Lecture 16 - Chick embryo Flashcards
(17 cards)
What genes are expressed at HH24?
Shh, Grem1 & Fgf8
What happens at HH stage 23-24?
- somites which will give rise to the muscles
- Allantois - function is to store metabolic waste produced by embryo
What are areas of the brain?
Diencephalon is a division of the forebrain, between the telencephalon & the mesencephalon.
It differentiates into hypothalamus & pituitary.
Telencephalon contain cerebral cortex
Describe chick wing development
Full pattern of cartilage laid down by day 8
Describe a chick wing
It has 3 digits (anterior to posterior)
Digit 4&5 lost due to dinosaurs
Digit 1 - 2 phalanges
Digit 2 - 2 phalanges
Digit 3 - 1 phalanx
Describe a chick leg
It has 4 digits (anterior to posterior)
Digit 1 - 2 phalanges
Digit 2 - 3 phalanges
Digit 3 - 4 phalanges
Digit 4 - 5 phalanges
Describe how a limb bud is equipped with signalling centres
Shh is expressed in the ZPA (posterior)
Fgf8 is expressed in the AER (distal ‘membrane’ - periphery)
Wnt7a is expressed in the dorsal epithelium (entire inside)
Gremlin found in the mesenchyme (inside)
What was predicted about the ZPA?
Accidental discovery of the Zone of Polarizing Activity (ZPA) or polarizing region in the chick wing bud.
It was predicted that the ZPA produces a signal or morphogen which specifies AP position & digit identity.
What is the positional information model - French flag problem?
- Homogenous population of cells
- Concentration gradient of a morphogen forms over cells & at different thresholds gives cells a position
- Cells interpret their position & differentiate to form a pattern.
Mesenchymal cells (each cell can develop as blue, white or red - representing digit 1,2 & 3).
Higher concentration of morphogen posteriorly, low anteriorly. High concentration - digit 3, intermediate concentration - digit 2, low concentration - digit 1.
Cells now interpret their position, instructed by the concentration of morphogen they have received.
Genetic difference in receiving cells.
What is the affect of Retinoic acid?
Retinoic Acid bead induced digit duplication.
Retinoic acid, when applied to the anterior margin of chick wing bud, induced a new ZPA that produced a signal.
What is the morphogen thought to act in the ZPA?
Shh - a bead soaked in Shh protein or Shh expressing cells implanted to the anterior margin duplicates the pattern of digits.
When cells containing shh were grafted to anterior margin of chick wing bud, this resulted in a mirror image duplication of the digit pattern, similar to ZPA.
Shh could operate via other signals - e.g. BMPs
How does shh signalling operate?
Shh signalling operate through Gli transcription factors:
1 - No Shh signalling - Ptch1 inhibits Smoothened allowing Gli activators (A) to be processed into Gli repressors (R)
2 - Shh signalling - Shh relieves the Ptch1 inhibition of Smoothened preventing Gli activators being processed into repressors.
This results in a GliA/R gradient across the limb bud & activation of target genes including Gremlin.
Describe the 3 Gli genes
3 Gli genes - Gli3 most important in limb - shh results in gradient of Gli3 activator & repressor across the limb bud.
High levels of Gli3 activator & low levels of repressor are found at posterior-side of limb bud, where Shh levels are higher. This results in activation of genes, including Grem1.
The Gli3 repressor constitutively inhibits gene transcription in the absence of Shh - loss of repressor of Gli3 that allows gene transcription downstream of Shh signalling.
What is the role of Gremlin?
The Gremlin gene encodes a BMP antagonist. It is expressed in limb bud mesenchyme. It is known as the Apical Ectodermal Ridge (AER) maintenance factor (AERMF).
AER-FGFs are required for Shh expression.
Reinforces the principal that genetic signalling loops are important in Dev Bio
Gremlin expressed in middle of limb bud (yellow).
Gremlin inhibits BMPs. This allows AER/FGF expression which induces Shh, which induces Gremlin.
How does a ZPA graft induce Gremlin expression?
Quail ZPA placed on the anterior margin of chick wing bud. Quail ZPA induces ectopic Gremlin expression in anterior chick wing tissue, but Shh expression turned off, as it is expressed in a shorter duration in quail than in chicken.
Shh from quail ZPA duplicates pattern of Gremlin expression in chick wing bud (host chick wing tissue).
What signal does the AER produce?
Fibroblast Growth Factors are the main signal produced by the AER.
Elements were truncated when AER was removed. Can be rescued by a small bead soaked in FGF protein.
An FGF bead implanted into the flank of a chick embryo can induce an extra limb.
Ectopic limb had grown.
FGF signalling is sufficient to establish all the signalling centres & gene regulatory networks required for limb development.
Summarise chick embryo
- Shh specifies the pattern of digits across the A/P axis
- Shh induces Gremlin which maintains the AER that are essential PD (proximo-distal) outgrowth of the limb
- FGFs in the AER are required for Shh expression.
KEY POINT: signalling regions express mRNAs that produce secreted proteins, which operate together to control axial growth & patterning
