PLNT3140 INTRODUCTORY CYTOGENETICS    

Due in class, Tuesday Oct. 23.

To create a model system for studying chromatin folding and unfolding, a cell line was established for which two constructs were stably inserted at separate chromosomal loci:

an array of about 200 tandem copies of a 9 kb sequence. The 9kb sequence consists of the mouse mammary tumor virus (MMTV) promoter, driving the viral genes v-ras and BPV.
a gene encoding a chimeric protein NH3+-GFP-GR-COO- where GFP is the green fluorescent protein and GR is the glucocorticoid hormone receptor

Figure 2 shows that a) In cells without hormone treatment, GFP-GR  protein fluorescence is only detected in the cytoplasm. Hybridization of the same slide with a v-ras/BPV probe (RNA-FISH) showed no transcript. b) In cells treated with the corticosteroid hormone dexamethasone, GFP-GR is only found in the nucleus, while the v-ras/BPV transcript (circled) is seen as a bright signal against a background of fluorescence. The same cell, visualized at the appropriate wavelength for RNA flourescence, shows that the v-ras/BPV transcript is only present at a single locus.

Note: Some of these figures do not reproduce well in grayscale, so you need to either print in color or view them on a screen.

1) (1 point) State a plausible hypothesis that links the observations about the glucocorticoid receptor and the MMTV/v-ras/BPV genes.

2) (3 points) For each of the following figures, write 1 or 2 declarative sentences that state the key observation from the figure(s) ie. what is the "take home lesson"?

• Figure 4, Figure 6 and Figure 5 taken together
• Figure 7
• Figure 8

3) (1 points) Write a concise and precise discussion putting all the observations from parts 1 and 2 together. In other words, what were these experiments designed to show us about the relationship between chromatin structure and gene expression? Some analysis is requried here; simply restating what you said in 1 and 2 is not sufficient. Think of 1 and 2 as the Results, and part 3 as the Discussion.

Submitting your assignment

• Make sure your assignment report includes your name, student number and UM email address.
• Feel free to include graphs, figures, tables, anything that gets your point across.
  
• Save your assignment report as a PDF file, and upload it to the PLNT3140 UMLearn dropbox site in the Assignment 2 folder. Files in word processing formats (.doc, .docx, .rtf, .odt) are NOT acceptable.

Note on grading: In assigning a grade, some consideration may be given to how the answer communicates your ideas. Keep in mind the following:

• Use precise and appropriate biological terms.
• Organization the ideas in your answer, so that a clear chain of logic is apparent. Formatting tools such as s subheadings or bullet points help to show the reader the structure of your ideas
  
• Ambiguity: Try to read your answers from the viewpoint of your reader. Are there several possible ways to interpret what you have said?
• Feel free to use figures or tables in cases where they will be useful get across your ideas.
  
• Your first step should NOT be to go into the literature and find papers on the topic. I want to see your interpretation of the data, not someone else's. All the information you need is given in this assignment.
• Be careful not to go beyond what the data is telling you. In other words, state only those conclusions that are actually supported by the data given here. It's perfectly legitimate to include sentences like "These results are consistent with what we know about glucocorticoid receptors...." but make it clear what you can actually conclude from this data, versus other points you wish to bring in.