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BMS237 - Advanced Developmental Biology > Lecture 19 > Flashcards

Lecture 19 Flashcards

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1
Q

How does the pathway of wing development in Drosophila differ from that of it vertebrate counterparts, particularly, in the way in which the Lmx1b homologue acts

A

Apterous acts upstream of the wnt7a homologue in Drosophila. Apterous acts to upregulate the expression of wingless leading to the adoption of a dorsal cell fate

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2
Q

From medial to lateral, recall the mesodermal tissues in the developing chick embryo

A

Axial mesoderm, Paraxial mesoderm, intermediate mesoderm and lateral mesoderm

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3
Q

As the limb grows its specifies the skeletal element that the cells adopt depending on how long they spend proliferating in the progress zone, T or F

A

T

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4
Q

Explain the interaction between tbx4 and 5 that act to restrict the limb fields

A

There is a negative regulation mechanism between the tbx 4 and 5 genes. Tbx5 can block the expansion of tbx4 expression which prevents the hindlimb field from progression anteriorly into the forelimb field

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5
Q

Which two genes show an exquisite complimentary expression pattern with one being uniquely expressed in the dorsal ectoderm whilst the other in the ventral ectoderm

A

Engrailed1 is only expressed in the dorsal ectoderm whilst wnt7a is expressed in the ventral ectoderm

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6
Q

Homothorax is the Drosophila homologue of Meis1/2. What is the importance of this gene in fruit fly development

A

Homothorax is important in controlling the patterning of the proximal wing

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7
Q

What is the result of lmx1b knockout

A

Lmx1b knockout results in a ventralised limb with no dorsal mesenchymal development

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8
Q

FGF was considered to have a role in formation of the limb bud. Describe an experiment that could be carried out in chicks to determine if indeed, FGFs are involved in limb formation

A

Implantation of FGF-soaked beads into the flank of chick embryos and observe if ectopic limbs develop

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9
Q

It was later found that ectopic grafting of the ZPA resulted in a mirror image duplication of autopod structures. What was seen in a similar experiment where grafting occurred earlier in limb development

A

There was also a duplication of the ulna

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10
Q

Describe how FGF10 expression in the lateral mesoderm at the level of the limb is induced by the intermediate mesoderm

A

The intermediate mesoderm produces large amounts of FGF8 which induces the FGF10 expression by the lateral mesoderm

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11
Q

Chick wings have three digits, describe the nomenclature of these digits

A

The chick wing has three digits named digit 2,3 and 4. There is no digit 1

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12
Q

Why does digit 1 often develop to some extent in the limbs of sonic hedgehog knockout mice

A

The anteriormost limb, or digit 1 forms independently of sonic hedgehog signalling. Ihh expression is seen in the most anterior region of the limb bud and acts independently of sonic hedgehog to dictate some autopod development. Ihh maintains the expression of sonic hedgehog target genes in the anterior region of the limb bud in the absence of sonic hedgehog hence leading to the digit development seen in sonic hedgehog -/- mice. Similarly Ihh -/- mice did not show development of the anteriormost digit

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13
Q

How does sonic hedgehog dictate digit formation in the chick wing

A

Each threshold of sonic hedgehog correlates with the formation of a specific digit. At the lowest sonic hedgehog concentration digit 2 forms. Intermediate sonic hedgehog concentrations correspond to digit 3 with the highest sonic hedgehog concentration dictating digit 4

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14
Q

Below is a list of limb bone structures and their embryonic precursors regions. Match up the corresponding bones and embryonic structures. A - humerus, B - Metacarpals, C - Radius, D - Carpals, E - Phalanges, F - Ulna. i - Stylopod, ii - Autopod, iii - Zuegopod

A

A - i, B - ii, C - iii, D - ii, E - ii, F - iii

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15
Q

The temporal model for patterning the vertebrate limb was determined to be incorrect, T or F

A

T

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16
Q

Which genes are mainily involved in proximo-distal patterning of the limb

A

RA and FGF

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17
Q

How does the chick forelimb differ to its hindlimb

A

The chick wing only has three digits whereas the hindlimb or leg has 4. The leg has digits 1,2,3 and 4

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18
Q

Recall the pathway of interaction between the genes involved in the d-v patterning of the vertebrate limb

A

BMP –> En1 –| Wnt7a –> Lmx1b

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19
Q

The key genes involved in specification of the limbs were determined to be, FGF8 and 10, Retinoic acid, Hox genes and tbx transcription factors. Put these genes in the sequence in which they act to dictate limb field development

