Bio 362 Human Genetics Spring 2008
Huntington study sheet for March 18th
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Bates, 2005:
Read this paper for a general overview only. Note the mouse models mentioned, and make a list of these and other Huntington mouse models (described in the next two review papers) that shed significant light on the pathology of the disease.

Gatchel and Zoghbi, 2005:
Read the first and third sections of this paper (loss of protein function and altered RNA function) only to get a general overview. Be able to describe the general situation regarding the diseases/genes in these categories, but don't worry about details. Focus on the middle section about disorders caused by gain of function.
What was originally thought about the effect of aggregates on the ubiquitin-proteasome system? What experiment suggests there is no such effect?
Do the authors think that aggregates are pathogenic? What experiments are cited in support of their position?
How might polyglutamine repeats affect transcription?
What might polyglutamine-containing proteins be doing out in the cytoplasm to cause disease symptoms?
What's the evidence that more of the protein besides the polyglutamine domain is causing problems?

Michalik and Van Broeckhoven, 2003:
What did Perutz propose as the basis for why repeats over a certain size are pathogenic? How did recent structural studies refute his proposal?
Given those results, why do only people with long polyglutamine tracts show symptoms?
Explain the multiple nucleation hypothesis. What factors influence aggregation rate?
Is a loss-of-function mechanism influencing Huntington's phenotypes?
What is the evidence for gain-of-function mechanisms in Huntington's?
Explain evidence for sequestration as the gain of function. What could be sequestered?
Explain evidence for toxicity as the gain of function.
Why are only neurons affected?
Why are only particular neuron types affected?
What evidence suggests that the aggregates are themselves toxic?
What evidence suggests that soluble mutant polyglutamine is toxic?
By what mechanisms could soluble mutant polyglutamine be toxic, and why would such toxicity lead to only late-onset disease?
How do you reconcile whether aggregates of polyglutamine proteins are a primary cause of toxicity, or whether they are protective against toxicity? Recap the various evidence in this paper and the Gatchel and Zoghbi paper, and decide what you think.

Arrasate et al., 2004:
What is an IB? What are the three possible roles of IBs in HD?
What system did the authors develop? 
What plasmids were transfected? What cell types were used?
How did they identify IBs in living neurons? For which polyQ-expansion lengths did IBs form?
Figure 1: What do the red and green indicate in part a? Which cell dies? How does part b validate the conclusion from part a?
How are Fig 1c-d different from 1e-f? What effect does repeat size have on survival?
Figure 2a-b: Does detergent treatment remove diffuse Htt, Htt in inclusion bodies, or both?
Figure 2c-f: Does the appearance of inclusion bodies correlate with a correspondingly increased risk of death?
Figure 3: How do panels a and b validate the methods for the rest of the figure? What correlation is shown in part c? How is part d a control for c?
Compare Figure 3e to 3c. What is consistent between the different cell types used?
What do Figure 4 a and b show about the relationship between IB formation and levels of diffuse Htt? 
Figure 4c-d: What two groups of neurons were compared? How do they differ with respect to survival and initial Htt?
Figure 4 e-f: What two groups of neurons were compared this time? How do they differ with respect to survival and intial Htt levels? What does this suggest about the role of inclusion bodies?  

DiFiglia et al., 2007 (questions from Mark and Brady):

Overall Q: should the siRNA be conjugated to cholesterol?  pros/cons?
Researchers put the human versions of the genes in with 10 and 100 repeats, however these are at the LOW and HIGH end of what is normal in actual HD cases.  Are we okay with this?

1) Why is turning off the gene in an inducible mouse model important?

2) What is the striata?  Why are researchers interested in this portion of the brain?

3) What is clasping and why is it important?

4) Explain short hairpin RNAs vs siRNA. Why even mention shRNA?

5) why did the researches conjugate the siRNA with cholesterol?

6) What is Cy3?

7) Explain the differences between the AAVHtt18Q vector and the AAVHtt100Q vector.

Fig 1 - What is to be gathered from panels a) and b)? How are they different from panels c) and d)? Why was panel e) included? What is the takeaway message of figure 1?

Fig 2 – How is the left image in panel a) different from the right? What is meant to be gathered from panel b)? What is the purpose of the figure on the right of panel c)? What is different between group 1 and group 2 mice? Annotate panel d). What is the takeaway message of figure 2?

8) what does EM48 do?

Fig 3 – Explain the differences between panels a), b) and c). In panel d), where are the results significant? What is suggested by panel e)? What is the takeaway message of figure 3?

9) Fig 4 - What are neuropil aggregates? How do neuropil aggregates related to Htt treatment? Why are there two groups of mice? What is the takeaway message from figure 4?

Fig 5 – What is clasping? Why were clasping and footslips studied? What is to be gathered from panel b)? What is the takeaway message of figure 5?