Organogenesis (Limbs and Eyes)

Question 1

1. What are the imaginal discs?
2. When are the imaginal discs set aside
3. What do the imaginal discs go through during metamorphasis?
4. Which genes control what is produced by each imaginal disc?
5. Name 2 TFs that repress the formation of the body wall so that the appendages can form?
6. From what do the imaginal discs inherit patterning from?

Answer 1

1. pockets of collumnar epithelial cells that form various structures in the adult.
2. in late stage embryogenesis following segmentation
3. extensive
4. Hox genes
5. elbows and no-ocelli
6. from the segments from which they are derived.

Question 2

cells at the centre of the appendage imaginal discs become what?

Answer 2

the most distal parts of the appendage

Question 3

1. what acts as the master regulator of AP patterning of the wing imaginal disc?
2. How does it act to pattern the disc?
3. What acts downstream of dpp?

Answer 3

1. engrailed
2. It is expressed in the posterior compartment of the wing disc and induces Hh expression here too. Hh is secreted and induces dpp at the anterior boundary in a thin strip
3. dpp is secreted. It activates the Pnt receptor, leading to the phosphorylation of the Mad TFMad activates Omb and Sal, depending on the dpp concentrationHigh [dpp] induces both Omb and SalLow [dpp] induces just Omb

Question 4

1. what is the master regulator for DV patterning of the wing imaginal disc?
2. In which compartment is the master regulator expressed
3. how does it pattern the imaginal disc?

Answer 4

1. apterous
2. dorsal compartment
3. the boundary between the DV compartments leads to the expression of Wg. Wg is secreted and acts in a gradient to induce DV genes at different thresholds.

Question 5

1. How does AP patterning of the Leg imaginal disc different to that of the wing?
2. in which region is dpp expression restricted to?
3. in which region is wg expression restricted to?
4. what is expressed in the region where dpp and wg are present at high concentrations? What region of the imaginal disc is this?
5. what concentrations do cells at the periphery usually recieve? What is induced in these cells?
6. What is expressed at the boundary of Q4 and Q5?

Answer 5

1. both dpp and wg is expressed at the anterior boundary
2. dorsally
3. ventrally
4. distalless
5. A high/low/medium concentration depending upon their position. Homothorax is induced
6. Dac is expressed at the Dll/Hth boundary.

Question 6

1. Describe the structure of the limb buds
2. What 2 TFs specift limb bud identity?
   
   • Which one is expressed in the forelimb bud?
   • which one is expressed in the hindlimb bud?
3. How do Hox genes control the positional expression of these TFs?

Answer 6

1. A core of loose mesenchymal cells from the lateral plate mesoderm and an outer layer of ectodermal epithelial cells
2. T box TFs - Tbx4 and Tbx5.

Tbx5 is expressed in the forelimb bud

Tbx4 is expressed in the hindlimb bud

3. synthesis of a retinoic acid gradient.

Question 7

1. What are the 2 main organising regions of the limb bud?
2. What is the role of FGFs in the initiation of limb formation
3. Describe the processes of how the 2 organising regions are induced.

Answer 7

1. the apical ectodermal ridge and zone of polarising activity
2. to induce and maintain the 2 organising regions
3. FGF8 is induced by the T-box TFs in a thin strip of intermediate mesodermFGF8 induces FGF10 in the adjacent lateral plate mesodermFGF10 causes the adjacent surface ectoderm to thicken and to express Wnt3a.Wnt3a induces FGF8 expression in the surface ecoderm which specifies the AERShh expression is induced in the posterior limb bud. This specifies the ZPA.

Question 8

1. Name the 3 PD regions
2. what does early removal of the AER result in?
3. what does late removal of the AER result in?
4. what does later removal of the AER result in?
5. What conclusions can be drawn from this experiment?
6. Describe a model for PD patterning.

Answer 8

1. Stylopods (humerus/femur)Zeugopods (radius and ulna/tibia and fibula)Autopods (carpals/tarsals, metacarpals/metatarsals and phalanges)
2. formation of the stylopods; partial development of zeugopods
3. formation of the stylopods and zeugopods
4. formation of the stylopods and zeugopods; partial development of autopods
5. the time spent in the progress zone dictates their PD identity
6. Antagonising activities of RA from the proximal regions and FGFs/wnts from the distal regions form opposing gradients which at different thresholds induce different PD genes.

Question 9

1. What is the AP axis patterned by and where is it produced from?
2. KOs of this gene have what phenotype?
3. How does this signal pattern the AP axis?

Answer 9

1. Shh produced from the ZPA
2. 1 zeudopod and 1 anterior most autopod (which usually develops independently of Shh)
3. It produces different combinations of Gli activators and repressors along the AP axis to determine the digit pattern. (each digit is determined by a specific combo of repressors and activators)

Question 10

1. What is DV patterning essential for?
2. What is expressed in a discrete band in dorsal ectoderm? ventral ectoderm?
3. What is the ventral ectoderm molecule induced by? and what does it inhibit?
4. What does the dorsal molecule induce? What is thsi a homologue of? Why does the expression of this molecule define the DV axis?

Answer 10

1. the specification of muscles, tendons and nerves
2. Wnt7a | En1
3. En1 is induced by BMPs and it inhibits Wnt7a expression in ventral regions
4. Lmx1b. It is a homologue of apterous. Because it is only expressed in the dorsal half of the limb bud.

Question 11

1. from what structures do drosophila eyes develop?
2. the drosophila eye is a _____ eye
3. what does each ommatidia contain (7 cell types)
4. what is the role of the dark pigment of the ommatidia?

