Question

If you are able to answer any or all of these that would begreat!!!

25. Talked about the warburg effect ( aerobic glycolysis) inwhich cancer cells preferentially use fermentation instead of goingthrough oxidative phosphorylation. Answer the followingquestions?

A) If cells are going to preferentially use fermentation, theyneed to transport higher levels of glucose into the cell. Explainwhy this is so?

B) Cells that preferentially use fermentation had higher levelsof the GLUT1 carrier protein at the membrane. explain the mechanismGLUT1 uses to bring glucose into the cancer cells?

C) If the GLUT1 transporter is composed of two main domains,what are their likely functions? what is the nature of the aminoacids within each domain?

D) What are two ways cells could increase the amount of GLUT1transporter on the cell membrane? Briefly explain each.

E) What is the one way the rate of movement of glucose into thecell through GLUT1 carriers could be altered ( either increase ordecreasd)? Describe a change you could make and be sure to indicatehow it would alter the rate of glucose uptake( either increase ordecrease)?

F) A drug called fasentin inhibits GLUT1 carriers. However, whenyou treat FTC133 cells with fasentin, they do not die. What is onehypothesis for how they are surviving in the absence ofglucose?

G What is a prediction that you could make based on yourhypothesis?

H) In the paper, the authors conclude that adding the PTEN backinto the cancer cells via transfection decreases the amount ofglucose that FTC133 cells take up. You hypothesize that HeLa cellsmight benefit from having additional PTEN protein. YOu transfectDNA coding for the PTEN protein into the HeLa cells and perform awestern blot to confirm PTEN expression. Unfortunately, you do notsee any expression of PTEN. What is one possible reason you dontget PTEN expression in your HeLa cells? ( HINT: you know yourantibodies are working and that transfection was successful, sothink about factors controlling gene expression)

I, Since you couldnt get PTEN expressions in the HeLa cells, youdecide to return to the FTC133 cells in order to test if the lipidphosphatase activity of PTEN is required for its ability todecrease glucose consumption. What type of mutation ( silent,missense, nonsense? frameshift or not?) would you introduce intothe PTEN gene in order to destroy the catalytic ability of PTENprotein while retaining the rest of the protein's structure andfunction? Explain your answer.

J) Next, you decide that you want to purify large amounts ofyour wild type and mutant PTEN proteins to test their catalyticabilities in in vitro. Your labmate suggests that you transform (insert) your DNA into bacterial cells because they are rapidlygrowing and are know to express large amounts of protein under theright conditions. You consider this but are unsure as to how wellyour human PTEN protein might be expressed in these cells. Based onwhat you know about the genetic code and about gene expression inprokaryotic and eukaryotic cells describe (i) one reason why yourgene might be able to be expressed in bacterial cells under theright conditions and (ii) two reasons why your PTEN gene might notbe expressed properly. (iii) Then, decribe one modification thatcould be made to your PTEN DNA to increase the likelihood that itwill be expresssed in the bacteria.