This exam consists of 7 pages and has a total of 50 points. Please make sure your name and student ID number are on all pages.

1. Provide concise definitions for each of the following (5 points)

a. ephrin:
 
 

b. Müllerian inhibitory substance:

 

c. silencer:
 
 

d. genomic imprinting:
 
 

e. cytoplasmic determinant:
 
 

2. An invertebrate embryologist friend of yours in Hawaii is studying a new species of ascidian. Although the fertilized eggs of this species have regions of brightly colored cytoplasm, she claims that embryos in this species do not have myoplasm that functions in the same way as that in other, "classic" ascidians, because they have a "red crescent" instead of a yellow one. You don't believe her theory, so you set out to disprove it. Describe one way you could you show that the red crescent functions like the yellow crescent in other ascidians (2 points).
 
 
 
 
 
 
 
 

3. Circle true (T) or false (F) for each of the following (6 points)

T       F	Lineage tracer injections have shown that individual neural crest cells are pluripotent
T       F	Alterations in the region of the Drosophila bicoid gene that encodes the 3' UTR of the bicoid RNA would be expected to alter the spatial distribution of bicoid mRNA during oogenesis.
T       F	Siamois is considered a candidate gene involved in differentiation of the Nieuwkoop Center on the ventral side of the early Xenopus embryo.
T       F	Alternative splicing of pre mRNAs does not lead to differences in the function of the  resulting mature mRNAs, since during splicing the introns are removed.
T       F	Roundabout is an important protein involved in axon guidance, because it helps regulate the crossing of neurons over the ventral midline of embryos.
T       F	The midblastula transition in Xenopus would occur earlier in an embryo derived from an oocyte to which a large excess of cytoplasm had been added.
T       F	Experiments involving guidepost neurons in grasshoppers are important  because they suggest that growth cones respond to long-range diffusible signals.
T       F	Unlike mRNAs, heterogeneous nuclear RNAs ("pre mRNAs") do not have 3’-UTR sequences.
T       F	Injection of synthetic bicoid mRNA into the posterior of a Drosophila oocyte would be expected to result in an increase in the production of anterior structures in the embryo.
T       F	Reporter constructs are useful because they allow us to examine how translational control elements in a mRNA function in generating the correct spatial and temporal expression of a gene.
T       F	Adrenal medulla cells form from neural crest that migrate over the dorsal surface of somites
T       F	Growth cones are guided along their migratory routes primarily by attractive signals.

4. Germ plasm is a common feature within animal embryos. Answer the following questions about the nature and formation of this material.

a. What is germ plasm? (1 points)
 
 
 
 

b. Oskar is a maternal effect gene required for formation of germ plasm in Drosophila. Describe one experiment that demonstrates this besides the mutant phenotype (2 points):
 
 
 
 
 
 

c. PIE-1 protein is thought to be important for the differentiation of the cells that form the germ line in C. elegans. What is PIE-1 thought to do? (1 point)

 

d. Recall that mex-1 gene function is important for regulating pie-1 gene function. You breed a hermaphrodite that is heterozygous for a complete loss-of-function mutation in the mex-1 gene with a male who is also heterozygous for the same mutation in mex-1. What would the phenotype of the embryos be who inherited two non-functional copies of the mex-1 gene (i.e., they're homozygous for mex-1), and why (1 point)?

5. Neural crest cells differentiate based on their position of origin, and/or in response to cues within their local environment.

a. Provide two pieces of evidence that neural crest cells respond to such local environmental cues (2 points)
 
 
 
 
 
 
 

b. You are studying a knockout mouse in which homozygous embryos are missing rhombomeres 2-6. What non-brain structures would you expect to be missing? (1 point)
 
 

6. You work in Marc Tessier-Lavigne's lab, and you are studying axon guidance. Please answer the following questions about guidance of axons in embryos.

a. You are reviewing previous axon guidance experiments. Cite two pieces of evidence that indicate netrins are required for attractive guidance of axons that migrate ventrally in animal embryos (2 points).
 
