Morphogens (L4) Flashcards
What is a morphogen?
A soluble, secreted molecules that act as a distance to specify fates of cells. A morphogen may specify more than one cell type by forming a conc. gradient.
Who suggested the idea of morphogens?
Alan Turing first suggested the concept of morphogens when he wrote a paper called the chemical bases of morphogenesis. Wolpert then expanded on the when he wrote positional information and spatial patterns of cellular differentiation. In the molecular age, (later than Turing and Wolpert). biologists cloned ligands that they thought might act as morphogens. E.g. a gene that when lost lead to no patterning. However, not all ligands involved in patterning are morphogens.
How do morphogen gradients work?
Morphogens can be produced and because they’re diffusable, they form a gradient. The conc. is higher at the source and lower at the sink. Cells along these gradients have different thresholds for the morphogen that cause them to become differentially fated cells. The morphogen gradient evolves over time due to the fact that the source may be producing more or other cells respond differently (e.g. by changing the number of receptors for the morphogen they have). If you change the gradient (e.g. make more from the course so the conc is higher, you can alter the pattern of cells that develop. If you’re making too much at the high end, you lose information here because more cells are receiving the same input.
What must a molecule be for it to be a morphogen?
For a morphogen to be a morphogen, it must induce different outputs at different concentrations. Morphogens are instructive signals because instruct cells to differentiate in a certain way. However, some signals may not be instructive, they may only be allowing cells that have already decided on a fate and are just telling it when to go (permissive factors) - perhaps from a previously inherited cell fate determinant. Permissive signals aren’t morphogens.
How can you test whether a signal is instructive or permissive?
To test whether a signal is instructive or permissive, you can ectopically implant to see if there’s a mirror image. The ectopically transplanted source will create bilateral symmetry due to 2 opposing gradients whereas a permissive signal will lead to the same pattern as in the wt because its concentration doesn’t matter, just that it reaches the cells to tell them to differentiate. For example, an ectopically transplanted bead of Shh put in a limb bud causes bilateral symmetry of the digits - showing that Shh is a morphogen. Another test you can do is to overexpress the signal everywhere and essentially dissipate that gradient. Again, this should have no effect on the pattern if it is a permissive factor. But if it is a morphogen, then all the cells will become the same as the one next to the source (where the highest conc is usually received). For a signal to be a morphogen, it also has to induce change from a distance, so only one source is needed
What is the significance of the signals being able to move?
For the gradient to be formed, the ligand has to be able to move. However, how do we know if the morphogen is diffusing and not just causing a domino of different signalling molecules through neighbouring cells (known as a bucket brigade) I.e. is it one signal or a chain?. To test this, you can tether the signalling molecule so it can’t diffuse, and is only available to the directly neighbouring cell, so it can differentiate the neighbour. If the system is a bucket brigade, then the differentiate of this cell would be enough to invoke expression in its neighbour, and so on. So you would get the same pattern. However, if it is not a bucket brigade, only the neighbour of the first cell would differentiate, and not the other ones because they are not receiving the morphogen (if you make it juxtacrine) Another way to test this is to make a genetic mosaic where some cells lack the receptor for the morphogen - again, a bucket brigade would be unaffected because the signalling molecule used is no longer specific to the starting molecule. Whereas, a morphogen model would cause the cells that lack the receptor to not differentiate.
Why is passive diffusion not enough to establish a good enough gradient? How is this overcome?
Passive diffusion would only generate a shallow gradient due to the morphogen being freely diffusible in all directions - so only a small fraction of the molecule diffused would actually go and affect the right cells, and its concentration would not be enough to get the detailed pattern required for the development. However, binding molecules in the extracellular matrix (e.g. heparin sulphate proteoglycans) and high concentration of receptor can generate a steep gradient. This is known as restricted diffusion. It enables the morphogen to diffuse but it keeps it in the right direction so makes the gradient stronger. Rapid degradation of the signal in the ENC may also generate a steep gradient, by making the lower conc lower, and means the high conc never gets too high and the cell can keep producing the morphogen without the area becoming saturated.
What are HSPGs?
Heparin sulphate proteoglycans. They are found in the extracellular matrix and are known to bind to many ligands. They are sometimes called co-receptors. They regulate morphogen diffusion by sequestering or slowing diffusion (e.g. BMP) or facilitating diffusion (e.g. Hh)
What is planar transcytosis?
Where pits form in the cell membrane and engulf the morphogen into a vesicle. Repeated cycles of exocytosis and resecretion allow certain morphogens to travel through the cell in a tissue e.g. Dpp. Helps play a role in morphogen gradients.
How can time play a role in establishing morphogen gradients?
Cells over time can change their receptor numbers to become more or less responsive. As the gradient is established, gene expression is changing with time. There must be a mechanism to block premature specification. The cell probably waits for the steady state of receptor activation to be achieved - but the molecular of this is not well understood.
What is the transcriptional readout model?
Higher concentrations of morphogen often result in a higher concentration of an activated transcription factor. In this model, receptor activation causes the transcription factor to enter the nucleus and direct transcription. It is the same TF in every cell - keep in mind these cells are initially identical. So, the amount of morphogen the cell receives is related to the amount of TF that is transported into the nucleus- however, sometimes the morphogen acts as both. E.g. bicoid is a morphogen and a TF. bicoid mRNA is localised at the anterior of the eff and translated into protein during early embryogenesis. Bicoid protein then diffuses through the cytoplasm and accumulates in nuclei of the syncytial blastoderm generation a concentration gradient (in Drosophila)
How is TF concentration interpreted at the DNA level?
Enhancers that regulate different cell fate genes have different affinities for the TF. Cells that receive a higher concentration of TF has a lower affinity for the TF than cells who receive low levels (they have a higher affinity). SO, at a medium concentration, the low-affinity genes (that would usually differentiate cells higher up the gradient) are unable to attract the TF due to their low affinity, but the high-affinity cells can so become the lower down gradient fate (all cells still possess the same gene so have the ability to become any of the fates). However, at the high end of the gradient, there is enough TF to not activate the low-affinity receptors - so why aren’t the high-affinity ones responding and cause a mixed up fate? Because when the lower affinity genes are expressed, they not only cause a higher gradient fate but make a repressor protein that blocks or turns off the high-affinity genes, meaning only 1 fate occurs. This is known as cross talk between cells
How are strict thresholds achieved when the gradient is not very steep?
Genes that have some TF binding to them positively feedback on themselves to turn themselves on more strongly. This also helps to turn off other genes due to increased production of repressors. Then other genes can negatively feedback on themselves to turn themselves off. Therefore, this causes genes to never be only partly on, they’re either very on or very off. This is what helps set the string distinct gradients.
