2) Given that E6 inactivates p53-dependent pathways, cervical cancer cells are p

Question

2) Given that E6 inactivates p53-dependent pathways, cervical cancer cells are particularly resistant to traditional treatments such as chemotherapy, the search is ongoing for new alternative therapeutics to be used in conjunction with current chemotherapeutic drugs. One such candidate is the plant flavonoid Luteolin. In 2014, Ham et al evaluate the effectiveness of Luteolin on sensitizing cervical cancer cells to apoptosis.

c) In Figure 2, panel B, the authors treated a cervical cancer cell line, HeLa, with increasing amounts of Luteolin. After 48 hours of treatment, the assayed cells via western blot for p53 and p21 as well as the S-phase Cyclins A & E.

i. Why do you think that the authors looked at both p53 and p21? I.e. what is the relationship between these two proteins.

ii. What would be 2 potential downstream effects of activating the p53-dependent pathway?

d. For each of the proteins indicate whether they showed increasing or decreasing levels with increased amounts of Luteolin.

p53

p21

Cyclin A

Cyclin E

i. Overall, the authors claimed that these results showed that cells were not proceeding into S-phase due to reactivation of p53-dependent pathways. How do these data support this claim?

Panel A: Figure 1: Ham et al E6 1.2 Panel B: 0.8 0.6 0.4 0.2 Luteolin (uM) 5 10 20 21 20 Luteolin (HM) Luteolin (LLM 5 10 20 Cyclin A Cyclin E

Explanation / Answer

Please find the answers below:

Answer c:

Part i: The cellular proteins p53 and p21 are actively involved in maintaining the cell cycle regulation, where p53 is universally known as the guardian of the cells. Here, it is important to note that p21 remains the main executionar molecule for conducting cell cycle arrest when present in active form in the cells. Since the authors over here are interested in investigating the cell-cycle arresting property of the novel molecule, they examined the effect of different concentration of their molecule of interest on the expression levels of p53 as well as p21 so that a direct inference about cell cycle arrest could be made.

Part ii: One of the potential pathways mediated by p53 downstream would be stabilization and activation of p21, thus leading to cell cycle arrest secondary to differential activation and deactivation of various cell cycle dependent kinases. The second most common pathway mediated by p53 downstreams would be self-transcriptional control of p53 where it itself acts as a transcription factor and regulates the cell cycle control.

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