PROBLEMS

WORKING WITH THE FIGURES

Question 16.1

In Figure 16-3a, what is the consequence of the new 5′ splice site on the open reading frame? In 16-3b, how big could the intron be to maintain the reading frame (let’s say between 75 and 100 bp)?

Question 16.2

Using Figure 16-4 as an example, compare the migration of RNA and protein for the wild-type gene and the mutation shown in Figure 16-3b. Assume that the retained intron maintains the reading frame.

Question 16.3

In the Ames test shown in Figure 16-17, what is the reason for adding the liver extract to each sample?

Question 16.4

Based on the mode of action of aflatoxin (Figure 16-16), propose a scenario that explains its response in the Ames test (Figure 16-18).

Question 16.5

In Figure 16-22, point out the mutant protein(s) in patients with Cockayne syndrome. What protein(s) is/are mutant in patients with XP? How are these different mutations thought to account for the different disease symptoms?

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Question 16.6

The MutH protein nicks the newly synthesized strand (Figure 16-23). How does it “know” which strand this is?

Question 16.7

What features of the bypass polymerase make it ideal for its role in translesion synthesis, shown in Figure 16-24?

BASIC PROBLEMS

Question 16.8

Consider the following wild-type and mutant sequences:

The substitution shown seems to have created a stop codon. What further information do you need to be confident that it has done so?

Question 16.9

What type of mutation is depicted by the following sequences (shown as mRNA)?

Question 16.10

Can a missense mutation of proline to histidine be made with a G · C → A · T transition-causing mutagen? What about a proline-to-serine missense mutation?

Question 16.11

By base-pair substitution, what are all the synonymous changes that can be made starting with the codon CGG?

Question 16.12

  1. What are all the transversions that can be made starting with the codon CGG?

  2. Which of these transversions will be missense? Can you be sure?

Question 16.13

  1. Acridine orange is an effective mutagen for producing null alleles by mutation. Why does it produce null alleles?

  2. A certain acridine-like compound generates only single insertions. A mutation induced with this compound is treated with the same compound, and some revertants are produced. How is this outcome possible?

Question 16.14

Defend the statement “Cancer is a genetic disease.”

Question 16.15

Give an example of a DNA-repair defect that leads to cancer.

Question 16.16

In mismatch repair in E. coli, only a mismatch in the newly synthesized strand is corrected. How is E. coli able to recognize the newly synthesized strand? Why does this ability make biological sense?

Question 16.17

A mutational lesion results in a sequence containing a mismatched base pair:

If mismatch repair occurs in either direction, which amino acids could be found at this site?

Question 16.18

Under what circumstances could nonhomologous end joining be said to be error prone?

Question 16.19

Why are many chemicals that test positive by the Ames test also classified as carcinogens?

Question 16.20

The Spo11 protein is conserved in eukaryotes. Do you think it is also conserved in bacterial species? Justify your answer.

Question 16.21

Differentiate between the elements of the following pairs:

  1. Transitions and transversions

  2. Synonymous and neutral mutations

  3. Missense and nonsense mutations

  4. Frameshift and nonsense mutations

Question 16.22

Describe two spontaneous lesions that can lead to mutations.

Question 16.23

What are bypass polymerases? How do they differ from the replicative polymerases? How do their special features facilitate their role in DNA repair?

Question 16.24

In adult cells that have stopped dividing, what types of repair systems are possible?

Question 16.25

A certain compound that is an analog of the base cytosine can become incorporated into DNA. It normally hydrogen bonds just as cytosine does, but it quite often isomerizes to a form that hydrogen bonds as thymine does. Do you expect this compound to be mutagenic, and, if so, what types of changes might it induce at the DNA level?

Question 16.26

Two pathways, homologous recombination and nonhomologous end joining (NHEJ), can repair double-strand breaks in DNA. If homologous recombination is an error-free pathway whereas NHEJ is not always error free, why is NHEJ used most of the time in eukaryotes?

Question 16.27

Which repair pathway recognizes DNA damage during transcription? What happens if the damage is not repaired?

Question 16.28

Where in a gene would a 4-bp insertion mutation have the least effect on gene expression?

  1. Introns

  2. Exons

  3. Regulatory regions

  4. Introns and exons

Question 16.29

Which of the following gene mutations is most likely to have the most severe impact on gene expression?

  1. A nonsense mutation in the last exon

  2. A point mutation in an exon

  3. A point mutation in the splice donor site of an intron

  4. A point mutation in the middle of an intron

Question 29.30

Which of the following is not possible?

  1. A nonsynonymous mutation in an intron

  2. A nonsynonymous mutation in an exon

  3. An indel mutation in an intron

  4. An indel mutation in an exon

Question 16.31

Which of the following is/are associated with spontaneous mutation?