A

Hox genes –> retinoic acid –> tbx4/5 –> FGF10 –> FGF8

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20
Q

What the name of the structure that forms in the posterior limb bud that expresses the sonic hedgehog morphogen and plays a key role in patterning of the limb

A

Zone of Polarising Activity (ZPA)

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21
Q

Which tbx gene is responsible for specifying the posterior or hindlimb bud

A

Tbx4

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22
Q

Explain how interactions between the two secreted signals in vertebrate limb patterning act to form the stylopod

A

The stylopod is the most proximal region of the limb. This is the regions that’s is exposed to the highest levels of RA. Retinoic acid acts to induce the expression of Meis1 and 2

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23
Q

Describe an experiment that could be carried out in chicks that would provide the evidence that the ZPA controls anterior-posterior patterning in the limb. What would you expect to see

A

Surgical removal of the ZPA from a donor chick embryo and implantation into the anterior limb bud of a donor embryo. This grafting would cause a mirror image duplication of distal limb structures

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24
Q

Describe the phenotype of a sonic hedgehog mutant with regards to the effects on limb development

A

Sonic hedgehog knockout mice exhibit a complete loss of the distal most skeletal elements as well as a loss of identity of the zeugopods

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25
Q

Which mesodermal region produces FGF10

A

Lateral mesoderm

26
Q

Explain how FGF10 signalling results in the induction of the AER

A

FGF10 from the lateral mesoderm acts to induce wnt3a expression in the overlying ectoderm which changes its identity to become thickened, producing the apical ectodermal ridge

27
Q

Hox11, Hox 13 and Meis1/2 are all downstream genes upregulated in the limb bud that act to pattern the vertebrate limb. What type of genes are these

A

Transcription factors

28
Q

Hox genes directly induce the expression of tbx transcription factors in the process of determining the limb field regions, T or F

A

F – Hox proteins lead to the expression of retinoic acid which in turn upregulates tbx transcription factors

29
Q

What experimentation methods could be employed to determine if sonic hedgehog is acting as a morphogen in anterior-posterior patterning of the distal limb

A

Carry out in situ hybridisation to look at where the sonic hedgehog mRNA is being transcribed in the limb bud. This would show a localisation of the mRNA in the cells of the ZPA. This could then be compared to immunohistochemistry experimentation which would stain for the sonic hedgehog protein using tagged antibodies. This would show a decreasing gradient of the protein from posterior to anterior throughout the limb bud and not localised to cells of the ZPA. This indicates that the sonic hedgehog protein is leaving the cells in which it is produced

30
Q

What is meant by the progress zone and what is its role

A

The progress zone is a region of mesoderm where cells proliferate and that will ultimately give rise to cells of the limb

31
Q

The anterior border of Hox6 expression was found to correlate with limb bud formation. As Hox 6 is a transcription factor, what type of experiment would be used to prove that at the anterior border of its expression, the limb fields would form

A

In situ hybridisation – identify the cells where the Hox6 mRNA transcript is produced

32
Q

Explain the role of lmx1b in patterning of the vertebrate limb

A

Lmx1b acts to pattern the dorsal region of the developing vertebrate limb

33
Q

Which genes are mainly involved in anterior-posterior patterning of the limb

A

Sonic hedgehog (and Ihh)

34
Q

Outline the role of the AER in limb patterning

A

The AER is required and necessary for the growth and maintenance of the progress zone

35
Q

What signalling acts to restrict expression of FGF10 in specification of the limb fields

A

Wnt signalling

36
Q

Explain how interactions between the two secreted signals in vertebrate limb patterning act to form the autopod

A

The autopod is the most distal region of the limb. This is the region that’s is exposed to the highest levels of FGF signalling. FGFs act to induce the expression of Hox13

37
Q

Describe a similar experiment to ZPA grafting the proves its role in anterior-posterior patterning of the distal limb based on the knowledge of the molecules that it secretes

A

Implantation of sonic hedgehog-soaked beads into the anterior limb bud. This should also create a mirror image duplication of distal structures

38
Q

What are the three genes involved in dorso-ventral axis formation in the developing limb

A

Wnt7a, engrailed1 and lmxb1

39
Q

Which tbx gene is responsible for specifying the anterior or forelimb bud

A

Tbx5

40
Q

Where is lmx1b expression only seen in the vertebrate limb

A

In the dorsal mesenchyme

41
Q

As the limb grows outwards during development, the more proximal skeletal and muscle regions are specified, T or F

