WNT

1) Wnts were discovered by Varmus and Nusse as oncogenes activated by MMTV(mouse mammary tumor virus) integration sites and encode secreted signalingfactors. There are about 20 Wnts in mammals. Wnts induce twinning in embryos.

2) Frizzled. Family of seven transmembrane serpentine receptors that transducethe signal. They contain an extracellular cysteine-rich domain (CRD) that issufficient to bind Wnt. Frzbs are Wnt antagonists consisting of a s ecreted CRDlacking the transmembrane portion.

COMPLESSIMULTIPROTEICI

WNT

Nucleotrascrizione

Proteasomadegradazione

Groucho: repressionecostitutiva del promotorebersaglio

beta-catenina: derepressione(alternativa a Groucho) e attivazione(reclutamento cofattori attivatori) delpromotore bersaglio

Partners di LEF

Ruolo del fattore(azione positiva/negativa)

Effetto in presenza di Wnt(attivazione/spegnimento)

HeparanSulfateProteoGlycans

Regolazione extracellulare dei ligandi Wnt

Frzb/FRP: falsirecettori checompetono per illigando Wnt

DKK DKK: falsi “ligandi” checompetono con Wntper il corecettore LRP

REARRANGEMENTS OF THE EGG CYTOPLASM

Sperm centriole directs the polymerization of themicrotubules so that they grow into the vegetal region ofthe egg. The off-center position of the sperm centriole asit initiates microtubule polymerization provides thedirectionality to the rotation. These cytoplasmicmovements initiate a cascade of events that determine thedorsal-ventral axis of the frog. Indeed, the parallelmicrotubules that allow these rearrangements to stretchalong what will become the dorsal-ventral axis.

DORSAL

CHORDINBMP4

Mesenchyme

B lood

Ep i de rm i s Neu ra l

Notochord

Prechordal pl.

ECTODERM

MESODERM

ENDODERM

}

}

}

VENTRALChordin

Wnt/b-catenin

Sperm entry

Effetto di blocco della via di Wnt (per esempio distruggendo imicrotubuli con raggi UV): mancanza di strutture dorsali

mRNA codificanteper beta-catenina

Effetto della espressione ventrale di beta-catenina: formazione di un asse secondario

Chd Chd

Embrione allo stadio di gastrula:Viene trascritto il gene Chordin ventralmenteSi genera un secondo Organizzatore dorsale

BMP

9) TCF-3/Lef-1 for “T-Cell” or Lymphocyte Enhancer Factors are DNA-binding proteins that were isolated because they bind to the T-cell receptorenhancer. However, they were c onsidered “architectural” HMG chromatinproteins because they could not activate transcription of reporter genes. A yeast2-hybrid screen found that the amino-terminal end of TCF bound !-catenin(very nice paper by Mol enaar et al., 1996). In co-transfection assays with areporter construct, both !- catenin and TCF-3 are required for activation. !-catenin is therefore a co-activator of the xTCF-3 transcription factor (ahomologue of Drosophila Pangolin). Deletion of the amino terminus of TCF-3(!N) generates a potent dominant-negative (DN-xTCF-3).

DNA binding

!N is a potent dominant negative (DN-xTCF-3) in co-transfection experimentsbecause it binds to DNA but not to !-catenin. The DN-xTCF-3 was able toblock axis formation by !-catenin mRNA injected in the ventral side (figure),as well as Goosecoid transcription and the endogenous dorsal axis wheninjected in the dorsal side.

Dominant-negative TCF3

Oncosoppressori, mutazioni inattivanti

Oncogene, mutazioni attivanti

Adenoma di un paziente affetto da FAP

Immunolocalizzazione di beta-catenina