QLanners Week 6

Answers to Assigned Questions

1. (Question 5, p. 110) Choose two genes from Figure 4.6b (PDF of figures on Brightspace) and draw a graph to represent the change in transcription over time. Create your plot in Excel (or other program that can do plots) and display the image up on your wiki page. Alternately, you can do it by hand, scan or take a photo of the plot, and display the image on your wiki page.
   • 
2. (Question 6b, p. 110) Look at Figure 4.7, which depicts the loss of oxygen over time and the transcriptional response of three genes. These data are the ratios of transcription for genes X, Y, and Z during the depletion of oxygen. Using the color scale from Figure 4.6, determine the color for each ratio in Figure 4.7b. (Use the nomenclature "bright green", "medium green", "dim green", "black", "dim red", "medium red", or "bright red" for your answers.)
   • 1 Hour
     • gene X = black
     • gene Y = black
     • gene Z = black
   • 2 Hours
     • gene X = dim red
     • gene Y = dim red
     • gene Z = dim red
   • 3 Hours
     • gene X = black
     • gene Y = dim green
     • gene Z = dim red
   • 4 Hours
     • gene X = medium green
     • gene Y = bright green
     • gene Z = dim red
3. (Question 7, p. 110) Were any of the genes in Figure 4.7b transcribed similarly? If so, which ones were transcribed similarly to which ones?
   • Genes X and Y appeared to be transcribed relatively similarly, as the ratios (and thus colors) were fairly similar to each other at each time point.
4. (Question 9, p. 118) Why would most spots be yellow at the first time point? I.e., what is the technical reason that spots show up as yellow - where does the yellow color come from? And, what would be the biological reason that the experiment resulted in most spots being yellow?
   • The technical reason that spots show up as yellow is because of a 1:1 binding ratio of the green and red cDNAs to the DNA chip. The biological reason for this is because the open reading frame for that gene is transcribed in both transcriptomes (in the presence and absence of oxygen). The reason that most spots are yellow at the first time point is because typically the gene can be transcribed in both environments at the very initial point, but will change as time goes on.
5. (Question 10, p. 118) Go to the Saccharomyces Genome Database and search for the gene TEF4; you will see it is involved in translation. Look at the time point labeled OD 3.7 in Figure 4.12, and find the TEF4 spot. Over the course of this experiment, was TEF4 induced or repressed? Hypothesize why TEF4’s change in expression was part of the cell’s response to a reduction in available glucose (i.e., the only available food).
   • TEF4 appears to be repressed. This makes sense, as a low level of available glucose would cause the cell to halt an unnecessary operations to conserve energy. Thus with such a low level of available energy, the translation of mRNA into proteins (which takes energy) would be repressed, thus repressing the TEF4 gene which is involved in translation.
6. (Question, 11, p. 120) Why would TCA cycle genes be induced if the glucose supply is running out?
   • With a low level of glucose, the genes 'Ald2' and 'Acs1' convert productss of alcohol dehydrogenase to acetyl-CoA. This acetyl-CoA is the primary entering molecule into the TCA cycle. Also, pyruvate is shifted away from acetaldehyde and towards oxalacetate, which also enters the TCA cycle. With the increased amount of molecules entering the TCA cycle, there is a need for an induction of the TCA cycle genes to increase TCA cycle activity and accommodate this necessity in the cell.
7. (Question 12, p. 120) What mechanism could the genome use to ensure genes for enzymes in a common pathway are induced or repressed simultaneously?
   • Having all the genes for enzymes in a common pathway be regulated by a similar transcription factor, and thus having similar promoter sequences, allows them to be induced or repressed simultaneously.
8. (Question 13, p. 121) Consider a microarray experiment where cells deleted for the repressor TUP1 were subjected to the same experiment of a timecourse of glucose depletion where cells at t0 (plenty of glucose available) are labeled green and cells at later timepoints (glucose depleted) are labeled red. What color would you expect the spots that represented glucose-repressed genes to be in the later time points of this experiment?
   • Red. Without the Tup1p repressor, these glucose-repressed genes would not be able to be repressed even when glucose was available. Thus the rate of this transcription would continue to increase without any controlling repression from the Tup1p.
9. (Question 14, p. 121) Consider a microarray experiment where cells that overexpress the transcription factor Yap1p were subjected to the same experiment of a timecourse of glucose depletion where cells at t0 (plenty of glucose available) are labeled green and cells at later timepoints (glucose depleted) are labeled red. What color would you expect the spots that represented Yap1p target genes to be in the later time points of this experiment?
   • Since Yap1p is a transcription factor associated with environmental stress, and the depletion of glucose is a definite stress to the cell, I would expect the spots that represented Yap1p target genes to be very bright red at later points in the experiment when the glucose levels were low. As the high level of Yap1p would induce a high amount of transcription.
10. (Question 16, p. 121) Using the microarray data, how could you verify that you had truly deleted TUP1 or overexpressed YAP1 in the experiments described in questions 8 and 9?
    • You could compare the colors of the spots of the target genes to that of a control cell that is known to have normal levels of Tup1p and Yap1p. If the dots of the Tup1p associated genes were red in the presence of glucose, as opposed to green (repressed) which they should be in the presence of glucose, you would know that the TUP1 had trult been deleted. And if the dots of the Yap1p associated target genes were much brighter red as the glucose diminished, this would indicate that there is more Yap1p ans thus an overexpressed YAP1.

Acknowledgements

• I communicated with my homework partner Eddie Azinge on multiple occasions throughout the weekend and into Monday night before the assignment was due. I offered help interpreting the article.
  

While I worked with the people noted above, this individual journal entry was completed by me and not copied from another source.
Qlanners (talk) 22:23, 9 October 2017 (PDT)

References

Campbell, A.M. and Heyer, L.J. (2003), “Chapter 4: Basic Research with DNA Microarrays”, in Discovering Genomics, Proteomics, and Bioinformatics, Cold Spring Harbor Laboratory Press, pp. 107-124.
LMU BioDB 2017. (2017). Week 6. Retrieved October 8, 2017, from https://xmlpipedb.cs.lmu.edu/biodb/fall2017/index.php/Week_6

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