Manifestation of Novel Social Challenges of the European Unionin the Teaching Material ofMedical Biotechnology Master’s Programmesat the University of Pécs and at the University of DebrecenIdentification number: TÁMOP-4.1.2-08/1/A-2009-0011

WNT SIGNALING

Tímea Berki and Ferenc BoldizsárSignal transduction

Manifestation of Novel Social Challenges of the European Unionin the Teaching Material ofMedical Biotechnology Master’s Programmesat the University of Pécs and at the University of DebrecenIdentification number: TÁMOP-4.1.2-08/1/A-2009-0011

TÁMOP-4.1.2-08/1/A-2009-0011

Discovery of Wnts

Wnt genes: • Wingless gene in Drosophila melanogaster• Int gene in mice• 24 has been discovered• 19 are expressed in mammals• 10 receptor genes - Frizzleds

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Wnt family proteins

Comprises of 19 secreted glycoproteins controlling a variety of developmental processes:• Cell fate specification• Cell proliferation• Cell polarity and cell migration• Different types of cancers • Various processes of aging

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Frizzled (Fz) family receptors• They are 7-TM receptors; however, assembly of an

active Wnt-Fz receptor complex also requires the presence of a co-receptors, the low-density lipoprotein related protein 5 and 6 (LRP5/6)

• Canonical pathway activators: Wnt1, Wnt3, Wnt3a, Wnt7a, Wnt7b, Wnt8

• Non-canonical pathway activators: Wnt5a, Wnt4, Wnt11

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Canonical pathway • In developing thymocytes or in thymic epithelium • Signals from the Wnt-Fz-LRP6 complex lead to the

phosphorylation of three domains of Dishevelled (Dvl), a family of cytosolic signal transducer molecules.

• Activation of Dvl ultimately leads to phosphorylation and consequently inhibition of GSK-3

• Inhibition of GSK-3 results in stabilisation and consequent cytosolic accumulation of -catenin, which then translocates into the nucleus,

• -catenin forms active transcription complexes with members of the T-Cell Factor (LEF1, TCF1, TCF3, TCF4) transcription factor family and transcription initiator p300.

• Successful assembly of the transcription complex leads to the activation of various target genes including cyclin-D1, c-myc, c-jun , Fra-1 VEGFR, etc.

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Non-canonical pathways• Independent from -catenin• Branches into the:

1Polar cell polarity (PCP) or c-Jun-N Terminal Kinase (JNK)/Activating Protein (AP1) dependent

2Ca2+ or Protein kinase C (PKC)/Calmodulin Kinase (CaMKII)/Nuclear Factor of Activating T- cells (NFAT) dependent pathways

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Wnt signaling pathwaysCanonical pathway Wnt/Ca2+

Cytoskeletalrearrangment

Planar cell polarity

Gene transcription

Nucleus

PKC

PLC

NFAT

Ca2+

Calcineurin

RacRhoA

JNK

G proteins?

CaMKII

NFATP

Cytoplasm

Plasma membrane

DIX PDZ DEP

Daam1

ROCK

Prickle

LRP5/6

-catenin

DIX PDZ DEP

AxinGSK3

APC

-TrCP

-cateninLEF/TCF

-catenin

Dsh Dsh

Wnt

Frizzled

Axin

Wnt5a

Frizzled

Wnt11

Frizzled Stbm

No Wnt signal

?

?

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Canonical Wnt pathway

Nucleus

Wnt8

Cytoplasm

Plasma membrane

LRP5/6

Frizzled

-catenin

Axin DIX PDZ DEP

TCF3

Dsh

Dkk1

Krm

Anterior genes

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-catenin in cellular adhesion

