Ch. 8 - Molecular Biology of Transcription and RNA Processing

Sanders - Genetic Analysis: An Integrated Approach 3rd Edition

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Sanders 3rd Edition

Ch. 8 - Molecular Biology of Transcription and RNA Processing

Problem 20b

Chapter 8, Problem 20b

Wild-type E. coli grows best at 37°C but can grow efficiently up to 42°C. An E. coli strain has a mutation of the sigma subunit that results in an RNA polymerase holoenzyme that is stable and transcribes at wild-type levels at 37°C. The mutant holoenzyme is progressively destabilized as the temperature is raised, and it completely denatures and ceases to carry out transcription at 42°C. Relative to wild-type growth, characterize the ability of the mutant strain to carry out transcription at 40°C

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Understand the role of the sigma subunit in RNA polymerase: The sigma subunit is a critical component of the RNA polymerase holoenzyme in E. coli. It helps the enzyme recognize and bind to specific promoter sequences to initiate transcription.

Analyze the mutation's effect: The mutation in the sigma subunit causes the RNA polymerase holoenzyme to become progressively destabilized as the temperature increases. At 37°C, the mutant enzyme functions normally, but its stability decreases at higher temperatures, and it completely denatures at 42°C.

Determine the temperature range of interest: The problem asks about the ability of the mutant strain to carry out transcription at 40°C. This temperature is between 37°C (where the mutant enzyme is stable) and 42°C (where it is completely denatured).

Predict the transcriptional activity at 40°C: Since the mutant enzyme is progressively destabilized as the temperature rises, its ability to carry out transcription at 40°C will likely be reduced compared to wild-type levels. However, it may still retain some activity, as it has not yet reached the complete denaturation point of 42°C.

Conclude the relative transcriptional ability: Relative to wild-type E. coli, the mutant strain's transcriptional activity at 40°C will be impaired but not entirely absent. This reduced activity will likely result in slower growth or reduced efficiency compared to the wild-type strain at this temperature.

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Key Concepts

Here are the essential concepts you must grasp in order to answer the question correctly.

RNA Polymerase Function

RNA polymerase is an essential enzyme responsible for synthesizing RNA from a DNA template during transcription. In E. coli, the holoenzyme consists of the core enzyme and a sigma factor that helps initiate transcription at specific promoters. Understanding how mutations in the sigma subunit affect the stability and function of RNA polymerase is crucial for analyzing the transcriptional capabilities of the mutant strain at different temperatures.

Temperature Effects on Enzyme Activity

Enzymes, including RNA polymerase, are sensitive to temperature changes, which can affect their stability and activity. Generally, as temperature increases, enzyme activity may rise to an optimal point before denaturing occurs. In this case, the mutant E. coli strain's ability to transcribe at 40°C must be evaluated in light of how the mutation impacts the holoenzyme's stability at elevated temperatures.

Mutations and Phenotypic Expression

Mutations can lead to changes in protein structure and function, which can manifest as altered phenotypes. In this scenario, the mutation in the sigma subunit of E. coli affects the stability of the RNA polymerase holoenzyme, influencing its transcriptional efficiency at varying temperatures. Understanding the relationship between the genetic mutation and its phenotypic consequences is key to characterizing the mutant strain's performance at 40°C.

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Mutations and Phenotypes

Related Practice

Textbook Question

Wild-type E. coli grow best at 37°C but can grow efficiently up to 42°C. An E. coli strain has a mutation of the sigma subunit that results in an RNA polymerase holoenzyme that is stable and transcribes at wild-type levels at 37°C. The mutant holoenzyme is progressively destabilized as the temperature is raised, and it completely denatures and ceases to carry out transcription at 42°C. Relative to wild-type growth, characterize the ability of the mutant strain to carry out transcription at 37°C

Textbook Question

Wild-type E. coli grows best at 37°C but can grow efficiently up to 42°C. An E. coli strain has a mutation of the sigma subunit that results in an RNA polymerase holoenzyme that is stable and transcribes at wild-type levels at 37°C. The mutant holoenzyme is progressively destabilized as the temperature is raised, and it completely denatures and ceases to carry out transcription at 42°C. Relative to wild-type growth, characterize the ability of the mutant strain to carry out transcription at 42°C

Textbook Question

Wild-type E. coli grows best at 37°C but can grow efficiently up to 42°C. An E. coli strain has a mutation of the sigma subunit that results in an RNA polymerase holoenzyme that is stable and transcribes at wild-type levels at 37°C. The mutant holoenzyme is progressively destabilized as the temperature is raised, and it completely denatures and ceases to carry out transcription at 42°C. Relative to wild-type growth, characterize the ability of the mutant strain to carry out transcription at What term best characterizes the type of mutation exhibited by the mutant bacterial strain? (Hint: The term was used in Chapter 4 to describe the Himalayan allele of the mammalian C gene.)

Textbook Question

A mutant strain of Salmonella bacteria carries a mutation of the rho protein that has full activity at 37°C but is completely inactivated when the mutant strain is grown at 40°C. Speculate about the kind of differences you would expect to see if you compared a broad spectrum of mRNAs from the mutant strain grown at 37°C and the same spectrum of mRNAs from the strain when grown at 40°C.

Textbook Question

A 1.0-kb DNA fragment from the end of the mouse gene described in the previous problem is examined by DNA footprint protection analysis. Two samples are end-labeled with ³²P, and one of the two is mixed with TFIIB, TFIID, and RNA polymerase II. The DNA exposed to these proteins is run in the right-hand lane of the gel shown below and the control DNA is run in the left-hand. Both DNA samples are treated with DNase I before running the samples on the electrophoresis gel.

Draw a diagram of this DNA fragment bound by the transcriptional proteins, showing the approximate position of proteins along the fragment.

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Textbook Question

A 1.0-kb DNA fragment from the end of the mouse gene described in the previous problem is examined by DNA footprint protection analysis. Two samples are end-labeled with ³²P and one of the two is mixed with TFIIB, TFIID, and RNA polymerase II. The DNA exposed to these proteins is run in the right-hand lane of the gel shown below and the control DNA is run in the left-hand. Both DNA samples are treated with DNase I before running the samples on the electrophoresis gel.

Explain the role of DNase I.

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