Chapter 18: Gene Mutation and DNA Repair Student Learning Objectives Upon completion of this chapter you should be able to: 1. 2. 3. 4. 5. 6.

Know the different classes of mutations and their effects upon the organism. Understand the key experiments that support the random mutation theory. Recognize how DNA can undergo spontaneous mutations. Know the different causes of induced mutations. Distinguish between mutation rate and mutation frequency. Understand DNA repair mechanisms and the types of mutations they recognize.

18.1 Effects of Mutations on Gene Structure and Function Overview The term mutation typically is associated with a negative change in the genetic material. However, detrimental or lethal mutations are actually but one class of mutations. A mutation represents any heritable change to the genetic material and can occur at the chromosome or gene levels. Chromosome variations were discussed in Chapter 8. This chapter examines mutations affecting single genes. The purpose of this first section is to establish the basic types of mutations and the terminology associated with the study of mutations. There is a significant amount of descriptive terminology. Table 18.1 provides a very good overview. It is important that you have a good understanding of this material before proceeding into the remainder of the chapter. Before you finish reading a section on a certain type of mutation, take a good look at the associated figure. As you do so, ask yourself if you understand how this mutation affects the gene or DNA sequence, and what possible effects it may have on the phenotype.

Outline of Key Terms Beneficial mutation Conditional mutants Suppressor mutations (Suppressors) Intergenic suppressor Intragenic suppressor Up promoter mutation Down promoter mutation Changes in chromosome structure Breakpoint effect Position effect Germ line Germ-line mutation Somatic cells Somatic mutation Genetic mosaic

Mutation Point mutation Base substitution Transition Transversion Silent mutation Missense mutation Neutral mutation Nonsense mutation Polar mutation Polarity Frameshift mutation Wild type allele Mutant allele Reversion Deleterious mutation Lethal mutation

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Focal Points   

Consequences of point mutations within the coding sequences (Table 18.1) Causes of position effects (Figure 18.2) Somatic versus germ-line mutations (Figure 18.4)

Exercises and Problems For questions 1 to 10, match the form of mutation with its correct definition. _____ 1. Frameshift mutation _____ 2. Transition _____ 3. Silent mutation _____ 4. Missense mutation _____ 5. Nonsense mutation _____ 6. Point mutation _____ 7. Transversion _____ 8. Genome mutation _____ 9. Chromosome mutation _____ 10. Neutral mutation a. A change in a single base pair in the DNA. b. A mutation that does not change the amino acid sequence of the polypeptide. c. The addition or deletion of one or two nucleotides. d. A change that alters a single amino acid in the polypeptide. e. A physical change in the structure of the chromosome. f. Changes a normal codon to a termination codon. g. A point mutation involving a change from a purine to a purine. h. A type of missense mutation that has no effect on protein function. i. A change in the number of chromosomes. j. A point mutation involving a change from a purine to a pyrimidine. For questions 11 to 20, choose the correct type of mutation that matches the description provided. _____ 11. Increases the chances of survival or the reproductive success of the organism. _____ 12. A second mutation that restores the wild-type condition. _____ 13. A suppressor that occurs within the same gene as the first mutation. _____ 14. Causes death. _____ 15. Decreases the chances of survival, but is not necessarily lethal. _____ 16. Temperature-sensitive mutations.

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a. b. c. d. e. f. g. h. i. j.

intragenic suppressor conditional mutants suppressor mutations reversion beneficial mutation deleterious mutation intergenic suppressor lethal mutation wild type forward mutation

_____ 17. Converts a variant back to the wild-type. _____ 18. A suppressor that occurs in a different gene than the first mutation. _____ 19. Converts the wild-type to a new variant. _____ 20. The most common genotype in the population. For questions 21 to 23, indicate whether the statement is true (T) or false (F). If false, correct the statement. _____ 21. A mutation in a regulatory region of a gene that causes a decrease in the rate of transcription is called an up promoter mutation. _____ 22. A mutation that converts a UAA codon into a UAC codon will make the protein shorter than normal. _____ 23. The movement of a regulatory region due to a chromosomal mutation is called a position effect. For questions 24 to 27, indicate whether the statement is associated with a germ-line mutation (G) or somatic cell mutation (S). _____ 24. May produce a genetic mosaic. _____ 25. Is heritable. _____ 26. Affects all of the cells of the organism. _____ 27. Would not be carried by the gametes.

