L4 - Morphogens Flashcards
What is a morphogen?
A soluble secreted molecule that acts at a distance to specify the fates of cells
May specify more than one cell type by forming a concentration gradient
Increasing/decreasing morphogen concentration continually alters the pattern
What led to the discovery of morphogen gradients?
In the molecular age, biologists cloned ligands that they thought might act as morphogens
E.g. a gene that when mutated results in loss of patterning
What are the two requirements of a morphogen?
Induce different outputs at different concentrations
Act directly at a distance
Requirements of a morphogen - induce different outputs at different concentrations
Instructive signal
Permissive signal – not morphogens
- Cell already knew its fate just needs a signal to tell it to assume this fate
- Red signal permits cells to respond to previously inherited cell fate determinant
Requirements of a morphogen - act directly at a distance
Red ligand from red cell
- One signal does everything
Bucket brigade – not used by morphogens
- Each signal induces another
What are the two ways to distinguish between instructive and permissive signals
Provide a second source of red signal
Provide red signal at a uniform concentration
What happens if you provide a second source of red signal?
Instructive signal – get a mirror image patterned effect
Permissive signal – no effect
Experiments often done in chick wings – get a mirror image
What happens if you provide a red signal at a uniform concentration?
Instructive signal – one cell fate induced
Permissive signal – no effect
What are the two ways to distinguish between one signal and bucket brigade?
Genetically engineer proposed morphogen into a juxtacrine
Make a genetic mosaic that lacks the receptor for the red signal in one of the cells
What happens if you genetically engineer proposed morphogen into a juxtacrine?
Add a transmembrane domain – signal cannot diffuse
One signal – only neighbouring cell receives signal
Bucket brigade – no effect
What happens if you make a genetic mosaic that lacks the receptor for the red signal?
One signal – cell without receptor does not receive signal and does not differentiate
Bucket brigade – no effect (unless green cell lacks receptor)
What establishes a shallow morphogen gradient?
Passive diffusion
What establishes a steep morphogen gradient?
Binding to molecules in the ECM (heparan sulphate proteoglycans) and high concentrations of receptor - restricted diffusion
Rapid degradation of signal in the ECM
Where are heparan sulphate proteoglycans found?
Found in the ECM and bind to many ligands
How do HSPGs regulate morphogen diffusion?
Sequestration or slowing diffusion – e.g. BMP - TGFbeta
Facilitating diffusion - Hh
What does planar transcytosis have a role in?
Role in establishing some morphogen gradients
Planar transcytosis method
Pit forms in cell membrane and engulfs the morphogen in a vesicle
Repeated cycles of endocytosis and resecretion allows certain morphogens to travel through cells
Evidence for transcytosis in Dpp (TGFbeta) signaling
Antibody staining shows that Dpp is found in vesicles
Mutations that block vesicle formation cause Dpp to act in a juxtacrine manner
What is the role of timing in establishing morphogen gradients?
As the gradient is established, gene expression is changing
- Must be a mechanism to block premature specification
- Cell waits for the steady state of receptor activation to be achieved - molecular mechanism is poorly understood
What is the transcriptional read out model used for?
Used to model how cells read or interpret a gradient to make a cell fate decision
Transcriptional read out model
Higher concentration of morphogen = higher concentration of activated transcription factor
Receptor activation causes TF to enter the nucleus and direct transcription
- The same TF in every cell
- Concentration of 3 TF per nucleus results in green cell fate decision
What is bicoid?
Is a morphogen and transcription factor
Where is bicoid mRNA localised?
mRNA is localised at the anterior of the egg and is translated into protein during embryogenesis
What happens to bicoid protein?
Diffuses through the cytoplasm and accumulates in nuclei of the syncytial blastoderm -generates a concentration gradient
How is transcription factor concentration interpreted at the DNA level?
Enhancers that regulate green cell fate genes have lower affinity for TFs than those of blue cell fate genes
Transcriptional activation in blue cell
Both sets of genes see the same medium levels of TF
However only high affinity enhancers can bind enough TF to activate gene expression
Blue cell fate genes are activated - high affinity
Green cell fate genes are inactive - low affinity
Transcriptional activation in green cell
Green cell has high levels of TF in the nucleus
Both high affinity (blue) and low affinity (green) enhancers are activated
Why doesn’t the green cell have blue characteristics?
Block blue gene expression by having one of the green cell fate genes encode a repressor – crosstalk regulation
How are strict thresholds achieved when the gradient is not steep?
One of the blue genes encodes a transcription factor that activates its own expression – positive feedback regulation
