1) Where in the cell are trimeric G proteins found? (2pt)
2) What are the enzyme and the substrate needed to form cAMP? (3pt)
3) When considering the activation of a receptor as the 'starting point', how is production of cAMP similar to production of cGMP? How is it different? (6pt)
4) Cells must be able to rapidly activate and inactivate intracellular signaling pathways. Why are both of these changes important? (3pt)
5) Briefly describe how increases and decreases in the concentration of the second messenger IP3 are controlled within the cell. (3pt)
6) The insulin receptor is an example of what type of receptor? (2pt)
7) The insulin receptor continues to activate intracellular proteins even after insulin has been cleared from the bloodstream. How is this possible? (4pt)
8) Compare and contrast a graded response with a threshold response. (5pt)

Section II
All of the parts in each question relate to the experimental scenario developed in that question. Be sure to answer the questions in light of the information you have been given. (Also, as always, answers must be in complete sentences.)

Question I parts A-G (27pt total)

You run a lab interested in the molecular basis of cystic fibrosis. Joe Scientist is a graduate student in your lab. His project is interested in the effects of physical activity on the function of CFTR protein. Using cells grown in a petri dish (tissue culture) he is trying to determine the effects of adrenaline on the mucus of 'normal' cells and CFTR DF508 mutant cells.

A) The CFTR gene is found in all cells but order to study CFTR protein function Joe must study certain cell types. Give an example of a cell type that you would recommend. Why must he use a cell type like the one you named? (4pt)
B) Where in the cell is 'normal' CFTR protein found and what is its function? (5pt)
C) Joe hopes to mimic the effects of physical activity by adding adrenaline. What is adrenaline? How does adrenaline have an effect on the cell? (i.e. what does it activate or inhibit?) (5pt)
One day Joe sheepishly comes to you with data that do not make sense to him and the following story. He was working late one night determining extracellular ion concentrations. By accident he spilled coffee into a few of the petri dishes of cells. Not wanting to scrap a week's work he continued his experiments and made a table of his results.

Extracellular anion concentration under varying experimental conditions
 

Treatment	Normal cells	CFTR DF508 mutant cells
None (just cells)	100%*	50%
Added adrenaline	120%	50%
Added adrenaline and coffee	140%	50%

* the extracellular anion concentration for normal, untreated cells was set to 100%. The concentrations seen under all other conditions are shown as a percentage of that control.

Joe was quite happy with the results seen with adrenaline but is confused by the effect of 'adrenaline + coffee'. He comes to you for advice.

D) What lab safety rule did Joe break when completing this experiment? (2pt)
E) Joe was happy with the 'added adrenaline' results. Why could he predict that added adrenaline could increase anions in the extracellular matrix? (3pt)
F) Why is the extracellular anion concentration so low in the CFTR DF508 mutant? What is wrong with the mutant CFTR protein that results in these data? (4pt)
G) You tell Joe that the results are consistent with other known facts. You explain by saying that one of the reasons that coffee wakes you up that caffeine is a phosphodiesterase inhibitor. Why would treatment with a phosphodiesterase inhibitor cause an increase in the effects caused by adrenaline? (4pt)
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Question 2 parts A-D (14pt total)

You and your lab partners have been diligently working to characterize your mating type a mutants. You have determined that Mutant I can secrete pheromone but does not shmoo. Mutant II shmoos but only shows 30% of the expected mating efficiency. You have log phase and stationary phase cultures growing in the incubator when member of the lab accidentally gets alcohol on the outside of your culture tubes, erasing the names. There is no time to start new cultures so you decide to use what you have and experimentally determine the identity of each of you cultures. To help you, your lab technician provides stock cultures of wildtype a and a cells that she had growing. She also says you can have access to her stocks of plates and liquid media.

A) What is the name of the pheromone secreted by Mutant I? (2pt)
B) Briefly describe the difference between a log phase and a stationary phase culture? (3pt)
C) What is a shmoo and why is the lack of a shmoo consistent with a defect in mating? (3pt)
D) Design one experiment that will allow you to determine the identity of the cells in your log phase cultures. (Volumes are not necessary. Be sure to include how you will be able to definitively interpret the results.) (6pt)
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Question 3 parts A-H (33pt total)

(Disclaimer--The following scenario is purely hypothetical and only designed for this review. Don't go asking for genetic testing--this condition is much more complicated than described here)

Hypostudiosis (common name procrastination) is a disease that effects over 75% of the world's population at some time during their life. In 'normal' people exposure to sunshine and warm weather causes the release of a peptide hormone. The hormone is bound by the "RR" (relax receptor), a tyrosine kinase associated receptor. Activation of RR and its related pathway causes feelings of lethargy and well-being and results in a lack of focus and concentration. It has been known that people suffering from mild hypostudiosis have a mutant form of RR (called RRM) that always showed low levels of activity. This low level of activity results causes RRM people to feel continually unfocused and content without ever having the hormonal stimulus.

A) What general class of receptors does RR belong to? Name 2 other types of receptors found in this class. (specific examples are not necessary, just general types) (4pt)
B) What is the basic function of a kinase? Why is this function focused on 3 particular amino acids? (3pt)
BONUS: Draw any one of the three amino acids that can be effected by a kinase (2pt)
C) RR is involved in activating an enzyme found on the Golgi membrane called 'vegin'. Describe a possible pathway resulting in vegin activation. (Begin with hormone binding. You can assume vegin activation uses the most direct pathway) (5pt)
D) A proposal to the NIH suggests using tissue culture to see if various compounds will reduce the activity of RRM in the mutant cells. Their list includes cholera toxin. Would you predict that cholera toxin will reduce RRM activity? Why or why not? (4pt)

Recent discoveries have determined that severe hypostudiosis also requires a mutation in a second separate protein. This protein, called CRAM (component required for accessing memory), is a tyrosine phosphatase. A molecular biologist has determined that the mutant form of CRAM found in hyperstudiosis cells has a much shorter half-life than the normal form. Her current hypothesis is that CRAM is falling under the N-end rule for protein stability.

E) What is the N-end rule? What information could she supply that would support this hypothesis? (3pt)
F) CRAM is a cytosolic protein. Describe the general structure of the cytosolic complex involved in protein degradation? (Include its name.) (5pt)
G) Describe the modification to CRAM that must occur so that it will be targeted to the complex named in F. (Include the name of the modification.) (4pt)
H) Hypothesize how a decrease in CRAM stability could result in a more severe case of hyperstudiosis. (5pt)