18.2 Random Nature of Mutations Overview The second section of the chapter explores the debate over whether mutations are physiological adaptations, or represent random changes in the genetic material. These opposing ideas were first proposed in the 19th century. They were tested in bacterial studies in the 1940s and 1950s. Both the Luria-Delbrück fluctuation test (Figure 18.6) and the Lederbergs’ replica plating technique (Figure 18.7) supported the random mutation theory.

Outline of Key Terms Random mutation theory Fluctuation test Replica plating Hot spots

Focal Points  

The Luria-Delbrück fluctuation test (Figure 18.6) The Lederbergs’ replica plating technique (Figure 18.7)

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Exercises and Problems Complete the following statements with the most appropriate word or phrase: 1. The two opposing ideas regarding the nature of mutations have been dubbed ________ and ________. 2. Lamarck’s theory of use and disuse favored the ________ idea. 3. The _______ test of Luria and Delbrück distinguished between the opposing ideas regarding the nature of mutations. 4. Luria and Delbrück investigated the ability of E. coli to resist infection by ________; this resistance is designated by the term ________. 5. In the 1950s, Joshua and Esther Lederberg developed a technique called ________, which involved the use of a velvet cloth to make copies of a master plate. 6. A region of a gene that is more likely to mutate than another region is called a ________.

18.3 Spontaneous Mutations Overview Mutations can be caused in one of two ways. Spontaneous mutations are changes in DNA structure that result from natural biological or chemical processes, whereas induced mutations are caused by environmental agents (Refer to Table 18.4). This section examines spontaneous mutations, and the following will address induced mutations. Spontaneous mutations can arise via several different mechanisms. These include depurination (Figure 18.8), deamination (Figure 18.9), tautomeric shifts (Figure 18.10), and even oxidative stress (Figure 18.11). The last part of this section discusses trinucleotide repeat expansions (TNRE). Once the length of a trinucleotide repeat increases above a certain critical size, it becomes prone to frequent expansion. This phenomenon is called dynamic mutation. The repeats can expand due to hairpin formation during DNA replication (Figure 18.12). TNRE disorders include Huntington disease, fragile X syndrome, and myotonic muscular dystrophy (Refer to Table 18.5).

Outline of Key Terms Trinucleotide repeat expansions Dynamic mutations Anticipation

Induced mutations Spontaneous mutations Depurination Apurinic site Deamination Tautomeric shift Tautomers Reactive oxygen species Oxidative stress Oxidative DNA damage

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Focal Points     

Causes of mutations (Table 18.4) Spontaneous depurination (Figure 18.8) Spontaneous deamination of cytosine and 5-methylcytosine (Figure 18.9) Tautomeric shifts and their ability to cause mutation (Figure 18.10) Proposed mechanism of trinucleotide repeat expansion (Figure 18.12)

Exercises and Problems For questions 1 to 10, complete the sentence with the most appropriate term(s): 1. During spontaneous depurination, the bond between an A or G and the deoxyribose sugar is broken, thereby releasing the base. This leaves a(n) _______ in the DNA. 2. The deamination of cytosine produces ________. 3. The deamination of 5-methylcytosine produces ________. 4. The most common tautomeric form for of guanine and thymine is the ________ form. At a low rate, G and T can interconvert to the _______ form. 5. The most common tautomeric form for of adenine and cytosine is the ________ form. At a low rate, A and C can interconvert to the _______ form. 6. Reactive oxygen species (ROS) include ________, ________, and ________. 7. ________ refers to an imbalance between the production of ROS and an organism’s ability to break them down. 8. The length of a trinucleotide repeat is prone to frequent expansion once it has increased above a certain critical size. This is known as a ________ mutation. 9. Some TNRE disorders have the unusual feature of a progressively worsening severity in future generations. This phenomenon is called ________. 10. Trinucleotide repeats can expand due to ________ formation during DNA replication. For questions 11 to 16, match the statement with the appropriate spontaneous mutation. a. deamination b. tautomeric shift c. depurination _____ 11. Sites where this occurs tend to be hot spots of mutation. _____ 12. A temporary change in base structure. _____ 13. This typically occurs at a cytosine in the DNA. _____ 14. This must occur immediately prior to replication in order to cause a mutation. _____ 15. This is the most common form of spontaneous mutation. _____ 16. Caused by a spontaneous reaction with water which releases the base from the sugar.

