Antibody staining (Lecture 3, p.4). In situ hybridization (Lecture 4 p.4)

Antibody staining (Lecture 3, p.4)

In situ hybridization (Lecture 4 p.4)

mRNA depletion (“knockdown”) methods (Lecture 3, p.4)

-catenin

-catenin builds up

Axin

Wnt is absent. -catenin mRNA is continuouslytranslated and the protein is continuously degraded.

Wnt is present. -catenin protein is not degraded. It builds up, binds to Tcf/LEF and enters the nucleus.

Wnt11 protein

Outside cell

Inside cell

-cateninmRNA

Wnt, beta-catenin, and organizer formation

*

*

*

**

*

*

Endo-mesoderm induction. Pieter Nieuwkoop 1968

(Nieuwkoop center)

(Nieuwkoop center)

Nodal signaling pathway(belongs to TGF-beta family of pathways)

phospho-Smad2

A dominant negative form of the Nodal (or Bmp)

receptor can block Nodal (or Bmp) signaling

Deplete and replace vegT mRNAvegT depletedNo mesoderm or endoderm,No blastopore

vegT mRNA restored by injection into vegetal hemisphere

vegT depleted and restored

VegT depleted embryos: rescue with mRNA for xnr1, 5, 6, or derriere, injected into vegetal pole cells

Normal late blastula:xnr1 expressed in vegetal cells

derriere rescue

xnr5, xnr6 rescue

xnr1 rescue

xnr1