lecture 5- oncogenes

Question 1

1) What is an Endogenous retrovirus and what is the theory behind how cancer may arise with their presence?

Answer 1

• transcriptionally silent viruses that have integrated into our genome and may be passed down
• they are thought to then become activated with the presence of carcinogen and cause the viral particles to spread throughout the body and cause cancer

Question 2

2) Describe the step-wise procedure of Transfection. Also, it provided strong indication that cancer cells arise from ________ and not just viral infections.

Answer 2

• First, DNA is extracted from cancer cells and mixed with calcium phosphate which forms a precipitate. This precipitate is then placed unto healthy monolayer cells on a plate. If an oncogene was present in the donor DNA, it may be uptake by the healthy cell, causing it to transform and proliferate making a tumor. Some tumor cells are extracted and injected into a healthy mouse and if a tumor forms, it confirms that cells were transformed.

Question 3

3) What experiments were performed to suggest that oncogenes (eg: HER2) are amplified in cancers? What was the survival rate of those with increased HER2 expression?

Answer 3

• Found increased expression of protein using immunoprecipitation
• southern blots found that some people with the cancer carried extra copies of the gene
• Microarrays showed overexpression of HER2 and genes close to HER2
• FISH shows the cancerous cells contain much more HER2 than normal cells
• According to the Kaplan-Meier plot, those with amplified HER2 had much lower disease free survival rate.

Question 4

3) In what way does the Ras gene become oncogenic and what tests were used to suggest such a mechanism? Explain how this theory was tested.

Answer 4

• single base pair mutation (GT) which results in a point mutation of glycine changing to valine
• It prevents Ras from being turned off
• This theory was tested using cloned oncogene and proto-oncogene DNA strands to form recombinants. Then cleaved by restriction enzymes to see which recombinants caused transformed cells using the transfection test, and then those recombinant segments were sequenced to see where exactly the change in the DNA had occurred.
• mutations in Ras are limited to very few areas

Question 5

4) Name 3 ways MYC may be amplified and state why myc amplification is so detrimental to the body.

Answer 5

1) a viral promotor integrated in front of myc gene
2) Errors made during the cell cycle result in gene amplification
3) Chromosomal translocation, where the myc gene is translocated into a chromosomal region that is expressed at higher levels
Since Myc is responsible for the production of many other proteins, it’s overexpression will lead to the overexpression of those other proteins.

Question 6

5) Describe the process in which Myc expression is amplified in Burkitt’s Lymphomas

Answer 6

• Chromosomal translocation between chromosome 8 and 14 occur where the myc gene is translocated within the heavy chain of the immunoglobin gene in chromosome 14. These genes are highly expressedwhich causes myc to become expressed and produce too many proteins.

Question 7

6) Why is Burkitt’s Lymphoma very common in areas where malaria is common too?

Answer 7

• cells need to repair more rapidly, increased rates of proliferation lead to increased chances of translocation occurring

Question 8

7) What is the role of the HMGA2 gene and how does chromosomal translocation effect its function?

Answer 8

• produces chromatin modifiying protein. Increases the rate of cell proliferation during embryonic stages.
• inhibited by Let7 miRNA.
• if another segment of chromosome is translocated to HMGA2, the binding sites for Let7 may be removed and it will no longer be under Let7 control, leading to its expression of chromatin modifying protein and increasing cell proliferation.

Question 9

8) Describe how structural changes in receptors may lead to oncogenic behavior.

Answer 9

• If the extracellular part of the receptor that binds to the ligand is deleted or truncated, the receptor will fire signals continuously without the ligand even being present.

Question 10

9) Discuss on example of hybrid proteins being formed that portray oncogenic activity.

Answer 10

• Translocation of the abl and bcr genes produce a abl-bcr protein complex that isn’t over expressed but lacks regulatory regions so it acts in an oncogenic fashion.