Drosophilla appendage development Flashcards
What are the different types if Drosophila appendages and what causes defects in them?
Wings, legs, antennae and halteres (balance organs). All defects in limb developments in Drosophila (and vertebrates) are due to defects in the Hh signalling pathway.
WHen does limb development occur in flies?
During metamorphosis from larvae to adult fly. Most adult structures come from the imaginal discs in the larvae. These discs are set aside during embryogenesis. Different discs are found in different sets of the larvae.
What is the structure of the wing imaginal disc?
The wing imaginal disc starts as a 2D epithelium of about 30 cells. It undergoes replication and proliferation so that by the 3rd larval instar stage it contains about 50,000 columnar epithelial cells. However, not much patterning occurs yet. This happens via signal transduction pathways and morphogens. Eventually, the wing becomes split into 4 quadrants. Dorsal, ventral, anterior and posterior.
How does the imaginal disc change shape to form the wing - which bits become which?
The central part of the wing disc becomes the blade, the surrounding region becomes the hinge. The disc unfolds into its adult shape via a process known as eversion. The future wing blade starts iff as 2D, it pinches up and moves out into the 3rd dimension to become the blade. This means the cells that were surrounding the future wing blade are all at one end (the hinge)
How are the axes of the wing established
Wingless is expressed along with the AP axis (horizontally) and Patched is expressed along the DV axis (vertically). Nubbin is expressed in the wing pouch (middle of the disc). The wing pouch moves out along the horizontal line and becomes the wing blade. The sensory organ precursors express Scarbrous and are seen in the specific places in the wing disc e.g. theres a double layered row of it located just adjacent to Wg expression. The vertical line is established by morphogen signalling mechanism, like Hh and Dpp. It is this vertical line of expression that actually patterns the AP axis because it causes a different expression along that way
How is the initial difference in anterior/posterior expression in the wing disc started?
It starts when the 1st imaginal discs appear in specific segments of the embryo. The engrailed TF defines the posterior compartment of the segment. You can see that where engrailed expression occurs, wing discs develop. Therefore, presumptive imaginal discs inherit their AP identity from the segmentation machinery. Engrailed is a nucelar protein (not a signalling molecule in its own right) But can be visuallised ysing en-LacZ reporter gene. So, you incorporate the Lac-Z gene in place of the engrailed protein but under the control of engrailed enhancers which care controlled by the segmentation system - so they’re still expressed in the same place. You then convert the lacZ from a soluble colourless molecule into x-gal which is an insoluble blue precipitate. Therefore, this shows that compartments in the developing fly are defined by differential cell affinities. So we now have a difference in the A and P, but more info is needed to be able to tell the cells exactly where they are along the axis, this is where Hh comes in.
Explain how Hh tells the cells what position they’re in - what is its transduction pathway?
Hh has a signal transduction pathway - it is controlled by 2 receptors. Ptc and Smo. When Hh is present - Ptc no longer inhibits Smo - meaning that Smo can cause the production of a molecular complex that allows for Ci to act as a gene activator. It activates things like dpp Wg and En. So overall, engrailed suppresses the expression of Ci. En drives expression of Hh. Therefore Ci is only expressed in the absence of En. Ptc is the Hh receptor and pathway target gene. Hh levels are high in the posterior, but a bit also diffuses into the anterior, making a bit of a gradient - these cells in the anterior, that have Hh, Ptc and Ci (because there’s no En) can carry out the pathway in the diagram (look at notes) so are the only cells that transduce the genes - one of which is Ptc itself. So, these cells can increase their Ptc. This is what creates the line up the middle of the wing disc. En prevents posterior cells from responding to their own Hh. cells in the anterior compartment can respond to Hh made next door, but can’t make their own because that requires En.
Why is the line of expression up the middle of the wing disc made of dpp and not Ptc?
Dpp is a transcriptional target of Hh signalling. I.e. Hh increases expression of dpp (dpp is the fly version of TGFb) So in the same place the cells are making Ptc because of responding to Hh, they’re also making dpp. This dpp moves to both the anterior and posterior, forming a morphogen gradient which tells the cells where they are in relation to the midline.
What are the properties of dpp? what does it signal?
Dpp is an extracellular ligand. It interacts with Type I and type II receptors which form a heterodimer when dpp binds. (in drosophilla these receptors are called Tkv and Pnt) this activates a TF called mad, which when in its active form is phosphorylated and acts on target genes brinker (which it inhibits) and omb and sal (which it activates). Dpp can be detected at a distance from its expression using dpp-lacz (in the same way as engrailed) High concentrations of Dpp induce sal (aka spatzle) and omb expression, whereas lower levels of Dpp induce only omb. This allows wing structure to be patterned and for growth to occur. (look at the thought experiment in notes if you’ve got time bc I think it might come up)