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18.4 Induced Mutations Overview Having discussed spontaneous mutations we now move on to induced mutations, which are those caused by environmental agents. These agents are called mutagens, and can be physical or chemical. As you proceed through this section, you should become familiar with the major classes of mutagens, which are listed in Table 18.6, and the different ways they can alter the structure of DNA. It is also necessary for you to understand the difference between mutation rate and mutation frequency. Mutation rate is the likelihood that a gene will be changed by a new mutation, while frequency represents the abundance of the mutant gene in a given population. The final part of this section examines the Ames test (Figure 18.16), which remains a commonly used procedure for determining whether a compound is actually a mutagen.

Outline of Key Terms Mutation rate Mutation frequency Ames test

Mutagens Nitrous acid Nitrogen mustard Ethyl methanesulfonate Acridine dyes Proflavin 2-aminopurine 5-bromouracil Thymine dimers

Focal Points     

Examples of mutagens (Table 18.6) Mispairing of bases modified by nitrous acid (Figure 18.13) Base pairing of 5-bromouracil and its ability to cause mutation (Figure 18.14) Formation and structure of a thymine dimer (Figure 18.15) The Ames test for mutagenicity (Figure 18.16)

Exercises and Problems For questions 1 to 11, complete the sentence with the most appropriate term(s): 1. A(n) _______ mutation is one that is caused by an environmental agent. 2. A mutagen may be either a chemical compound or a ________. 3. The compound 2-aminopurine is a ________ that becomes incorporated into daughter strands during DNA replication. 4. During alkylation, ________ or _______ groups are covalently attached to DNA bases. 5. Gamma rays are an example of ________ radiation.

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6. Ultraviolet light causes the formation of cross-linked ________ in the DNA. 7. The probability that a new mutation will alter a given gene is called the ________. 8. The average mutation rate is _______ to _______ per cell generation. 9. Mutation ________ indicates the prevalence of a specific mutation in the population. 10. The Ames test uses a strain of the bacterium ________. 11. ________ is an alkylating agent that was used as a chemical weapon in World War I.

For questions 12 to 17, match each of the following mutagens with its correct description. _____ 12. Nitrous acid _____ 13. 5-bromouracil _____ 14. Ethyl methanesulfonate _____ 15. Acridine dye _____ 16. X-rays _____ 17. UV light a. b. c. d. e. f.

A nucleotide base analog that enters into the DNA during replication. A form of nonionizing radiation. Distorts the double-helix structure of DNA by integrating into the helix. An alkylating agent. A form of ionizing radiation. Changes cytosine to uracil by adding a keto functional group.

18.5 DNA Repair Overview The final section of this chapter examines the various mechanisms by which cells may repair damage to the DNA caused by spontaneous or induced mutations. This section builds upon the information provided in the earlier sections. A summary of the major mechanisms of DNA repair is given in Table 18.7. Note that each repair system is composed of one or more proteins that play specific roles in the repair mechanisms, and that in most cases, the DNA repair is a multistep process. You will notice below that the focal points for this chapter include every diagram. That is because it is almost impossible to understand DNA repair by reading the text. As you review this material you should frequently refer back to the diagrams. As you do so, remember that these graphics actually represent three-dimensional structures in the cell and what you are observing are three-dimensional interactions.

