Name: ___________________________ 1 Recitation Section or TA: 7.013 Exam Three May 2, 2007 Exam starts at 11:05 am and ends at 11:55 am. There are 9 pages including this cover page. Please write your name on each page. Only writing on the FRONT of every page will be graded. (You may use the backs, but only as scratch paper.) Questions that call for short answers should be limited to 15 words or fewer. Page 2 5 pts Page 3 12 pts Page 4 17 pts Page 5 17 pts Page 6 15 pts Page 7 16 pts Page 8 12 pts Page 9 6 pts TOTAL out of 100_______Name: ___________________________ 2Topic 1. Ectodermin is a transcription factor found in the 500-cell stage frog embryo. It is expressed in a graded fashion, and levels of expression correlate with future fate of the embryo as follows. Animal PoleVegetal PoleNeuronMuscleGutRelative EctoderminConcentrationLowNoneHighAnimal PoleVegetal PoleNeuronNeuronMuscleMuscleGutGutRelative EctoderminConcentrationRelative EctoderminConcentrationLowNoneHigh Mutants were isolated, which have the following distribution of the Ectodermin protein. Animal PoleVegetal PoleAnimal PoleVegetal PoleNoneNoneNoneLowNoneNoneAnimal PoleVegetal PoleAnimal PoleVegetal PoleNoneNoneNoneNoneNoneNoneLowNoneNone 1a. (2 pts) For each, write next to the diagrams above what cell types you predict will form and where. Within the cells that make high levels of Ectodermin, Ectodermin activates expression of the Chordin and Noggin genes, both of which encode secreted proteins. 1b. (3 points) On the diagram of a normal embryo to the right, indicate on each line whether Noggin RNA is expressed, and whether Noggin protein would be detected. (As above, shading indicates relative Ectodermin concentration.) Animal PoleVegetal PoleNoggin RNA? Noggin Protein?Animal PoleVegetal PoleNoggin RNA? Noggin Protein?Name: ___________________________ 3 You isolate a mutant that does not express either Chordin or Noggin. 1c. (3 pts) Which of the following scenario(s) would be most likely to explain this phenotype: Mutant Ectodermin protein, increased methylation of Noggin promoter, or failure of Noggin splicing? (Circle one and explain in 15 words or fewer.) You isolate another mutant that fails to express Noggin, but expresses Chordin normally. 1d. (3 pts) Which of the following scenario(s) would be most likely to explain this phenotype: Mutant Ectodermin protein, increased methylation of Noggin promoter, or failure of Noggin splicing? (Circle one and explain in 15 words or fewer.) Noggin and Chordin are required for activation of the NeuroD gene, encoding a transcription factor that is required for development of neurons. 1e. (3 pts) Loss of NeuroD function leads to absence of all neurons as well as absence of the pancreas. Explain how NeuroD can direct formation of these two different cell fates. (Explain in 15 words or fewer) Treatment of early embryos with 5-azaC, an inhibitor of DNA methylation on cytosine, leads to a highly abnormal newborn, including increased numbers of neurons and a larger pancreas, but also abnormalities in almost every other organ and tissue. 1f. (3 pts) Explain the mechanism by which 5-azaC treatment leads to abnormalities? (15 words or fewer).Name: ___________________________ 4In a separate experiment, you isolate a mutant with an altered NeuroD promoter. The mutation leads to an increased neuron number and a pancreas that is larger than normal, but in other respects the animal is normal. Wild type and mutant promoter regions have the following sequences (altered bases are underlined): wild type mutant 5’ TAACCTCGAATCGATTG 3’ 5’ TAACCTAAAATTTATTG 3’ 3’ ATTGGAGCTTAGCTAAC 5’ 3’ ATTGGATTTTAAATAAC 5’ 1g. (3 pts) What is a likely mechanism by which the change in the neuroD promoter causes the associated phenotype? (15 words or fewer) Topic 2. Classical assays suggested that the brain contained no stem cells whatsoever. Recently, however, a seminal experiment indicated that cell division occurs in certain small regions of the brain. 2a. (2 pts) For researchers who had been trying to isolate neural stem cells, why was this finding so important? (15 words or fewer) Most stem cells express the cell surface protein Sca-1, whereas non-stem cells do not. 2b. (2 pts) Assuming that neural stem cells express Sca-1, what piece of laboratory equipment would likely be used to isolate neural stem cells? Once a population of cells that might be neural stem cells (“putative stem cells”) had been isolated, researchers performed the following assays to ask whether the cells had stem cell properties. 2c. (4 pts) Putative stem cells were placed in culture for several days. If these were neural stem cells, list the two key properties of stem cells that you would observe. 2d. (2 pts) It has been shown that some neural stem cells can produce several functional types of neurons. Cells with this ability are called 2e. (2 pts) Other neural stem cells can produce only one kind of neuron. Cells with this ability are called 2f. (2 pts) Are neural stem cells committed or differentiated? Explain in 15 words or fewer.Name: ___________________________ 5 In another assay, researchers introduced putative neural stem cells isolated from a newborn mouse into the brain of another newborn mouse, which normally produces many neurons, and later showed that the introduced cells had become neurons. 2g. (2 pts) How might researchers have done this experiment so that they could determine whether the introduced cells, specifically, had formed neurons? Explain in 15 words or fewer. In a further assay, neural stem cells from a newborn mouse were introduced into adult mice, where very little neurogenesis occurs, but these stem cells did not form neurons. 2h. (2 pts) What is likely to have changed between the newborn and adult mouse to give this result? Explain in 15 words or fewer. Any neurons that do form from stem cells need to set up correct circuits in order to function. 2i. (2 pts) What part of the neuron decides where to go, during circuit formation? 2j. (2 pts) What is chemotaxis? (15 words or fewer) 2k. (2 pts) Attractants stabilize the growing neuron by organizing what part of the cytoskeleton? 2l. (3 pts) Are migratory neurons mesenchymal or epithelial? Explain in 15 words or fewer. A recent analysis has used the technique of
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