The Wnt Signalling Network Flashcards
Describe paracrine cell signalling
Cells in all multicellular organisms must communicate with each other in order to
organise themselves into a functioning
unit
What are the prominent paracrine signalling systems
Growth factors
* Wnt/Frizzled signalling pathway
* Hedgehog Signalling pathway
* Fgf signalling pathway (RTK Pathway)
* SMAD Signal Transduction Pathway
(TGFβ-BMP)
* mTOR Signalling
Cytokines
Neurotransmitter
Describe the history of Wnt
- Screen for genes, which can
lead to cancer in mouse - Retroviral insertion leads to
activation of downstream
genes - Identification of an insertion
Int1 - Screen for genes, which
regulate embryonic
development in fruit flies
Chemical screen leads to
knock-out of genes - Identification of a gene
named after the phenotype
Wingless - Wingless + Int1 = Wnt1
Describe the Wnt Signalling pathway in 1990
Functions:
Embryogenesis: generation of wings in drosophila
Diseases: Oncogene in mouse
Describe the different examples that show the Wnt / B-catenin pathway is highly conserved
- Localisation of Wnt1-
mRNA in Hydra
-Localisation of armadillo
(=B-Catenin) in Drosophila
- Wnt activity in mouse
embryo shown by a
Wnt-reporte - Mutation in PTK7 lead to
congenital skeletal disorder - Mutation in APC lead to bowel cancer
(familial adenomatous polyposis)
Describe the Wnt Signalling network in 2020
Functions:
Embryogenesis:
Gastrulation
Body axis formation
Patterning of nervous system
Neural crest induction
Hair follicle growth
Limb polarity
Muscle development
Regeneration:
Limb bud
Heart
Brain
Diseases:
Colon carcinoma
Mamma carcinoma
Melanoma
Alzheimer’s diseases
Diabetes
Describe the Wnt Signalling network
- Planar Cell Polarity
signalling:
-> Cell polarity and
tissue migration - β-Catenin signalling
-> Cell differentiation
and cell proliferation - Wnt-Ca2+ signalling
-> Cell fate acquisition
and cell migration
Describe the structure of Wnt ligands [slides]
Multiple Wnt pathway genes in any animal genome
e.g. in human 19 Wnt ligands, 10 Frizzled
receptors and 5 co-receptors
mWnt8
* ca. 350 aa
* 22 Cysteines
* Serine 187 > palmitoleic acid (lipid adduct)
* Asparagine 87, 298 > glycan (glycolisation)
Describe production and secretion of the Wnt ligand
- Wnt is lipid modified in the ER by the
membrane bound Oacetyltransferase Porcupine - Palmitolylated, lipidated Wnt is
transported from ER -> Golgi by p24 - Wnt is transported from the Golgi to
the membrane by Wntless/Evi (check
point). - Wnt is loaded on signalling filopodia,
so-called cytonemes - Or Wnt is re-internalised in
endosomes, routed to the multivesicular bodies, packaged on
vesicles and released.
Describe the interaction of the Wnt ligand with its receptor
- Frizzled belongs to the family of
GPCR. Frizzled has a 7 transmembrane domain and a
cysteine-rich domain (CRD) - Wnt binds to the CRD of Frizzled
as a monomer. - Two finger-like domains grasping
Frizzled at two binding sites. - The ‘lipid thumb’ of Wnt (Nterminal domain) is dominated by
the palmitoleic lipid projecting
into a groove in the Frizzled CRD. - Cysteine residues are engaged in
intramolecular disulphide bonds
(not post-translationally modified).
Describe the Wnt-OFF state
- Receptor Fzd and co-receptor Lrp6 are
continuously endocytosed and recycled
back to the membrane -> this
determines the overall level of
receptors at plasma membrane. - The β-catenin destruction complex
consists of three structural proteins:
adenomatous polyposis coli (APC),
Dishevelled (Dvl) and Axin1 and the
main kinase, glycogen synthase kinase
3β (GSK-3β) - β-catenin is phosphorylated by GSK-3β
phosphorylates and ubiquitylated by βTRCP (“kiss of death”) -> degradation in
proteasome
-> Intracellular level of β-catenin is kept
low in the Wnt-OFF state
Describe B-Catenin/armadillo
Key effector of Wnt signaling
- β-catenin is the main effector of
canonical Wnt signalling. - β-catenin interacts with the
scaffold proteins Axin1 and APC - β-catenin can be phosphorylated
by GSK-3β and ubiquitylated by βTRCP - β-catenin can translocate to the
nucleus, engage with the
transcription factors TCF/Lef to
regulate gene transcription. - β-catenin has a further role in cell
adhesion together with Ecadherin
Describe activation of the pathway - the Wnt-ON state
- Formation of the ligand-receptor: Wnt
binds to Frizzled and Lrp6 -> Lrp6 is
phosphorylated - Recruitment of components of the βCatenin destruction complex to the
plasma membrane: Frizzled binds to
Dishevelled and Lrp6 to Axin1. coreceptor and both structural proteins are
phosphorylated. - Deactivation of the destruction complex:
Confirmation change of Axin1 blocks
phosphorylation of β-catenin by GSK-3β - Intracellular level of β-catenin increases
in the Wnt-ON state and β-Catenin
translocates into the nucleus - β-Catenin binds to DNA to regulate the
transcriptional profile (i.e. c-Myc -> cell
growth and proliferation).
Describe the “Handbrake system” mechanism
- Temporal control is essential for all
signalling pathways. - All components have been produced
and are in place, effective complexes
have been assembled - System is ready to go!
- Wnt ligand-receptor interaction
releases the “handbrake” - Other examples are the Hedgehog
pathway, TGFβ/BMP pathway.
Describe the “axin loop” neagtive feedback loop
- Wnt signalling activates the
transcription of Axin - Axin is an inhibitor for Wnt/βcatenin signalling
- Axin helps to keep Wnt/βcatenin signalling at bay by
acting in a negative feedback
loop - Ribosyltransferase Tankyrase
mediates poly-ADP
ribosylation of Axin ->
degradation - Chemical inhibitors of Tnks
block Wnt signalling
Describe engineering of LRP6-FZD heterodimers
Based on the structure of receptors and
co-receptors, Wnt surrogates can be
generated.
Wnt surrogates activate signalling by
clustering essential receptors
independent of a ligand.
- can be easily produced
- are soluble (non-lipidated agonists)
- facilitate functional studies of Wnt
signalling
- allow the exploration for translational
applications in regenerative medicine.
Describe Wnt/B-catenin signalling in diseases
- The large intestine (colon, large bowel)
includes a mucosa with 10 millions
intestinal villi and crypts. - Per day 10 billion cells (ca. 200g) are shed
of the villi tips into the gut lumen per day. - Wnt signalling is important for tissue
homeostasis in the intestinal crypt - Wnt ligands are expressed in Paneth cells
and stroma cells. - Wnt signalling regulates proliferation of
the adjacent stem cells - Cells differentiate to enterocytes and
Goblet cells.
The APC protein functions a tumour suppressor gene (or an anti-oncogene)
1. Colon cancer may occur when the APC gene or β-catenin gene are mutated
2. APC can no longer keep β-catenin out of the nucleus
3. Inside the nucleus β-catenin can displace proteins including SMAD4, Groucho etc to
upregulate proteins (c-Myc) for cell division > uncontrolled proliferation
4. Ultimately, this can lead to tumour formation, more mutations, invasion and metastasis.
