Eyanosch Week 7

(Question 5, p. 110) Choose two genes from Figure 4.6b (PDF of figures on MyLMUConnect) and draw a graph to represent the change in transcription over time. You can either create your plot in Excel and put the image up on your wiki page or you can do it in hard copy and turn it in in class.
- File:Change in transcription Week7 HW EY 20151915.xlsx
(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.)
- Gene X would be Black(100%), dim red(220%), black(100%), medium green(15%) at each corresponding ratio (Hour 1, 3, 5, and 9 respectively). Gene Y would be black(100%), medium red(450%), black(95%), light green(5%) at each corresponding ratio (Hour 1, 3, 5, 9). Gene Z would be black(100%), black(150%), dim red(200%), dim red(200%) at each corresponding ratio (Hour 1, 3, 5, 9)
(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?
- The genes X and Y showed a similar transcription pattern of an early induction phase followed by increased repression. Although the rates of induction and repression are never identical after hour one, they follow a similar pattern.
(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?
- Most spots are yellow because not enough time has passed to see the results of the interaction between the control and experimental group. The yellow dots are due to an even mixture of red and green (1:1). Neither an viewable repressed or induced amount of transcription is occurring, the genes affected are only in small amount at this early stage in viewing.
(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).
- The TEF4 gene is being repressed over time. The initial color is yellow, but with each time lapse progression becomes increasingly green, starting as a light green within the second frame (Growth OD .46) and eventually a light green in the final frame (Growth OD 6.9). TEF4 is a gene that codes for translation elongation factors, which allow for polypeptide production. This process requires energy and a reduction in the available energy source for a cell, glucose, would inhibit the gene for coding those protiens.
(Question, 11, p. 120) Why would TCA cycle genes be induced if the glucose supply is running out?
- The TCA Cycle, also known as the Krebs or Citric Acid Cycle, utilizes Acetyl-CoA (produced when pyruvate is oxidized after being formed from the breakdown of glucose in glycolysis) to produce energy the cell can use. As the cell runs out of glucose, it will have to turn towards the catabolism of carbohydrates, fats, and proteins for potential precursors to be utilized in the TCA cycle. This will cause an induction in the TCA cycle genes so the cell will have enough available energy to supply the necessary processes to survive.
(Question 12, p. 120) What mechanism could the genome use to ensure genes for enzymes in a common pathway are induced or repressed simultaneously?
- Common pathways would be affected by the same transcription factors (repressors/inhibitors). Altering the amount of repression or induction through transcription factors would ensure genes for the enzyme produced on the pathway are affected simultaneously.
(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?
- I would expect the spots that represented glucose-repressed genes to be red. Without the repressor TUP1 gene transcription will not be repressed and induction of transcription will occur yielding a red color on the microarray. The lack of glucose available will cause the glucose-repressed genes to induce transcription.
(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?
- I would expect the spots with that represent the Yap1p target genes to be red at the end of the experiment. Over expression of a gene that codes for the promotion of transcription in response to the cells environment will induce transcription and utilize available glucose storages.
(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?
- If TUP1 was truly deleted then the microarray would be red at the end of the experiment because available glucose would not inhibit transcription but be utilized by it. Overexpression of YAP1 in cells would allow for a larger response to environmental changes within the cell, a larger response calls for more energy. Therefore I would predict the overexpresison of YAP1 to look red.