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Ch. 16 - Regulation of Gene Expression in Bacteria
Chapter 16, Problem 20

A bacterial operon is responsible for the production of the biosynthetic enzymes needed to make the hypothetical amino acid tisophane (tis). The operon is regulated by a separate gene, R. The deletion of R causes the loss of enzyme synthesis. In the wild-type condition, when tis is present, no enzymes are made; in the absence of tis, the enzymes are made. Mutations in the operator gene (O⁻) result in repression regardless of the presence of tis. Is the operon under positive or negative control? Propose a model for (a) repression of the genes in the presence of tis in wild-type cells and (b) the mutations.

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Identify the type of control: Since the deletion of the regulatory gene R leads to loss of enzyme synthesis, this suggests that the operon is under negative control. In negative control, a repressor protein (encoded by R) binds to the operator to prevent transcription.
Model the wild-type repression: In the presence of tis, the repressor protein (produced by gene R) is active and binds to the operator region, preventing RNA polymerase from transcribing the operon genes, thus no enzymes are produced.
Model the wild-type activation: In the absence of tis, the repressor protein is inactive or unable to bind to the operator, allowing RNA polymerase to transcribe the operon genes, leading to enzyme production.
Explain the operator mutation (O⁻): The mutation in the operator gene (O⁻) results in a change that allows the repressor to bind to the operator regardless of the presence or absence of tis, leading to constant repression of the operon.
Summarize the model: The operon is negatively controlled by a repressor that is active in the presence of tis, preventing transcription. The operator mutation causes constitutive repression, overriding the normal regulatory mechanism.

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Key Concepts

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

Operon Structure and Function

An operon is a cluster of genes under the control of a single promoter, allowing coordinated regulation of gene expression. In bacteria, operons typically consist of structural genes, a promoter, and an operator. The operator is a regulatory sequence where repressor proteins can bind, influencing whether transcription occurs. Understanding operon structure is crucial for analyzing how genes are turned on or off in response to environmental signals.
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Negative Control in Gene Regulation

Negative control refers to the mechanism by which a repressor protein binds to the operator region of an operon, preventing transcription of the downstream genes. In the context of the question, the presence of the amino acid tisophane (tis) leads to the binding of a repressor, inhibiting enzyme synthesis. This mechanism is essential for conserving resources, as the cell only produces enzymes when tis is absent, highlighting the operon's regulatory response to metabolic needs.
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Mutations and Their Effects on Gene Regulation

Mutations in the operator gene can alter the binding affinity of the repressor, leading to changes in gene expression. In the case of the O⁻ mutation mentioned, the operator is modified such that the repressor cannot bind, resulting in continuous repression of the operon regardless of tis presence. This illustrates how genetic mutations can disrupt normal regulatory mechanisms, providing insights into the operon's control and the consequences of genetic alterations.
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Related Practice
Textbook Question
Keeping in mind the life cycle of bacteriophages discussed earlier in the text (see Chapter 6), consider the following problem: During the reproductive cycle of a temperate bacteriophage, the viral DNA inserts into the bacterial chromosome where the resultant prophage behaves much like a Trojan horse. It can remain quiescent, or it can become lytic and initiate a burst of progeny viruses. Several operons maintain the prophage state by interacting with a repressor that keeps the lytic cycle in check. Insults (ultraviolet light, for example) to the bacterial cell lead to a partial breakdown of the repressor, which in turn causes the production of enzymes involved in the lytic cycle. As stated in this simple form, would you consider this system of regulation to be operating under positive or negative control?
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Textbook Question
Bacterial strategies to evade natural or human-imposed antibiotics are varied and include membrane-bound efflux pumps that export antibiotics from the cell. A review of efflux pumps [Grkovic, S., et al. (2002)] states that, because energy is required to drive the pumps, activating them in the absence of the antibiotic has a selective disadvantage. The review also states that a given antibiotic may play a role in the regulation of efflux by interacting with either an activator protein or a repressor protein, depending on the system involved. How might such systems be categorized in terms of negative control (inducible or repressible) or positive control (inducible or repressible)?
356
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Textbook Question
In a theoretical operon, genes A, B, C, and D represent the repressor gene, the promoter sequence, the operator gene, and the structural gene, but not necessarily in the order named. This operon is concerned with the metabolism of a theoretical molecule (tm). From the data provided in the accompanying table, first decide whether the operon is inducible or repressible. Then assign A, B, C, and D to the four parts of the operon. Explain your rationale. (AE=active enzyme; IE=inactive enzyme; NE=no enzyme.) Genotype tm Present tm Absent A⁺B⁺C⁺D⁺ AE NE A⁻B⁺C⁺D⁺ AE AE A⁺B⁻C⁺D⁺ NE NE A⁺B⁺C⁻D⁺ IE NE A⁺B⁺C⁺D⁻ AE AE A⁻B⁺C⁺D⁺/F'A⁺B⁺C⁺D⁺ AE AE A⁺B⁻C⁺D⁺/F'A⁺B⁺C⁺D⁺ AE NE A⁺B⁺C⁻D⁺/F'A⁺B⁺C⁺D⁺ AE+IE NE A⁺B⁺C⁺D⁻/F'A⁺B⁺C⁺D⁺ AE NE
594
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Textbook Question
A marine bacterium is isolated and shown to contain an inducible operon whose genetic products metabolize oil when it is encountered in the environment. Investigation demonstrates that the operon is under positive control and that there is a reg gene whose product interacts with an operator region (o) to regulate the structural genes, designated sg. In an attempt to understand how the operon functions, a constitutive mutant strain and several partial diploid strains were isolated and tested with the results shown in the following table. Host Chromosome F' Factor Phenotype Wild type None Inducible Wild type reg gene from mutant strain Inducible Wild type Operon from mutant strain Constitutive Mutant strain reg gene from wild type Constitutive Draw all possible conclusions about the mutation as well as the nature of regulation of the operon. Is the constitutive mutation in the trans-acting reg element or in the cis-acting o operator element?
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Textbook Question
The SOS repair genes in E. coli (discussed in Chapter 15) are negatively regulated by the lexA gene product, called the LexA repressor. When a cell's DNA sustains extensive damage, the LexA repressor is inactivated by the recA gene product (RecA), and transcription of the SOS genes is increased dramatically. One of the SOS genes is the uvrA gene. You are a student studying the function of the uvrA gene product in DNA repair. You isolate a mutant strain that shows constitutive expression of the UvrA protein. Naming this mutant strain uvrAᶜ, you construct the diagram shown above in the right-hand column showing the lexA and uvrA operons: Describe two different mutations that would result in a uvrA constitutive phenotype. Indicate the actual genotypes involved. (Leader sequence for Problem 24 above)
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Textbook Question
Figure 16.13 depicts numerous critical regions of the leader sequence of mRNA that play important roles during the process of attenuation in the trp operon. A closer view of the leader sequence, which begins at about position 30 downstream from the 5' end, is shown below, running along both columns. Within this molecule are the sequences that cause the formation of the alternative hairpins. It also contains the successive triplets that encode tryptophan, where stalling during translation occurs. Take a large piece of paper (such as manila wrapping paper) and, along with several other students from your genetics class, work through the base sequence to identify the trp codons and the parts of the molecule representing the base-pairing regions that form the terminator and antiterminator hairpins shown in Figure 16.13.
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