Lecture 15: vertebrate limb development Flashcards
Describe the morphology of a limb bud.
The limb buds arise from the flank of the embryo body wall. They are composed of mesoderm and an outer cover of ectoderm.
The tip is called the apical ectodermal ridge and the core is a lose mesenchyme made from the lateral plate mesoderm and the somites.
The apical ectodermal ridge is characterised by the thickening of the ectoderm and by the expression of a range of FGFs including FGF8 and FGF10.
Behind the AER is the progress zone where cells are highly proliferative.
The zone of polarising activity is a small area in the posterior mesenchyme which expresses Shh to instruct the limb of A-P activity.
Describe the different axes of the limb.
Proximal-distal = shoulder-finger tips Anterior-posterior = thumb-little finger Dorsal-ventral = back of hand-palm
Describe the genes expressed in the hind and forelimb.
Hox genes establish a permissive environment for RA to be synthesised on the lateral plate mesoderm mesenchymal cells. RA induces the T-box TFs.
The forelimbs express Hox5 and 6 which control the expression of Tbx5.
The hindlimbs express Hox8, 9 and 10 which control the expression of Tbx4.
What kind of TFs are Tbx genes?
T-box TF. They have a DNA binding motif called the T domain that binds DNA in a sequence-specific manner.
Tbx4 and Tbx5 control the production of FGF that initiates limb development.
Explain the Tbx4 and Tbx5 downstream pathways and their role in establishing two regions of the limb bud. Draw the pathways.
Tbx4 and Tbx5 control the production of FGF that initiates limb development. They establish and maintain the apical ectodermal ridge, important for limb outgrowth and patterning, and the mesodermal polarising region, needed for A-P patterning of the limb.
In the forelimb: RA induces TBX5 which is involved in a feedback loop, with the expression of FGF10 which result in induction of wnt3a which induces fgf8 in a particular area of the ectoderm, important for establishing the proximal-distal axis.
In the hindlimb: TBX4 is involved in a feedback loop, with the expression of FGF10 which results in the induction of wnt3a which induces fgf8 in a particular area of the ectoderm, important for establishing P-D axis.
What are the two models of proximal-distal patterning. Which one is most likely true and why?
- Progress zone model: the length of time that a cell spends proliferating in the progress zone dictates the identity of the cells they give rise to. Early removal of the AER = cells don’t spend much time proliferating = small protrusion. This is not likely because cells found early in development are already fated to develop in the limp.
- Two-signal model (current theory). Antagonising signals of FGF (from distal) and RA (proximal) establish various areas of the limb. This is likely because RA and FGF show precisely these activities. This is also a model conserved with Drosophila.
Explain the result of anterior transplantation of the ZPA.
Transplant in anterior of host embryo => animal has a mirror image duplication.
There is a single signalling molecule that causes this = Shh. It is expressed in the ZPA.
Transplant Shh cells instead of entire ZPA produced the same results.
Shh has the ability to diffuse, important for establishing identity of muscles, bones etc.
Explain the process of limb A-P patterning.
- ZPA expresses Shh (morphogen)
- Shh diffuses to establish bone/muscle identity (P→A)
- Closest to ZPA = highest level of Shh = posterior structures
- Time of exposure and concentration of Shh establishes different cell fates
- Digits 2, 3, 4 and 5 depend on timing of Shh exposure. Digit 1 is too far away so develops independently of Shh
- Feedback loops between ZPA (Shh) and progress zone (FGF) for coordination of A-P and P-D patterning
Explain the result of a Shh KO.
Loss of identity in the zellgopods (ulna/radius).
Explain the process of D-V patterning. Draw the pathway.
Not much is known about this but Wnt7a is expressed in the dorsal ectoderm which induces Lmx1b (homolog of drosophila apturus), and engrailed is expressed in the ventral ectoderm (default pathway).
What is the result of a Wnt7a KO?
Loss of dorsal identity.