Cytoplasm

-cateninAxin

GSK3

APC

-catenindegradation

Wnt

Frizzled

No Wnt signal

Dsh

Nucleus

-catenin

LEF/TCFTranscription

-catenin

P

P

-catenin

Cadherin

-catenin

-catenin

Cadherin

-catenin

-c

aten

in

Cadherin

-ca

teni

n

-c

aten

in

Cadherin

-ca

teni

n

Adherensjunction

+ Wnt signal

Plasma membrane

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Alzheimer’s disease I

Development of NFTsApoptosis

NO

Activated microglia

DNA damage

Excitotoxicity

Cell-cycle activation

Abnormal DNA synthesis

AP

p53

Fast AP toxicity Delayed AP toxicityWnt

Dkk1Bax-

+

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Alzheimer’s disease II

Late stage

↓Phosphorylation of tau

GSK3

Wnt

Frizzled

Akt

PI3K

Dkk1

Krm

↑Phosphorylation of tau

GSK3

Wnt

Frizzled

Akt

PI3K

Dkk1

Krm

Early stage

P

GD3 synthase-cyclin D1

-catenin

GSK3

Wnt

Frizzled

Dsh

Nucleus

-cateninLEF/TCF

bAP

Akt

TÁMOP-4.1.2-08/1/A-2009-0011Inhibition of Wnt and Tcf signaling in the canonical pathway

Growth

-catenin

GSK3

Wnt

Frizzled

Dsh

Nucleus -catenin

TCF4

Frat

PP2A

AxinAPC

b-TrCP

TCF4

Frp

Nkd 1 and 2

Dominant negative Dsh-s

Dominant negative Frizzleds

ICAT

CK-1,2

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PKC isoforms in Wnt signalling• PKC• PKCd• PKCz

TÁMOP-4.1.2-08/1/A-2009-0011The classical view of three independent Wnt signalling pathways1 The canonical pathway is the first and best

characterized Wnt pathway. Signals are coming through the 7 transmembrane domains of Frizzled-receptors, than Dsh is phosphorylated and signal is transmitted via -catenin to TCF/LEF in the nucleus.

2 Ca-dependent Wnt signaling is transmitted by Frizzled-s and G-proteins and the intracellular signaling molecules are CaMKII and different izotypes of PKCs. Inhibitory signals can use TAK and NLK to get into the nucleus. One of the key targets is NF-AT.

3 Planar cell polarity pathway is Ca dependent and using JNK as well as PKCs to transduce signals to the AP1 complex.

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In what diseases are Wnt signalling pathways involved ?• Inflammation• Fibrosis• Cancer

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Wnt Target genesCanonical pathway(-catenin/TCF)

Cyclin D1, MMP-s, c-myc, Cox-2, c-jun, Fra-1, VEGFR, EGFR

Ca2+ pathway(NFAT, NFkB)

IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-8, IL-15, IFN-, GMCSF, TNF-, ICAM-1

PCP pathway(AP1)

Cyclin D1, MMP-s, FasL, Bim, Bcl3, FL1, GMCSF

INFLAMMATION TISSUE REPAIR AND REMODELLING

IL-1, Il-8, IL-6, MMP-s FGF10, TGFb, BMP4, MMP-s

Wnt target genes

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Rheumatoid arthritis

CD34+ bone marrowprogenitorcell infiltration

Inflammatorystimuli

Leukocyte infiltration

Increased Wnt5a

IL6, IL8, IL15,metallo-proteinases

Joint destruction

Wnt1Wnt5aWnt11Wnt13

Fz-2Fz-5Fz-7

Synoviocytes

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Molecular changes of Wnt4 signaling in the aging thymus• Wnt4 uses mainly the -catenin dependent canonical signaling

pathway• Wnt4 expression is decreasing during aging in the thymus• The receptors of Wnt4 signaling are Frizzled-4 and Frizzled-6• The expression pattern of Wnt4 receptors is changing during

thymic senescence• During aging the balance moves towards the Fz-6, transducing

negative Wnt signals• PKCd is modulating intracellularly the Wnt4 signaling

mechanism• CTGF - a target gene of Wnt4 signals- expression is increasing• CTGF is a negative regulator of b-catenin dependent signaling• CTGF and its recently described receptor Fz-8 is functioning as

a secondary negative feedback mechanism of Wnt4 signaling