Bio 362 Human Genetics Spring 2008
Narcolepsy study sheet for April 15th
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Chabas et al., 2003:
For background information on MHC/HLA loci and allele nomenclature, see http://pathmicro.med.sc.edu/ghaffar/mhc2000.htm .
What are the normal stages of sleep? What goes wrong in narcolepsy?
What is the structure and function of class II MHC proteins? In what cells are they expressed?
What is relative risk?
In a very general sense, what do tables 1 and 2 suggest about different MHC genotypes and their link to narcolepsy?
Can you figure out how relative risks were calculated in table 2?
What do twin studies suggest about narcolepsy susceptibility?
Where in the brain is hypocretin made? Where are hypocretin receptors expressed?
After you've read the Lin paper, compare its text with the canine narcolepsy section of this review.
What mutations were found in different narcoleptic dogs (see Fig 3)?
How is the HLA DQA1*0102 DQB1*0602 association with human narcolepsy related to the hypocretin story??

Black, 2005:
What experiments on humoral immune responses were done to explore autoimmunity in narcoleptic patients?
Which of these experiments gave negative results, and which gave some positive results? What do the results mean for the etiology of narcolepsy?

Lin et al., 1999:
Explain in general why backcrosses (see top of second page) were necessary to prepare for this linkage study.
The researchers thought they found linkage to an immunoglobulin gene, but what was actually the case?
How did they use the idea of synteny to help with their chromosome walking efforts?
Once they had isolated the DNA across the region, what additional features were found to help with finer mapping?
What are all the features depicted in Figure 2?
For the dogs mentioned in the middle of Figure 2, what does it mean for the assocated line to be blunt at one end and have an arrow at the other end?
Which two dogs initially defined the genomic region of interest?
Which dogs further narrowed the region of interest? How many known genes were in that region?
How did Southern blotting initially suggest that there were mutations in HCRTR2?
How did PCR help clarify the nature of the mutation? What were the templates used in the different panels of Figure 5?
What are the different mutations in narcoleptic Dobermans and Labradors?
What was intriguing about the hypocretin system?
Don't try to muddle through all the brain anatomy in the discussion. Just get a general sense of whether hypocretin-synthesizing neurons are widespread or localized in the brain, and whether the regions they project to are widespread or localized.

Chemelli et al., 1999:
What stages of the generation of knockout mice were examined in Figure 1, panels B and C? (In other words, were embryonic stem cell lines or live mouse tissue examined in each case?)
Relate the probes and/or primers used in Figure 1B and C to the diagram in panel A.
What part of the brain was examined for orexin expression in Figure 1D-E? What kind of probe is used in each experiment, and what exactly is being detected?
In Figure 2, why are there two different Y axes in the same graph? Which data refer to each axis?
Use Figure 2B to give a narration of mouse F's dark phase (normally an active period).
What parts of the brain are examined in Figure 5? Why are these data important to include in the paper?
What is modafinil? What does Fos immunoreactivity have to do with anything (Figure 6)? Why are the Figure 6 data important?
What specific mechanism might you predict for modafinil?
Glance at the Hara et al. 2001 paper in the supplementary folder enough to answer the following two questions:
Why might the mouse model generated here be a better model of human narcolepsy than a prepro-orexin knockout?
What were the phenotypes in the mice generated in this study?

Peyron et al., 2000:
What subset of narcolepsy patients had their DNA examined? What were the researchers looking for?
What one mutation was found?
How was the allele in that one mutant patient tested in cultured cells (Figure 1)? What happens to the altered preprohypocretin?
Does the mutant allele have a dominant effect at the cell level (Figure 1e)? How does that fit with the fact that the disease seems dominant in that patient (see discussion)?
What defect in narcoleptic brains is demonstrated in Fig 3a-d and 4a-b? Where did these samples come from? How can this phenotype exist if the specific genetic defect is not in preprohypocretin or its receptors?
What phenomenon is shown NOT to be occurring in mutant brains in Fig3e-g? How can that make sense with the mechanism we infer must have led to narcolepsy in these patients (see discussion)?