2005_pset7 - 7.06 Problem Set#7 Spring 2005 1 a You are a...

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1 7.06 Problem Set #7, Spring 2005 1. a. You are a grad student studying genetic alterations that lead to tumorigenesis. While plating primary cell cultures from a healthy person and a tumor from a cancer patient, you mix up the samples. A colleague suggests that you use the differences in growth properties between the two samples to distinguish them. Describe how the growth properties of cancer cells differ from those of healthy primary cells. b. When studying the cancer cells from part a) , you discover that there is high Ras activity in the cells, but you can find no mutations in the Ras gene. You then start investigating different Receptor Tyrosine Kinases. You find by immunofluorescence, using a monoclonal antibody against the kinase domain, that one specific RTK is now located throughout the cytosol instead of in the plasma membrane. You then perform Western blot analysis on protein extracts from these cancer cells, and find that the RTK is much larger than normal. What kind of mutation in the RTK is indicated by these results, and how does that specific type of mutation lead to cancer? c. In addition to differences in growth properties, cancer cells also have morphological differences, such as a disorganized cytoskeleton. The Rho family of small GTPases is involved in cytoskeleton regulation, and the pathways that Rho proteins are involved in are implicated in cancer development. Their regulation of the cytoskeleton is thought to be important in cell migration during cell division and invasion of other tissues. In an experiment performed a decade ago, it was found that transfecting NIH 3T3 cells with a construct that expressed a specific mutant form of RhoA could transform these cells. Based on this experiment, would you predict that RhoA is a proto-oncogene or a tumor suppressor gene? Would you predict that the RhoA mutation is a gain-of-function mutation or a loss-of-function mutation? Explain your answers. d. In class, we learned about the experiment by the Weinberg lab, in which an oncogenic version of Ras was discovered in cells from a human bladder carcinoma.
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