W11L1 Flashcards
Cells need to rapidly respond to changes in the environment
Cells need to rapidly respond to changes in the environment, whether this a normal process (such as development) or exposure to foreign substances (carcinogen, pharmaceuticals, etc.)
- can happen quickly in minutes!
- cell needs to be ready to adapt
What’s needed for a rapid response (i.e. change) in gene expression due to an environmental stimulus?
- Something on the surface to recognize the environment change – receptors (ligands) or channels (ions, chemicals)
- Existing networks of proteins that can rapidly switch from an active to inactive state (or vice versa)
- Mechanism for entering the nucleus quickly and efficiently
- Genes that are poised to be activated (or repressed)
WNT signaling
WNT signaling was first identified in 1982 in cancer biology; refers to Wingless and Int-1
- most work was done in drosophila
Allows cross talk between cells (paracrine or autocrine signaling) or from one developmental structure to another
There is a canonical and three non-canonical pathways
- Pathways trigger changes in gene expression in direct or indirect way
- all pathways lead to effect on transcription
There are common parts to each pathway including:
(a) WNT ligand (19 in total)
(b) Frizzled receptor (FZD) for recognizing WNT
Differential WNT expression, different FZD receptors and co-factors, and downstream mediators dictate WNT signaling outcomes
- e.g. WNT5 activates non-canonical pathways.
WNT signalling - 1 canonical and 3 non-canonical pathways
- Canonical
- Non-canonical
- planar cell polarity
- WNT11 - Non-canonical
- Wnt5/calcium - Non-canonical
- Wnt5/Fzd2
Canonical signalling - WNT/β-catenin - OFF state
OFF STATE
RSPO is floating extracellularly, WNT is bound to sFRP.
Cell surface proteins bind to frizzled (FZD) and target it for degradation (by ubiquitination; Ub)
- this is because WNT is not bound to FZD
Disheveled (DVL) is ubiquitinated and targeted for degradation
β-catenin is bound by the β-catenin destruction complex (Axin, APC and GSK)
β-catenin is phosphorylated and targeted for degradation by the proteasome
TCF is bound to target genes but is not active
Canonical signalling - WNT/β-catenin - ON state
ON STATE
RSPO is bound to LGR4, then gets internalized into the cell. WNT is not bound to sFRP, WNT is bound to FZD
R-spondin (RSPO) binds to surface co- factor and prevents ubiquitination of FZD
- RSPO is cofactor for WNT to allow it to be activated
WNT binds to FZD and triggers activation of the pathway
Disheveled (DVL) is not ubiquitinated and binds to FZD
Active DVL de-activates the β-catenin destruction complex
- β-catenin is not phosphorylated
β-catenin translocates to and enters the nucleus and binds T Cell Factors (TCF) to activate target genes
Overall, Wnt signaling inhibits the β-catenin destruction complex, preventing its degradation and allowing translocation to the nucleus
Factors for rapid response with WNT signalling - Canonical
- Something on the surface to recognize the environment change
- Frizzled receptor recognizing WNT ligands (e.g. WNT3) - Existing networks of proteins that can rapidly switch from an active to inactive state
- β-catenin stabilization and release from the destruction complex - Mechanism for entering the nucleus quickly and efficiently
- Several mechanisms have been reported that involve interaction with other transcription factors, the nuclear pore complex and importins - Genes that are poised to be activated (or repressed)
- Some target genes are maintained in a “poised” state by epigenetic modifications. β- catenin may also be able to displace repressive factors binding LEF1/TCF
β-catenin in pathway = canonical
WNT/β-catenin in Development
Canonical WNT signaling is required for most developmental processes
- Deletion of β-catenin or WNT3 causes early embryonic lethality BUT…
- Need the appropriate amount of WNT signaling
WNT signalling is important for early development
- DKK is a repressor that prevents WNT signalling btw
- Delete an inhibitor of WNT signaling leads to loss of anterior neural development
Dosage is important
WNT/Non-canonical pathways - WNT11
- Something on the surface to recognize the environment change
- Frizzled receptor recognizing WNT ligands (WNT 11) - Existing networks of proteins that can rapidly switch from an active to inactive state
- JUN (Mitogen Activated Protein Kinase; MAPK) phosphorylation - Mechanism for entering the nucleus quickly and efficiently
- Phosphorylation allows nuclear localization (JUN)
- JUN enters the nucleus - Genes that are poised to be activated (or repressed)
- JUN target genes are numerous and may already be expressed at low levels
WNT/Non-canonical pathways - WNT5/Calcium
- Something on the surface to recognize the environment change
- Frizzled receptor recognizing WNT ligands - Existing networks of proteins that can rapidly switch from an active to inactive state
- Protein Kinase C (PKC) Activation - Mechanism for entering the nucleus quickly and efficiently
- Phosphorylation leads to activation of a nuclear transcription factor (PKC-NFAT)
- PKC phosphorylates NFAT to allow it to enter the nucleus
- NFAT is very responsive to calcium and causes change in gene expression and effects inflammatory processes - Genes that are poised to be activated (or repressed)
- NFAT contributes to existing transcriptional complexes
Loss of WNT5 expression
Loss of WNT5 expression leads to
- truncated Anterior to Posterior axis (with incomplete outgrowth of distal limbs, genitals and tail)
- shortening of limbs (skeletal defect)
- impaired distal lung morphogenesis
Increased WNT5 expression
Increased WNT5 expression leads to
- cranial facial defects
- abnormal skull development
- loss of hair follicles
DOSAGE IS IMPORTANT!
Dominant β-catenin mutations
Dominant β-catenin mutations (β-catenin is not phosphorylated and is always active due to the mutations) cause…
- intellectual disability with recognizable syndromic features
- microcephaly, a full tip of the nose, and thin upper lip
- Recapitulate in mice
Different role for β-catenin
β-catenin was originally as a component in the adherens junction complex
β-catenin interacts with other cadherins to help it signal from the periphery
- this was the original discovery of the function of β-catenin, it was only later that they discovered it is in WNT signalling to the nucleus
The membrane bound complex allows signaling and attachment to the cytoskeleton
β-catenin is mostly found in cell membrane
β-catenin is part of the complex that links the cell membrane to the actin cytoskeleton
Loss of function and gain of functions in β-catenin can have multiple affects including cell communication, signaling and gene expression
WNTs in stomach and intestinal epithelium
Stomach and intestinal epithelium is constantly turning over (cells die every 3-5 dies)
- Stem cells sit at the base of the glands to maintain the cells
WNT signalling at base of glands to keep cells in undifferentiated state
High levels of WNT signaling allow cells to proliferate and maintain an un-differentiated fate
- As cells divide and move up the gland, WNT signaling decreases to allow for differentiation
