6) A mutant yeast strain stops proliferating when shifted from 25°C to 37°C. Whe

Question

6) A mutant yeast strain stops proliferating when shifted from 25°C to 37°C. When these cells are analyzed at the two different temperatures, using a machine that sorts cells according to the amount of DNA they contain, the graphs below are obtained.

Which of the following would not explain the results with the mutant?

A         inability to initiate DNA replication

B         inability to begin M phase

C         inability to activate proteins needed to enter S phase

D         inappropriate production of a signal that causes the cells to remain in G1

Please explain your choice:

            i) What is the expected DNA content of cells unable to start DNA replication? What would this look like on the graph?

ii) What is the expected DNA content of cells stuck in G2, unable to start M phase? What would this look like on the graph?

iii) What would the graph look like if cells could not produce proteins that block /slow down the start DNA synthesis? What would this look like on the graph?

BIOL111 -Problem Set 1 Name 6) A mutant yeast strain stops proliferating when shifted from 25°C to 37°C, When these cells are analyzed at the two different temperatures, using a machine that sorts cells according to the amount of DNA they contain, the graphs below are obtained Which of the following would not explain the results with the mutant? inability to initiate DNA replication inability to begin M phase inability to activate proteins needed to enter S phase inappropriate production of a signal that causes the cells to remain in G Please explain your choice: i) What is the expected DNA content of cells unable to start DNA replication? What would this look like on the graph? ii) What is the expected DNA content of cells stuckin G2,unable to start M phase? What would this look like on the graph? iii) What would the graph look like if cells could not produce proteins that block slow down the start DNA synthesis? What would this look like on the graph?

Explanation / Answer

The analysis of the DNA content of the cell can be determined by Flow cytometry or fluorescence activated cell sorter or FACS. It's principle is dependent on the basis of fluorescence emission by cells which the amount of DNA is directly proportional to the amount of DNA of each cell.

The cells are analyzed by staining them with a fluorescent dye and passing them one by one within a capillary tube. The amount of fluorescence emitted by each cell is analyzed by recorder.

From analysis we get

If we plot a graph taking the relative amount of DNA per cell along x axis and number of cells along y axis ---

1) at G1 stage there is a peak in fluorescence indicating that highest number of cell are in G1 phase. In this stage the duration is longest at cell cycle that's why maximum number of cell are there . Though their DNA content is not duplicated at G1 phase.

2) then there is a plateau indicating intermediate amount of DNA in S phase cells.In S phase there is a mixed population of cells having some cells duplicated amount of DNA , some cell's DNA content is being duplicated, replication is not completed.

3) after that there is a small peak , smaller than G1 peak this indicates these cells are in G2 or M phase. Their DNA content is duplicated hence more amount of DNA than S phase.

Now in this question when mutant yeast strain was shifted from 25 degree Celsius to 37 degree Celsius , it stopped proliferation.

The graph at 25 degree Celsius shows ( if we consider it as the flow cytometry graph) then there is G1 , S , G2 and M phase.we can know there is proliferation by measuring the amount of fluorescence against DNA content of each cell.In this graph as previously mentioned a peak in G1 , then a plateau then G2 And M phase cells.

But at 37 degree Celsius we can only see A large peak at G1 , highest level fluorescence but after that no peak or plateau formation. From this we can conclude the cell cycle stops at G1 phase ,so no entrance into S phase. And no further proliferation.

Option A ,C and D would explain the result with mutants because of their inability to DNA synthesis or inability to activate protein needed to activate replication or inappropriate production of factor which will keep the cells in G1 phase.

But if we see normal cells ( option B) then analyzing the graph we would know there is further progress in cell cycle.

So option B is correct.

1) the expected DNA content will be in the cells stuck in G1 phase is 2C , cells are not duplicated.

2) the expected DNA content of each cell stuck in G2 phase is 4C , coz DNA is duplicated.

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