Switch Proteins – GTP-binding proteins

that act as molecular switches in signal transduction pathways“ON” when bound to GTP“OFF” when bound to GDP.Two classes of GTPase switch proteins:a)     Trimeric G protein – coupled directly to activated receptorsb)     Monomeric Ras and Ras-like proteins – linked indirectly via other proteins

INTRACELLULAR SIGNALING PROTEINS

2) PROTEIN KINASES – carry out the process of phosphorylation-opposed by the activity of protein phosphatases

3) ADAPTER PROTEINS – no catalytic activity- contain domains as docking sites for other proteins

MAJOR INTRACELLULAR SIGNALING MECHANISMS

SIGNALING VIA G-PROTEIN-COUPLED RECEPTORS (GPCR)

G-Proteins – GTP-binding proteins

Trimeric proteins ( α β γ )

Coupled directly to activated receptors

GTPases – convert GTP to GDP + Pi

ACTIVE- when GTP is bound

INACTIVE – when GDP is bound

G-PROTEIN –COUPLED RECEPTOR

AC

hormone signal outside GPCR plasma membrane

GTP GDP ATP cAMP + PPi

cytosol

GDP GTP

G-PROTEIN-COUPLED RECEPTORS

Activate events altering concentrations of intracellular mediators (SECOND MESSENGERS)

Common second messengers:

cyclic AMP (cAMP)

Ca++

MAJOR PATHWAYS TO GENERATE SECOND MESSENGERS

CYCLIC AMP (cAMP)

Second messenger produced from hydrolysis of pyrophosphate from ATP

Synthesized by Adenylyl Cyclase

Degraded by cAMP phosphodiesterase to form 5’AMP.

CARBOHYDRATE METABOLSIM REGULATION BY cAMP

cAMP activates glycogen phosphorylase (glygenolysis)cAMP inhibits glycogen synthase (Glycogenesis)Insulin inhibits cAMPGlucagon and Epinephrine activates cAMP

PHOSPHOINOSITIDES

Second messengers derived from phosphorylation of inositol by PI kinase

1. Phosphatidyl inositol (PI)

2. PI 4-phosphate (PIP)

3. PI 4,5-Biphosphate (PIP2)

4. Inositol 1,4,5-triphosphate (PI3)

TWO BRANCHES OF INOSITOL PHOSPHOLIPID PATHWAY

Activated Phospholipase C-ß cleaves PIP2 to generate IP3 and DAG(diacylglycerol)

1. IP3 releases Ca++ from ER

2. DAG together with bound Ca++ activates C-Kinase

C-Kinase phosphorylates cell proteins

Ca++

ATP ADP + Pi

Ca++

IP3

calmodulin

endoplasmic reticulum

Ca++

Ca++-ATPase

Ca++-release channel

SIGNALING BY RECEPTOR TYROSINE KINASES AND RAS

LIGANDS- soluble or membrane-bound protein hormones

NGF, PDGF, FGF,EGF, insulin

Binding stimulates the receptor’s intrinsic protein kinase activity

Functions:cell proliferation,differentia-

tion,cell survival and metabolism

RECEPTOR TYROSINE KINASE (RTK)

RAS – the GTPase monomeric protein that transduce signals from RTK

ACTIVE – when bound to GTP

INACTIVE – when bound to GDP

Not directly linked to RTK

KEY LINKS OF RAS TO RTK

GRB2 – adapter protein for receptor

SH2 domain- binds to phosphotyrosine residue in activated receptor

SH3 domains- bind to and activate Sos

Sos – functions as GEF(guanine nucleotide exchange protein)

- converts GDP-Ras to GTP-Ras

CYCLING OF RAS BETWEEN ACTIVE AND INACTIVE FORMS

1. Guanine Nucleotide Exchange Factor (GEF) facilitates dissociation of Ras from GDP

2. GTP binds while GEF dissociates yielding active Ras*GTP

3. Hydrolysis of bound GTP to regenerate inactive Ras*GDP.

ACTIVATION OF RAS FOLLOWING BINDING OF LIGAND TO RTK

1. Binding of ligand causes dimerization and autophosphorylation of tyrosine residues

2. Binding GRB2 and Sos couples receptor to inactive Ras

3. Sos promotes dissociation of GDP from Ras

4. GTP binds and Sos dissociates from active Ras

SIGNALING BY MAP KINASE PATHWAY

MAP KINASE – serine/threonine kinaseTranslocates into nucleus to phosphorylate proteins involved in transcriptionInduced by activated RasOther proteins involved:Raf – serine/threonine kinaseMEK- a dual-specificity protein kinase

CASCADE OF PROTEIN KINASES

1. Activated Ras binds to N-terminal of Raf

2. Raf binds to and phosphorylates MEK3. MEK phosphorylates and activates

MAP kinase4. MAP kinase phosphorylates nuclear

transcription factors mediating cellular responses

SIGNALING FROM PLASMA MEMBRANE TO NUCLEUS

CRE ( cAmp-response element) – cis-acting DNA sequence in genes regulated by cAMP

CREB (CRE-binding protein)- a transcription factor to which CRE binds

CBP/300 – a co-activator allowing CREB to stimulate transcription

CREB links cAMP to Transcription

1. cAMP activates cAmp-dependent protein kinase (cAPK)

2. cAPK translocates to nucleus and phosphorylates CREB

3. CREB interacts with CBP/300

4. CREB-CBP/300 complex binds to and activates transcription of target genes

MAP KINASES REGULATE TRANSCRIPTION

MAP kinase is activated via RTK-Ras pathway

translocates to the nucleus and phosphorylates activators and repressors of transcription

NF-kß TRANSCRIPTION FACTOR

A heterodimerIn resting cells, found in cytoplasmbound to an inhibitor I-kßIn response to extracellular signal, I-kß is phosphorylated and degradedNF-kß translocates to nucleus and binds to DNA and regulates transcription

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