A

F – the more distal skeletal elements are specified as the limb grows outwards

42
Q

Explain how interactions between the two secreted signals in vertebrate limb patterning act to form the zeugopod

A

The zeugopod is the intermediate region of the limb. This is the region that’s is exposed to intermediate levels of both RA and FGF signalling. A combination of this signalling acts to induce expression of Hox11

43
Q

What is the effect of en1 knockout on vertebrate limb development

A

Engrailed knockout results in a dorsalisation of the limb as wnt7a expression isn’t inhibited. This is because wnt7a expression is driven throughout entire d-v axis ectoderm. En1 acts to prevent wnt7a expression from expanding ventrally

44
Q

Apterous is the invertebrate homologue of lmx1b. What is the significance of this in appendage development

A

Apterous is present in the Drosophila wing disc and controls dorsal fate by inducing wingless expression in Drosophila

45
Q

Distal-less (Dlx) is a gene found in Drosophila identified as playing a role in development of appendages, in particular, the leg. What is the significance of dlx in vertebrate limb development

A

Dlx is the Drosophila homologue of Hox13

46
Q

Explain how the complimentary genes involved in d-v pattering of the vertebrate limb interact

A

Engrailed1 prevents the expression of wnt7a.

47
Q

In the two-signal model of proximal-distal limb patterning, what are the signals involved

A

The distal signal from the AER are FGFs, whereas the proximal signal is retinoic acid

48
Q

Which region of mesoderm gives rise to the urogenital structures such as the kidneys

A

Intermediate mesoderm

49
Q

In vertebrate limb development, hox genes are expressed in the lateral and paraxial mesoderm, which ultimately leads to the expression of tbx4 and 5, T or F

A

T

50
Q

Outline the two-signal model of proximal-distal limb patterning

A

The AER is only important in establishing the distal area of the limb. Another signal acts to establish the proximal regions. Its the interactions between proximal and distal elements that determines the intermediate regions

51
Q

Explain the coordination between the developing anterior-posterior and proximo-distal axis development

A

The AER releases FGF8 which plays an important role in maintaining sonic hedgehog expression by the ZPA. Sonic hedgehog then in turn plays an important role in maintaining FGF expression in the AER. This positive feedback loop allows for a mutual maintenance of both sonic hedgehog and FGF expression

52
Q

After induction of the AER by FGF signalling from the lateral mesoderm and wnt3a expression, another set of FGF ligands are expressed at high levels. Which FGFs are these

A

FGF4 and 8

53
Q

What molecule is secreted by the ZPA to control anterior-posterior patterning of the limb

A

Sonic hedgehog

54
Q

The molecules that act to specify proximal and distal regions of the vertebrate limb are secreted products that act as morphogens, T or F

A

T

55
Q

Tbx4 and tbx5 were wound to be the downstream genes activated that specify the limb fields. What type of genes are tbx genes

A

Tbx genes are T-box containing transcription factors

56
Q

What is the ZPA and its role in limb development

A

The Zone of Polarising Activity (ZPA) is an area of mesenchyme in posterior limb bud that controls anterior-posterior limb patterning

57
Q

What are the names given to the structures that initially form and demarcate where the limb buds will form

A

Limb fields

58
Q

Explain the temporal patterning of the proximal-distal axis of the vertebrate limb

A

There is a clock present in the progress zone of the limb bud that measures the number of cell divisions. Cells that have undergone a small number of cell cycles will form proximal skeletal elements, whereas those that have undergone more cycles will form distal skeletal elements

59
Q

What set of genes dictate where the early limb regions are positioned

A

Hox genes

60
Q

It was later found that ectopic FGF did in fact lead to the development of an ectopic limb in the chick embryo. What is the significance of the proximity of this ectopic signalling to either of the normal developing limb fields

A

Depending of the proximity of the FGF signalling (due to bead implantation) to either the hind or forelimb fields determined the ultimate identity of the ectopic limb. If it was closer to hindlimb field an ectopic leg would develop, similarly, if it was closer to forelimb field an ectopic wing would develop

61
Q

What is the result of forced tbx4 expression in the forelimb field at the anterior limb bud in chicks

A

This will lead to the development of a leg (or hindlimb) instead of a wing (or forelimb)

BMS237 - Advanced Developmental Biology (32 decks)

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