This question is related to how growth rate affect the interpretation of a study

Question

This question is related to how growth rate affect the interpretation of a study with respect to gene expression reporter.

You want to study the various domain of a transcription factor, TF605. TF605 has been shown to be an activator/enhancer. Your strategy is to first obtain the promoter that the TF605 binds and places it in from of a reporter gene, such as green fluorescence protein, in a plasmid. Next you will clone various mutants of the TF605 in an expression plasmid and transform it into the cell. You made 4 version of TF605.

WT Full length TF605

Mutation that is believed to disrupt DNA binding

Mutation that is believed to disrupt binding to transcription initiation complex (activating domain)

Mutation that is between the DNA binding and activating domain

You transformed each version of TF605 in different transformation reaction, and picked a colony from each plate and grew them in liquid culture. Next you took out the same number of cells for each version of TF and measured the GFP fluorescence intensity.

A. What do you think data would look like if the assumption for version 2 and 3 is correct and the mutation in version 4 didn’t affect the function of TF605 in any meaningful way? Can you draw a bar chart of what the data will look like? For example, the y-axis is the GFP fluorescence intensity and the x-axis is the different version of TF605.

B. Assuming the GFP degrades with a first order kinetics with a rate constant of kd. Also the cell’s growth rate is mu. The functional form of the GFP synthesis by TF605 follow a simple hill equation with hill coefficient of 2, maximum synthesis rate of alpha, and concentration of TF to achieve half maximal activity is Km. Can you write out a differential equation describing the synthesis of GFP?

C. What is the equation for steady state GFP concentration?

D. You realize that the cells that carry version 4 of TF605 actually grow slower than the other. If the GFP level for version 1 and version 4 is similar, and assuming that version 4 does not affect GFP degradation and that the degradation rate is very slow compared to the growth rate, what does that mean in terms of TF605 version 4’s ability to induce GFP expression?

Explanation / Answer

Based on the given data, the function of transcription factor TF605 is activator/enhancer of gene expression. The assumptions of four versions of TF605 are:

A)

If the assumption for version 2 and 3 is correct, the expression of reporter gene would not takes place. If the mutation in version 4 didn’t affect the function of TF605 in any meaningful way, then the gene expression of GFP takes place properly.   

The graph would look like,

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