Ch. 16 - Regulation of Gene Expression in Bacteria
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Problem 1
In this chapter, we focused on the regulation of gene expression in bacteria. Along the way, we found many opportunities to consider the methods and reasoning by which much of this information was acquired. From the explanations given in the chapter, what answers would you propose to the following fundamental questions? What evidence established that lactose serves as the inducer of a gene whose product is related to lactose metabolism?Problem 1
In this chapter, we focused on the regulation of gene expression in bacteria. Along the way, we found many opportunities to consider the methods and reasoning by which much of this information was acquired. From the explanations given in the chapter, what answers would you propose to the following fundamental questions? What led researchers to conclude that a repressor molecule regulates the lac operon?Problem 1
In this chapter, we focused on the regulation of gene expression in bacteria. Along the way, we found many opportunities to consider the methods and reasoning by which much of this information was acquired. From the explanations given in the chapter, what answers would you propose to the following fundamental questions? How do we know that the lac repressor is a protein?Problem 1
In this chapter, we focused on the regulation of gene expression in bacteria. Along the way, we found many opportunities to consider the methods and reasoning by which much of this information was acquired. From the explanations given in the chapter, what answers would you propose to the following fundamental questions? How do we know that bacteria regulate the expression of certain genes in response to the environment?Problem 1
In this chapter, we focused on the regulation of gene expression in bacteria. Along the way, we found many opportunities to consider the methods and reasoning by which much of this information was acquired. From the explanations given in the chapter, what answers would you propose to the following fundamental questions? How do we know that the trp operon is a repressible control system, in contrast to the lac operon, which is an inducible control system?Problem 2
Write a brief essay that discusses why you think regulatory systems evolved in bacteria (i.e., what advantages do regulatory systems provide to these organisms?), and, in the context of regulation, discuss why genes related to common functions are found together in operons.Problem 3
Contrast positive versus negative control of gene expression.Problem 4
Contrast the role of the repressor in an inducible system and in a repressible system.Problem 5
For the lac genotypes shown in the following table, predict whether the structural genes (Z) are constitutive, permanently repressed, or inducible in the presence of lactose. Genotype Constitutive Repressed Inducible I⁺O⁺Z⁺ x I⁻O⁺Z⁺ I⁻OᶜZ⁺ I⁻OᶜZ⁺/F'O⁺ I⁺OᶜZ⁺/F'O⁺ IˢO⁺Z⁺ IˢO⁺Z⁺/F'I⁺Problem 6
For the genotypes and conditions (lactose present or absent) shown in the following table, predict whether functional enzymes, nonfunctional enzymes, or no enzymes are made.Problem 7
The locations of numerous lacI⁻ and lacIˢ mutations have been determined within the DNA sequence of the lacI gene. Among these, lacI⁻ mutations were found to occur in the 5′-upstream region of the gene, while lacIˢ mutations were found to occur farther downstream in the gene. Are the locations of the two types of mutations within the gene consistent with what is known about the function of the repressor that is the product of the lacI gene?Problem 8
Describe the experimental rationale that allowed the lac repressor to be isolated.Problem 9
What properties demonstrate that the lac repressor is a protein? Describe the evidence that it indeed serves as a repressor within the operon system.Problem 10
Predict the effect on the inducibility of the lac operon of a mutation that disrupts the function of (a) the crp gene, which encodes the CAP protein, and (b) the CAP-binding site within the promoter.Problem 11
Erythritol, a natural sugar abundant in fruits and fermenting foods, is about 65 percent as sweet as table sugar and has about 95 percent fewer calories. It is 'tooth friendly' and generally devoid of negative side effects as a human consumable product. Pathogenic Brucella strains that catabolize erythritol contain four closely spaced genes, all involved in erythritol metabolism. One of the four genes (eryD) encodes a product that represses the expression of the other three genes. Erythritol catabolism is stimulated by erythritol. Present a simple regulatory model to account for the regulation of erythritol catabolism in Brucella. Does this system appear to be under inducible or repressible control?Problem 12
Describe the role of attenuation in the regulation of tryptophan biosynthesis.Problem 13
Attenuation of the trp operon was viewed as a relatively inefficient way to achieve genetic regulation when it was first discovered in the 1970s. Since then, however, attenuation has been found to be a relatively common regulatory strategy. Assuming that attenuation is a relatively inefficient way to achieve genetic regulation, what might explain its widespread occurrence?Problem 14
Neelaredoxin is a 15-kDa protein that is a gene product common in anaerobic bacteria. It has superoxide-scavenging activity, and it is constitutively expressed. In addition, its expression is not further induced during its exposure to O₂ or H₂O₂ [Silva, G. et al. (2001). J. Bacteriol. 183:4413–4420]. What do the terms constitutively expressed and induced mean in terms of neelaredoxin synthesis?Problem 15
The creation of milk products such as cheeses and yogurts is dependent on the conversion by various anaerobic bacteria, including several Lactobacillus species, of lactose to glucose and galactose, ultimately producing lactic acid. These conversions are dependent on both permease and β-galactosidase as part of the lac operon. After selection for rapid fermentation for the production of yogurt, one Lactobacillus subspecies lost its ability to regulate lac operon expression [Lapierre, L., et al. (2002). J. Bacteriol. 184:928–935]. Would you consider it likely that in this subspecies the lac operon is on or off? What genetic events would likely contribute to the loss of regulation as described above?Problem 16
Both attenuation of the trp operon in E. coli and riboswitches in B. subtilis rely on changes in the secondary structure of the leader regions of mRNA to regulate gene expression. Compare and contrast the specific mechanisms in these two types of regulation with that involving short noncoding RNAs (sRNAs).Problem 17
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?Problem 18
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)?Problem 19
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 NEProblem 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.Problem 21
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?Problem 22
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)Problem 24
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.
