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706_F06exam2 - Name 7.06 Cell Biology EXAM#2 This is an...

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Name: ___________________________ 1 7.06 Cell Biology EXAM #2 October 24, 2006 This is an open book exam, and you are allowed access to books and notes but not computers or any other types of electronic devices including cell phones. Please write your answers in pen (not pencil) to the questions in the space allotted. Please write only on the FRONT SIDE of each sheet. And be sure to put your name on each page in case they become separated! Remember that we will Xerox all of the exams. Good luck! Question 1. 25 pts __________ Question 2. 30 pts __________ Question 3. 25 pts __________ Question 4. 20 pts __________
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Name: ___________________________ 2 Question 1 (25 points) You are interested in studying the enzyme Air1 responsible for Histone H3 phosphorylation in mam- malian tissue culture cells. Your friend raised antibodies against the enzyme and is kind enough to send you an aliquot. A. (6 points) Describe how you could use this antibody to examine whether the protein that is recog- nized by the antibody is indeed responsible for Histone H3 phosphorylation. Be sure to include details on the experimental process and controls. You decide to characterize the regulation of Air1 in more detail by first examining the levels of the protein during the cell cycle in synchronized wild type cell cultures. You arrest the cells in G1 by se- rum starvation and release them into the cell cycle synchronously by re-adding serum. To monitor cell cycle progression you measure DNA content. This is the result that you obtain. B. (3 points) Which stages of the cell cycle is Air1 absent from? When is the protein present at high levels and when at low levels?
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Name: ___________________________ 3 C. (6 points) How could you test the hypothesis that ubiquitin-proteasome dependent protein degra- dation is responsible for the loss of Air1 in certain stages of the cell cycle? Include controls. Draw the expected result if the hypothesis is true and explain your reasoning. Do NOT write in this space.
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Name: ___________________________ 4 Intrigued by Air1 you now wonder whether the protein has functions in addition to the regulation of Histone H3. To examine this possibility you use a novel method called RNAi to knock down the lev- els of Air1 in cells. Using this method you achieve knock down of Air1 protein in serum starved cells, which are then released from block. Here is the result you obtain. E. (6 points) What goes wrong in cells lacking Air1 (other than checkpoint failure)? Provide two po- tential explanations as to why this phenotype may have occurred.
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