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Outline of Key Terms DNA double-strand break (DSB) Homologous recombination repair (HRR) Non-homologous end-joining (NHEJ) Transcription-coupled DNA repair Transcription-repair coupling factor (TRCF) Translesion synthesis (TLS) Error-prone replication SOS response

Direct repair Photolyase Photoreactivation Alkyltransferase Base excision repair (BER) DNA-N-glycosylases AP-endonuclease Nick translation Nucleotide excision repair (NER) Base mismatch Mismatch repair system

Focal Points  

DNA repair systems (Table 18.7) DNA repair mechanisms (Figures 18.17-18.19 and 18.21-18.24)

Exercises and Problems For questions 1 to 10, choose the repair mechanism that is best associated with the statement. Some statements may have more than one answer. a. nucleotide excision repair b. mismatch repair c. translesion synthesis d. recombinational repair e. base excision repair _____ 1. Uses an enzyme called the DNA-N-glycosylases. _____ 2. Utilizes a group of proteins called Mut. _____ 3. The disease xeroderma pigmentosum is caused by a defect in this system. _____ 4. This is associated with the SOS response in bacteria. _____ 5. Removes a short segment of DNA around the damaged bases. _____ 6. Removes a single abnormal base from the DNA. _____ 7. This may be used to repair an unreplicated gap in the DNA. _____ 8. Identifies the parental strand by methylation patterns, then uses it as a template for repairing the damage to the unmethylated strand. _____ 9. Abnormal nucleotides are recognized by AP-endonuclease. _____ 10. Uses the Uvr proteins in the response.

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Chapter Quiz 1. Which of the following types of mutations changes a normal codon to a stop codon? a. silent mutation b. frameshift mutation c. nonsense mutation d. missense mutation e. none of the above 2. A _______ mutation returns a variant to the wild type condition. a. forward b. reverse c. lateral d. vertical e. horizontal 3. A mutation that influences the phenotype only under specific environmental conditions is called a ________ mutation a. conditional b. silent c. transitional d. missense 4. Which of the following enzymes can DIRECTLY repair thymine dimers? a. alkyltransferase b. pyrimidine kinase c. phosphatase d. DNA polymerase e. photolyase 5. Which of the following mechanisms may cause spontaneous mutations? a. deamination b. depurination c. tautomeric shift d. A and B e. A, B, and C 6. Ionizing radiation includes which of the following? a. UV light and X-rays b. UV light and gamma rays c. Gamma rays and X-rays d. UV light, gamma rays, and X-rays 7. Which of the following tests is used to determine if a chemical is a mutagen? a. fluctuation test b. replica plating c. tautomeric test d. Ames test

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8. Which of the following compounds is an example of a base analog? a. acridine dyes b. ethyl methanesulfonate c. 5-bromouracil d. nitrous acid e. nitrogen mustard 9. Gaps in the DNA are best repaired by which of the following repair mechanisms? a. base excision repair b. nucleotide excision repair c. SOS response d. recombinational repair 10. DNA-N-glycosylases and AP-endonucleases are used in which of the following systems? a. base excision repair b. nucleotide excision repair c. recombinational repair d. thymine dimer repair

Answer Key for Study Guide Questions This answer key provides the answers to the exercises and chapter quiz for this chapter. Answers in parentheses ( ) represent possible alternate answers to a problem, while answers marked with an asterisk (*) indicate that the response to the question may vary. 15. f 16. b 17. d 18. g 19. j 20. i 21. F, down promoter mutation 22. F, longer 23. T 24. S 25. G 26. G 27. S

18.1

1. c 2. g 3. b 4. d 5. f 6. a 7. j 8. i 9. e 10. h 11. e 12. c 13. a 14. h

18.2

1. physical adaptation and random mutation 2. physical adaptation 3. fluctuation 4. bacteriophage T1; tonr 5. replica plating 6. hot spots

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18.3

1. apurinic site 2. uracil 3. thymine 4. keto; enol 5. amino; imino 6. hydrogen peroxide, superoxide, hydroxyl radical 7. oxidative stress 8. dynamic

9. anticipation 10. hair-pin (stem-loop) 11. a 12. b 13. a 14. b 15. c 16. c

18.4

1. induced 2. physical agent 3. base analog 4. methyl or ethyl 5. ionizing 6. thymine (pyrimidine) dimers 7. mutation rate 8. 10-5, 10-9 9. frequency

10. Salmonella typhimurium 11. Nitrogen mustard (mustard gas) 12. f 13. a 14. d 15. c 16. e 17. b

18.5

1. e 2. b 3. a 4. c 5. a

6. e 7. d 8. b 9. e 10. a

Quiz 1. c 2. b 3. a 4. e 5. e 6. c 7. d 8. c 9. d 10. a

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