Answer 11

1. imaginal discs
2. compact eye
3. 8 rhabdomeres/photoreceptors | 4 cone cells | 11 pigment cells | sensory bristle | neuron | socket | sheath
4. provides light insulation

Question 12

1. which gene is necessary and sufficient to make an eye?
2. what is the name of its vertebrate homologue?
3. what is the phenotype of Q1 mutants?
4. What are Q2 mutations responsible for
5. ectopic expression of Q1 or Q2 leads to what?

Answer 12

1. eyeless
2. pax6
3. they lack eyes
4. human aniridia
5. ectopic eye formation

Question 13

1. which rhabdomeres are the inner rhabdomeres
2. which rhabdomeres are the outer rhabdomeres
3. how are the rhabdomeres organised?
4. what does the equator divide?
5. Why do ommatidia either side of the equator produce mirror images?
6. what area of the brain do outer photoreceptors project to; inner photo receptors?

Answer 13

1. R7 & R8
2. R1-R6
3. in a hexagonal crystaline lattice structure
4. D and V ommatidia
5. because they are rotated by 90^o in opposite directions
6. lamina | medulla

Question 14

1. what is the differentiation of rhabdomeres triggered by?
2. which ommatidia are the first to differentiate? Why?
3. describe the proliferative state of cells before, during and after their encounter with the trigger
4. describe the order in which rhabdomeres differentiate (5 stages)
5. the restriction of which gene marks differentiation. Where is its expression restricted?
6. how does the restriction of these gene occur in an evenly spaced pattern?

Answer 14

1. a morphogenetic furrow
2. posterior ones. As the morphogenetic furrow moves in a posterior to anterior direction
3. cells anterior to the furrow divide asynchronouslycells within the furrow do not dividecells that leave the furrow undergo 2 synchronous divisions before differentiating
4. R8 | R2&R5 | R3&R4 | R1&R6 | R7
5. atonal. restricted to R8 cells
6. lateral inhibition via notch. R8 expresses a higher level of notch leading to increased delta expression, leading to an increased repression of R8 identity to its neighbours.

Question 15

1. what secreted molecule is responsible for recruiting rhabdomeres? In which rhabdomeres is it expressed? Which rhabdomeres does it recruit?
2. cells that express Q1 also express what? What is the role of this molecule?
3. What is the phenotype of Boss and Sev mutants?
4. From which cells can Boss and Sev be lost without the loss of R7?
5. In which cells is Boss and Sev absolutely required for R7 recruitment?
6. Why dont other cells respond to boss in the same way as R7?

Answer 15

1. Spitz. Expressed in R8, R2 and R5. Recruits R3, R4, R1 and R6
2. Argos. It inhibits more distant cells from responding to Spitz
3. preference to white light (rather than UV light)
4. Outer photoreceptors
5. Boss is required in R8, and Sev is required in R7
6. because they have already been recruited.

Question 16

1. how does the optic vesicle form?
2. how does the lens placode form?
3. how does eye development occur?

Answer 16

1. evagination of the diencephalon
2. the extension of the optic vesicle to the surface ectoderm induces a thickened region of ectoderm
3. through reciprocal interractions between the optic vesicle and the lens placode.

Question 17

1. which part of the paraxial mesoderm maintain anterior identities of the neural tube?
2. this mesoderm produces signals which activate the expression of which 3 genes?
3. where are these genes expressed?
4. in which direction does the floor plate push as it is induced?
5. which signal mediates the bilateral splitting of the expression domain? What is this domain split into?
6. mutants of this signal have what phenotype?

Answer 17

1. prechordal mesoderm
2. Pax6, Six3 and Rx1
3. in a crescent pattern at the anterior end of the neural tube
4. anteriorly
5. Shh splits the anterior crescent domain into 2 oval domains
6. Shh mutants have 1 eye

Question 18

1. What does the lens placode do after it has been induced? What does this form?
2. What does this newly formed structure (Q2) tell the optic vesicle to do? What does this form?

Answer 18

1. invaginates and detatches from the surface ectoderm forming the lens vesicle
2. invaginate, forming the optic cup.

Question 19

1. what does the outer layer of the optic cup produce and eventually become?
2. what does the inner layer of the optic cup consist of, and what do these cells eventually become?
3. How do cells in the inner layer migrate out of this layer. Using what mechanism do they differentiate?
4. What are stem cells important for.

Answer 19

1. produces melanin, and becomes the retinal pigment
2. consists of a residual pool of stem cells which give rise to progenitors which differentiate into the cells of the neural retina
3. they migrate out of the layer in waves, depending upon the class of cell that has differentiated. They differentiate using a clock mechanism
4. adaptation of the eye to new environments (namely on birth; dark to light environment)

Question 20

1. on which side of the lens vesicle are proliferating cells found?
2. what do centrally found cells produce? What does this protein do?
3. Why are stem cells important in the maintenance of the lens?
4. Why is lens degeneration common in the elderly?

Answer 20

1. anterior side
2. crystalline proteins. They force the nucleus and mitochondria out of cells to make them truely transparent
3. because the cells lack the nucleus and mitochondria, they don’t last very long, thus frequent regeneration is required
4. stem cell pools become depleted over time thus lost lens cells can’t be replaced.

Question 21

1. what is the cornea?
2. what is the inner cornea layer formed by?
3. what is the outer cornea layer formed from?

Answer 21

1. a transparent epithelium that seals the front of the eye
2. mesenchymal neural crest cells that migrate to the anterior eye chamber
3. surface ectoderm adjacent to the lens that is induced by the lens placode.