A researcher studying a line of cancer cells notices that cellular content of RhoBTB3 and Cyclin E1 increase and decrease through the cell cycle. a. He hypothesizes that RhoBTB3 signaling regulates the production of Cyclin E1 and the entry of the cell into mitosis. Based on the data below, explain why his results do not support this hypothesis. Identify what conclusions are supported by this data. b. Suppose that the researcher can engineer a cell line in which RhoBTB3 expression can be induced by the addition of a drug. Design an experiment using this cell line to test whether RhoBTB3 regulates the production of Cyclin E1. Describe what results he can expect if his hypothesis is correct. A line graph is shown along with immunoblots showing protein levels of RhoBTB3 and Cyclin E1. The top of the image shows a line graph with x-axis labeled h and marked from 0 to 18 in increments of 2 and y-axis labeled Protein Level (AU) and marked from 0 to 1 in increments of 0.2. Two lines on the graph show the protein level of RhoBTB3 and Cyclin E1. The line for RhoBTB3 is at approximately 0.2 AU between 0 to 10 h and begins rising at 12 h and increases to 18 h. The line for Cyclin E1 is at 0 Au until 8 h and rises until 16 h before decreasing at 18 h. Two immunoblots are shown at the bottom of the image. The first shows light bands for RhoBTB3 from 0 to 12 h and getting darker at 14, 16, and 18 h. The second shows no bands for Cyclin E1 until 10 h. The band is at medium intensity at 10 h and darker at 12, 14, and 16 h. The band at 18 h is at medium intensity. Hours 0 and 2 are labeled M. Hours 4, 6, 8, and 10 are labeled G1. Hours at 12, 14, and 16 are labeled S. Hour 18 is labeled G2.

Question

A researcher studying a line of cancer cells notices that cellular content of RhoBTB3 and Cyclin E1 increase and decrease through the cell cycle. a. He hypothesizes that RhoBTB3 signaling regulates the production of Cyclin E1 and the entry of the cell into mitosis. Based on the data below, explain why his results do not support this hypothesis. Identify what conclusions are supported by this data. b. Suppose that the researcher can engineer a cell line in which RhoBTB3 expression can be induced by the addition of a drug. Design an experiment using this cell line to test whether RhoBTB3 regulates the production of Cyclin E1. Describe what results he can expect if his hypothesis is correct. A line graph is shown along with immunoblots showing protein levels of RhoBTB3 and Cyclin E1. The top of the image shows a line graph with x-axis labeled h and marked from 0 to 18 in increments of 2 and y-axis labeled Protein Level (AU) and marked from 0 to 1 in increments of 0.2. Two lines on the graph show the protein level of RhoBTB3 and Cyclin E1. The line for RhoBTB3 is at approximately 0.2 AU between 0 to 10 h and begins rising at 12 h and increases to 18 h. The line for Cyclin E1 is at 0 Au until 8 h and rises until 16 h before decreasing at 18 h. Two immunoblots are shown at the bottom of the image. The first shows light bands for RhoBTB3 from 0 to 12 h and getting darker at 14, 16, and 18 h. The second shows no bands for Cyclin E1 until 10 h. The band is at medium intensity at 10 h and darker at 12, 14, and 16 h. The band at 18 h is at medium intensity. Hours 0 and 2 are labeled M. Hours 4, 6, 8, and 10 are labeled G1. Hours at 12, 14, and 16 are labeled S. Hour 18 is labeled G2.

Answer 1

a. The researcher's hypothesis states that RhoBTB3 signaling regulates the production of Cyclin E1 and the entry of the cell into mitosis. However, the data shows that the levels of RhoBTB3 and Cyclin E1 do not directly correlate with each other throughout the cell cycle. Specifically, Cyclin E1 levels increase until 16 h and then start decreasing at 18 h, while RhoBTB3 levels start increasing at 12 h and continue to increase until 18 h. If RhoBTB3 signaling regulated the production of Cyclin E1, one would expect the levels of Cyclin E1 to increase or decrease in tandem with the levels of RhoBTB3. Therefore, the results do not support the hypothesis that RhoBTB3 signaling regulates the production of Cyclin E1.

The conclusion that can be supported by this data is that there is no direct regulatory relationship between RhoBTB3 signaling and the production of Cyclin E1. Other factors may be involved in regulating the production of Cyclin E1, or Cyclin E1 production may be regulated independently of RhoBTB3 signaling.

b. To test whether RhoBTB3 regulates the production of Cyclin E1, the researcher can design an experiment using the engineered cell line in which RhoBTB3 expression can be induced by the addition of a drug.

Experimental design:
1. Divide the engineered cell line into two groups: a control group and a drug-induced group.
2. In the control group, do not add any drug to induce RhoBTB3 expression.
3. In the drug-induced group, add the drug to induce RhoBTB3 expression.
4. Collect cell samples at regular time intervals (e.g., every 2 hours) for a specific duration (e.g., 24 hours).
5. Measure the protein levels of Cyclin E1 in both groups using immunoblots.
6. Analyze the results to compare the levels of Cyclin E1 in the control group and the drug-induced group at different time points.

If the hypothesis is correct and RhoBTB3 regulates the production of Cyclin E1, the researcher can expect the following results:
- In the control group (without drug-induced RhoBTB3 expression), the levels of Cyclin E1 should remain constant or follow a pattern that is not significantly influenced by the cell cycle.
- In the drug-induced group (with drug-induced RhoBTB3 expression), the levels of Cyclin E1 should show variations that correlate with the cell cycle. Specifically, the levels of Cyclin E1 should increase during specific phases of the cell cycle, such as G1, S, and G2, when RhoBTB3 is expressed.

By comparing the protein levels of Cyclin E1 between the control group and the drug-induced group at different time points, the researcher can determine whether RhoBTB3 expression affects the production of Cyclin E1 and supports or does not support the hypothesis.