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706_F06exam3_key - Answer Key Name 7.06 Cell Biology EXAM#3...

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Name: ___________________________ 1 Answer Key 7.06 Cell Biology EXAM #3 November 21, 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. 25 pts __________ Question 3. 25 pts __________ Question 4. 25 pts __________
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Name: ___________________________ 2 Answer Key Question 1 (25 points) You have joined a lab that works on cell motility. One of the cell lines this lab studies is a keratinocyte line. Unsure as to how these cells crawl, you ask the graduate student in the lab a number of essen- tial questions. A. (1 point) What are the protrusions called at the leading edge of the crawling cell? Lamellipodia and/or filopodia B. (2 points) Propose an experiment to show that the movement of keratinocytes is mediated by the actin cytoskeleton. You do not need to describe extensive experimental detail. I could treat cells with an actin depolymerizing drug, i.e. latrunculin or cytochalasin D and ex- amine the effects of the drugs on movement. Alternatively I could express a dominant negative form of actin (if existed) or use siRNAs against actin, myo2, arp2/3 to perturb actin cytoskeleton function. C. (4 points) How do you measure the critical concentration of G-actin necessary for actin filament formation? Be sure to add experimental detail. I would measure filament mass at various G-actin concentrations. The concentration at which I first observe filament formation would be the critical concentration for actin filament forma- tion. I would use one of the following assays to measure filament formation (a) Ultracentrifugation (b) viscosity of the solution (c) measure filament length using a microscopic assay (in this case I would have to add fluo- rescent G-actin.
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Name: ___________________________ 3 Answer Key D. (4 points) How would addition of a seed affect the kinetics of filament formation? Adding seed would accelerate filament formation. The initial nucleus assembly is the rate-lim- iting step in filament formation. By adding seed, this rate-limiting (slow step) of the reaction would be eliminated. Armed with all the information necessary to get a start on your project you first decide to determine the critical concentration for the (+) end and the (-) end. E. (6 points) Describe an experiment that would allow you to determine the critical concentration for the two ends including experimental detail. I would measure filament mass at various actin concentrations as described for (3) except I would add a seed to the reaction. In addition to the seed I would add either a (+) end or (-) end capping protein to specifically determine the Cc for the (-) end and the (+) end, respectively.
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