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BIOL 309 > Early Vert Development (4) > Flashcards

Early Vert Development (4) Flashcards

1
Q

In summary, what did we find with the animal/vegetal sandwich experiments?

A
  • endoderm is able to induce mesoderm
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2
Q

In summary, what did we find with the rescue of UV ventralized embryos?

A
  • dorsal endoderm (Nieuwkoop center) can induce dorsal mesoderm (Spemann’s organizer)
  • Wnt signaling was driving force
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3
Q

Given all the information, what can we conclude about the UV effect?

A
  • microtubule formation is blocked by UV
  • this causes GBP (and disheveled and wnt11 mRNA) to not make it to the right location
  • end result is a ventralized embryo
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4
Q

What is the Nieukoop center doing wrt organizer formation?

A
  • GBP, disheveled –> TF siamois –> TF Goosecoid
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5
Q

What is Goosecoid?

A
  • transcription factor that specifies the Sp organizer
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6
Q

What do we see with a GBP rescue experiment?

A
  • injecting GBP into a UV treated ventralized embryo results in only half the average dorsal/anterior index
  • this indicates that GBP is not suficient for a complete rescue of a ventralized embryo and something else is needed
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7
Q

How is beta-catenin normally in each axis?

A
  • ventral: beta-catenin degraded

- dorsal: beta-catenin stabilized

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8
Q

On the dorsal side, what happens to the siamois gene?

A
  • b-catenin proteins bind upstream to siamois gene
  • this activates transcription of siamois protein
  • siamois protein activates the goosecoid gene and goosecoid protein is transcribed
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9
Q

On the ventral side, what happens to the siamois gene?

A
  • Tcf3 proteins bind upstream to the siamois gene

- this represses the siamois gene and no siamois protein is transcribed

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10
Q

What contributes to goosecoid expression?

A
  • siamois protein (but does not produce strong enough goosecoid expression alone)
  • Smad2/4 from TGF-B signal
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11
Q

What is the TGF-B pathway that contributes to Goosecoid protein transcription?

A
  • Vg1 –> goosecoid
  • Vg1 –>Xnr1,2,3 (xenopus nodal-related) –> goosecoid
  • (VegT) –> derriere activin nodal –> Xnr1,2,3 –> goosecoid
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12
Q

What does goosecoid specify/activate?

A
  • noggin, chordin and follistatin

- these specify/activate XBF-2 and Zic-r that define neural fate

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13
Q

What concentrations lead to ventral endoderm?

A
  • high concentrations of VegT and Vg1 (all endoderm specified by this)
  • low concentrations of B-catenin
  • nodal related low
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14
Q

What concentrations lead to dorsal endoderm?

A
  • overlap of VegT and Vg1 with b-catenin

- nodal related high

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15
Q

What is seen throughout the fully formed adult body?

A
  • high degree of diversification along axis (ex. brain, or bones in limb)
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16
Q

What is the Xnr gradient?

A
  • lower concentration on the ventral side and higher concentration on the dorsal side
  • within the endoderm and affects the mesoderm
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17
Q

Why is having a gradient advantageous?

A
  • allows for scale
  • different relations of response to signal (different response to low, medium or high)
  • different cell responses could lead to different fates
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18
Q

What are morphogens?

A
  • factors that organize a field of surrounding cells into patterns
  • very important mechanism in development
19
Q

What are the two criteria to define a morphogen?

A
  1. form a gradient of concentration (usually emanating from a localized source)
  2. act on cells in a concentration-dependent manner (i.e. different concentrations will lead to a different response from the same cell)
20
Q

What upregulates Xnr?

A
  • GBP and disheveled upregulate Xnr

- this is why it has a higher concentration in the dorsal region

21
Q

What is the factor in mesoderm? What is it’s gradient and why?

A
  • BMP4 in mesoderm
  • higher concentration in ventral region and lower in dorsal
  • because GBP and disheveled produce Xiro which inhibits BMP4
  • morphogen gradient in mesoderm
22
Q

What information do morphogen gradients give? How does this apply to BMP4?

A
  • “positional information”
  • high BMP4: ventral mesoderm
  • intermediate BMP4: intermediate mesoderm
  • low BMP4: dorsal mesoderm
23
Q

What are key experiments that need to be performed in order to demonstrate that a molecule functions as a morphogen?

A
  • can test the affect of different concentrations

- add more or knock down and then add back in different concentrations

24
Q

What experiment can be used to test whether BMP4 functions as a morphogen in the mesoderm?

A
  • test different levels of BMP4
  • assay: score for tissues
  • control: no BMP4 or simply a normal embryo
25
What exactly was done to examine BMP4?
- injected different amounts of BMP4 RNA to all cells - removed dorsal lip and cultured in isolation - examined what type of mesoderm was being formed
26
What tissues result for different amounts of BMP4?
``` (dorsal/low) - notochord - muscle - pronephros - blood (ventral/high) ```
27
Do morphogen gradients pattern the ectoderm during neural induction?
- yes/probably | - many different domains created by morphogens
28
What is the "einsteck" experiment?
- dorsal lip transplant | - remove dorsal lip from donor and put into host
29
What are the results of the "einsteck" experiment?
- depending what stage the dorsal lip is at (young or advanced), it can induce different axial regions - young dorsal lip: extra head - advanced dorsal lip: extra tail
30
What results from a transplantation experiment using mesoderm at the neural plate stage?
- take different sections of mesoderm and transplant to early gastrula - induce different axial regions depending on which part of mesoderm taken - indicates that mesoderm is not uniform in the types of tissues produced
31
What hypothesis does the mesoderm transplantation experiment lead to?
- hypothesis: there are molecules that act along the A-P axis in a graded manner - signals from underlying mesoderm (archenteron roof) instruct the neural plate in a graded manner
32
What did researchers find when searching for differentially-expressed mesodermal genes along the A-P axis?
- cerberus phenotype (named after three headed dog)
33
What does injection of cerberus into the vegetal blastomeres result in?
- ectopic head structure but no secondary axis
34
Where is cerberus expressed?
- anterior most endomesoderm | - first tissue that involutes in the organizer
35
What factors does cerberus act on?
- inhibits nodal, BMP4 and wnt
36
What results from a cerberus knockdown?
- head region is not lost following a cererus knockdown | - indicates there is functional redundancy
37
What other factors were found to be expressed in dorsal endomesoderm?
- looking for functional redundancy with cerberus | - found Dickopf (thick headed) and Frzb
38
What resulted from Dickkopf knock-down?
- diminished head formation
39
What resulted from Frzb gain-of-function experiments?
- increases head size
40
What was Frzb found to do molecularly?
- antagonizes wnt pathway by binding to wnt and blocking receptor
41
What do cerberus, Frzb and Dickkopf have in common? What hypothesis results?
- all expressed in head (anterior) - all inhibit wnt - wnt signaling is required for posterior (trunk) formation and is inhibited during anterior (head) formation
42
What might happen if wnt is overexpressed in endomesoderm?
- may override wnt inhibition - may form more tail structure - result: over-expression of wnt8 in dorsal mesoderm leads to loss of head structures
43
Hypothesis: Does wnt function as a morphogen on neural tissue? What experiment was done to test this?
- separate neural tissue from other influences to test wnt on it - isolate ectoderm: on own becomes skin - dissociate ectoderm and re-aggregate: becomes neural - assay: use RT-PCR to identify genes in specific areas
44
What genes were found to be expressed in the previous experiment?
``` (Anterior) - forebrain: Bf1 - forebrain+midbrain: Otx2 - hindbrain: Krox20 (posterior) ```

BIOL 309 (14 decks)

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