L15 - Drosophila Limb Formation Flashcards
What is the fundamental principle RE patterning
To give a cell unique identity about 3 body axes
What are the axes of the limb
A-P
D-V
P-D
What are the 3 skeletal elements of the vertebrate limb
Stylopod
Zeugopod
Autopod
What bone makes up the stylopod
Humeris
What bones make up the zeugopod
Ulna and Radius
What bones make up the autopods
Carpals and digits
Describe the appearance of the limb buds
Appear as protrusions from the flank at precise locations along the AP axis
The region where the limb will form is known as
The limb field
Describe the effect of transplanting cells normally making up the limb fields at the flank
Growth of ectopic limbs
Describe the effect of transplanting cells that normally make up the flank at the region where the limb normally develops
No limb development in that region
Tbx5 can be seen expressed in the
Forelimb buds
Tbx4 can be seen expressed in the
Hindlimb buds
What is the effect of forces expression of TBx5 in the hindlib bud
Tbx4 is turned off
What is the effect of a Tbx4 KO
Still get hind limbs developing
What is driving the identity of the hind limb
Pitx1
Describe the effects of knocking out Pitx1
No hindlimb development and expression of Tbx4 is lost
What does Tbx mean
T-box
What specifiies the AP axis
Hox genes
Describe the evidence that Hox genes are able to act directly without the need for RA intermediate
HOXC6 boundary
Marks the boundary between the thoracici and cervical vert
Also markss the position where the forelimb will form
What is able to mimic the action of the limb field to induce ectopic development
FGF soaked bead
Describe the process of the intitiation of the limb
FGF8 produced from the intermediate mesoderm
FGF10 produced from the lateral plate mesoderm
Wnt2b restricts the domain of FGF10 expression
Wnt3a induced in the limb field
FGF8 then produced at the AER acts on the progress zone
What are the three regions of the early limb bud
Progress zone
Apical ectodermal ridge
Zone of polarising activity
What type of cells make up the progress zone
Primarily mesenchymal cells
What does the ZPA express
Shh
Removal of the AER shwos the the AER is nescersary for
The progress zone and limb growth
Describe the specificaiton of more distal skeletal elements
Specified as the limb grows
How many models are the for the P-D patterning of the limb
Name them
2
Progress zone model
Two signal model
Describe the progress zone model
Suggests that the AER is driving outgrowth of the limb
AER then acts on the progress zone
The longer a cell spends in the progress zone then the more distal a structure is formed
Describe the two signal model of limb PD patterning
Initial progenitors are present for all of the skeletal derivatives
Recieve antagonistic signals according to the positions they are along the gradient
In the two signal model what are the two gradients
Retinoic acid proximally
FGF distally
What structures form under hi RA lo FGF , what genes are first expressed
Meis genes
Stylopods
What gene is expressed at intermeidate levels of FGF and RA - what structures form here
Hox11
Zeugopod
What gene is expressed under the influence of lo RA and hi FGF what strcutures then form
Autopods
Hox13 first expressed
FGF is hi
Dorsally
RA is hi
Proximally
Describe what happens to limb patterning when you KO the enzyme required for the synthesis of RA
Lose expression of the meis gene and you lose the prominal structures
Describe the effect of grafting the ZPA into an anterior position
Mirror image duplication of the autopods
Describe what happens if the ZPA is grafteed at a very early stage
duplication of the ulna also
Describe how the action of the ZPA may be mimiced
Using a bead soaked in Shh
OR forcing Shh expression in cells which don’t usually express
Shh acts as a ____________
Classic morphogen
Describe how Shh acts as a classic morphogen
Digits form depending on the concentration of Shh that they see
Describe how mirror image duplication is also evidence for Shh acting as a morphogen
Now get X^2 curve - high either sides - low in the middle
Describe the Shh mutant limb
Complete loss of distal most skeletal elements
Causes the loss of identity of the zeugopods - R and U forms as a single bone
Describe what Shh from the ZPA acts on
Diffuses P–>A
Acts on the ZPA
Describe what FGF from the AER acts on
FGF4 diffuees back to the proximal
FGF8 acts on the ZPA
What are the three genes which have an essential role in dorso-ventral patterning
Wnt7a
En1
Lmx1b
Describe where Wnt7a is expressed
In the dorsal edge
Describe when En1 is expressed
In the ventral edge
Describe where Lmx1b is expressed
In the dorsal HALF
What is the drosophila homologue of Lmx1b
What does this control
Apterous
Controls the dorsal cell fate
Describe the effect of a conditional KO of Lmx1b
Ventralisation of the limb
Fur forms on both sides
All cells adopt a ventral identity
Describe the process for the formation of dorsal patterns
No BMPR so no En-1 No supression of Wnt7a Expression of Wnt7a Wnt7a induces the expression of Lmx1b Lmx1b diffuses into the dorsal segment Cell take a dorsal pattern
Describe the process for the formation of ventral patterns
BMPR induces expression of En-1
En-1 confers a ventral pattern to cells
En-1 supresses Wnt7a
What is the effect of En-1
Supression of Wnt7a
What is the default cell fate
Dorsal (must supress this fate with En-1)
Aer secretes …. and the drives …
FGF4/8 driving the outgrowth of the limb
How many modes for the outgrowth
Progress zone and two signals
ZPA secretes … which forms a gradient along which axis
Shh - AP axis
How are PD and AP axes coordinanted?
Through the reciprocal control between the AER and the ZPA
Sum up the DV axis patternign
Wnt7a and BMP from dorsal and ventral cotnrol Lmx1b
What will form in presence of lmx1b
Dorsal