 
 
 
 
 
 
 

b. Your Ph.D. thesis work involves a set of cells in the ventral spinal cord that normally migrate dorsally. Design one experiment that would test whether netrin acts as a chemorepellant for your neurons (2 points)

 

 
 

7. Transcriptional regulators influence a wide variety of developmental processes in Drosophila.

a. Bicoid is a transcriptional regulator of zygotic hunchback expression in the early embryo. Describe what happens to levels of hunchback mRNA in the following situations. Use diagrams if you find these helpful (3 points)

(i) When an embryo is derived from a mother that is homozygous for a loss-of-function mutation in bicoid:
 
 

 

(ii) When an embryo is derived from a mother with 2 extra copies of the bicoid gene:
 
 
 
 

(iii) When an embryo is pricked at its anterior end to allow cytoplasm to leak out, and then the same embryo receives an injection of posterior cytoplasm into its anterior end:
 
 
 
 
 
 
 

b. Another well-studied example of transcriptional regulation in embryos is the way in which gap gene products regulate the expression of pair rule genes in Drosophila. In general terms, describe how this regulation occurs ( 2 points).
 
 
 
 

 

 

8. You are studying hedgehog proteins, which you may recall are important in pattern formation events in many embryos.

a. Based on what you know about the molecular basis of segment formation in Drosophila, why would mutations in hedgehog affect the formation of segments? (2 points)

 

 

 

 

b. Your friend is studying sonic hedgehog knockout mice. He wants to show that the only limb defects associated with the knockouts are due to failure of cells in the zone of polarizing activity (ZPA) to send the appropriate signals. Assuming that all experiments possible in chicks can be performed in mice, design an embryological experiment that would show this. You may not use engineered cells or introduce molecules into your experimental subjects (2 points)
 
 
 
 
 
 
 
 

c. Your friend wants to be sure that the normal dorsal-ventral polarity of the limb bud ectoderm is not affected in the knockouts. What molecule’s expression might he examine? (1 point)
 
 
 
 
 
 
 

 

9. You are assaying for mesoderm differentiation in Xenopus embryos under a variety of circumstances, using the loose lips gene as a marker. loose lips encodes a transcription factor normally expressed in the dorsal, anterior mesoderm in Xenopus. In each case, state whether you expect the levels of expression of loose lips to be higher than normal (high), normal, or lower than normal (low). (4 points).

 

Treatment	loose lips level
Inject large quantities of b-catenin mRNA into the one-cell zygote
Injection of high levels of chordin mRNA into ventral blastomeres at the four-cell stage
Injection of large quantities of BMP-4 mRNA into dorsal blastomeres of the early embryo
Combine animal cap cells with ventral, vegetal cells at the 16-cell stage
Isolate dorsal, vegetal cells with no further treatment
Injection of a mutant form of Brachyury that acts as a repressor

10. During limb development, the FGF family of proteins are thought to be crucial for the establishment of the limb bud.

a. Describe one experiment that demonstrates that FGF-10 is necessary for establishing the sites of limb bud formation in the flank of amniotes (2 points):
 
 
 
 

b. Describe one experiment that indicates FGF family members are sufficient as a functional substitute for the apical ectodermal ridge (AER) (1 point)
 
 
 
 

11. In an actual episode of the X-files called Postmodern Prometheus, the evil Dr. Pollidari has been experimenting with the homeotic gene proboscopedia in Drosophila.

a. In a terse exchange with Scully and Mulder, Pollidari asks Scully to explain to Mulder what "homeotic genes" are. If you had been Scully, what would you say? (1 point)
 
 
 
 

b. Pollidari goes on to say that humans have such genes, too.

Name three expected similarities between human homeotic genes and Drosophila homeotic genes, and one expected difference. (4 points)

Similarity #1:
 
 

Similarity #2:
 
 

Similarity #3:
 
 

Difference #1:
 
 

b. Imagine that in a later episode, the writers realize that with the availability of human embryonic stem cells, it might be possible for Dr. Pollidari to make a knockout of a human homologue of proboscopedia. After Pollidari performs the unethical experiment, he is enraged to find little phenotypic effect. Being a bright young developmental biologist, you are not surprised by this. Why? State your reasons (1 point)
 
 
 
 
 

12. It is possible to produce amphibian embryos that "exogastrulae", i.e., their marginal zone tissue does not involute, but instead elongates on the exterior of the embryo (see the figure). You reexamine neural induction in such embryos. What do you expect to see regarding expression of neural markers, and why (2 points)?

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