  1. An occurrence of lung cancer due to smoking

  2. A nonsense mutation in an exon caused by an error in DNA replication

  3. An indel mutation in an intron caused by replication slippage

  4. A nonsense mutation in an exon caused by an error in DNA replication, and an indel mutation in an intron caused by replication slippage

Question 16.32

Which of the following statements best describe the mismatch repair pathway?

  1. It is part of the 3′ to 5′ proofreading function of DNA polymerase.

  2. It acts after DNA replication by recognizing mismatched base pairs.

  3. It is activated by stalled replication forks.

  4. It is coupled to transcription.

CHALLENGING PROBLEMS

Question 16.33

  1. Why is it impossible to induce nonsense mutations (represented at the mRNA level by the triplets UAG, UAA, and UGA) by treating wild-type strains with mutagens that cause only A · T → G · C transitions in DNA?

  2. Hydroxylamine (HA) causes only G · C → A · T transitions in DNA. Will HA produce nonsense mutations in wild-type strains?

  3. Will HA treatment revert nonsense mutations?

Question 16.34

Several auxotrophic point mutants in Neurospora are treated with various agents to see if reversion will take place. The following results were obtained (a plus sign indicates reversion; HA causes only G · C → A · T transitions).

Mutant

5-BU

HA

Proflavin

Spontaneous reversion

1

−

−

−

−

2

−

−

+

+

3

+

−

−

+

4

−

−

−

+

5

+

+

−

+

  1. For each of the five mutants, describe the nature of the original mutation event (not the reversion) at the molecular level. Be as specific as possible.

  2. For each of the five mutants, name a possible mutagen that could have caused the original mutation event. (Spontaneous mutation is not an acceptable answer.)

  3. In the reversion experiment for mutant 5, a particularly interesting prototrophic derivative is obtained. When this type is crossed with a standard wild-type strain, the progeny consist of 90 percent prototrophs and 10 percent auxotrophs. Give a full explanation for these results, including a precise reason for the frequencies observed.

Question 16.35

You are using nitrosoguanidine to “revert” mutant nic-2 (nicotinamide-requiring) alleles in Neurospora.

You treat cells, plate them on a medium without nicotinamide, and look for prototrophic colonies. You obtain the following results for two mutant alleles. Explain these results at the molecular level, and indicate how you would test your hypotheses.

  1. With nic-2 allele 1, you obtain no prototrophs at all.

  2. With nic-2 allele 2, you obtain three prototrophic colonies A, B, and C, and you cross each separately with a wild-type strain. From the cross prototroph A × wild type, you obtain 100 progeny, all of which are prototrophic. From the cross prototroph B × wild type, you obtain 100 progeny, of which 78 are prototrophic and 22 are nicotinamide requiring. From the cross prototroph C × wild type, you obtain 1000 progeny, of which 996 are prototrophic and 4 are nicotinamide requiring.

Question 16.36

You are working with a newly discovered mutagen, and you wish to determine the base change that it introduces into DNA. Thus far, you have determined that the mutagen chemically alters a single base in such a way that its base-pairing properties are altered permanently. To determine the specificity of the alteration, you examine the amino acid changes that take place after mutagenesis. A sample of what you find is shown here:

Original:

Gln–His–Ile–Glu–Lys

Mutant:

Gln–His–Met–Glu–Lys

Original:

Ala–Val–Asn–Arg

Mutant:

Ala–Val–Ser–Arg

Original:

Arg–Ser–Leu

Mutant:

Arg–Ser–Leu–Trp–Lys–Thr–Phe

What is the base-change specificity of the mutagen?

Question 16.37

You now find an additional mutant from the experiment in Problem 36:

Original:

Ile–Leu–His–Gln

Mutant:

Ile–Pro–His–Gln

Could the base-change specificity in your answer to Problem 36 account for this mutation? Why or why not?

Question 16.38

You are an expert in DNA-repair mechanisms. You receive a sample of a human cell line derived from a woman who has symptoms of xeroderma pigmentosum. You determine that she has a mutation in a gene that has not been previously associated with XP. How is this possible?

Question 16.39

Ozone (O3) is an important naturally occurring component in our atmosphere, where it forms a layer that absorbs UV radiation. A hole in the ozone layer was discovered in the 1970s over Antarctica and Australia. The hole appears seasonally and was found to be due to human activity. Specifically, ozone is destroyed by a class of chemicals (called CFCs for chlorofluorocarbons) that are found in refrigerants, air-conditioning systems, and aerosols.

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As a scientist working on DNA-repair mechanisms, you discover that there has been a significant increase in skin cancer in the beach communities in Australia. A newspaper reporter friend offers to let you publish a short note (a paragraph) in which you are to describe the possible connection between the ozone hole and the increased skin cancers. On the basis of what you have learned about DNA repair in this chapter, write a paragraph that explains the mechanistic connection.