CHAPTER 16 GENE MUTION AND DNA REPAIR Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

INTRODUCTION The term mutation refers to a heritable change in the genetic material Mutations provide allelic variations On the positive side, mutations are the foundation for evolutionary change On the negative side, mutations are the cause of many diseases Since mutations can be quite harmful, organisms have developed ways to repair damaged DNA 16-2 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

16.1 CONSEQUENCES OF MUTATIONS Mutations can be divided into three main types 1. Chromosome mutations Changes in chromosome structure 2. Genome mutations Changes in chromosome number 3. Single-gene mutations Relatively small changes in DNA structure that occur within a particular gene Types 1 and 2 were discussed in chapter 8 Type 3 will be discussed in this chapter 16-3 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Gene Mutations Change the DNA Sequence A point mutation is a change in a single base pair It involves a base substitution 5’ AACGCTAGATC 3’ 3’ TTGCGATCTAG 5’ 5’ AACGCGAGATC 3’ 3’ TTGCGCTCTAG 5’ A transition is a change of a pyrimidine (C, T) to another pyrimidine or a purine (A, G) to another purine A transversion is a change of a pyrimidine to a purine or vice versa Transitions are more common than transversions 16-4 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Gene Mutations Change the DNA Sequence Mutations may also involve the addition or deletion of short sequences of DNA 5’ AACGCTAGATC 3’ 3’ TTGCGATCTAG 5’ 5’ AACGCGC 3’ 3’ TTGCGCG 5’ Deletion of four base pairs 5’ AACGCTAGATC 3’ 3’ TTGCGATCTAG 5’ 5’ AACAGTCGCTAGATC 3’ 3’ TTGTCAGCGATCTAG 5’ Addition of four base pairs 16-5 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Gene Mutations Can Alter the Coding Sequence Within a Gene Mutations in the coding sequence of a structural gene can have various effects on the polypeptide Silent mutations are those base substitutions that do not alter the amino acid sequence of the polypeptide Due to the degeneracy of the genetic code Missense mutations are those base substitutions in which an amino acid change does occur Example: Sickle-cell anemia (Refer to Figure 16.1) If the substituted amino acids have similar chemistry, the mutation is said to be neutral 16-6 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Gene Mutations Can Alter the Coding Sequence Within a Gene Mutations in the coding sequence of a structural gene can have various effects on the polypeptide Nonsense mutations are those base substitutions that change a normal codon to a termination codon Frameshift mutations involve the addition or deletion of nucleotides in multiples of one or two This shifts the reading frame so that a completely different amino acid sequence occurs downstream from the mutation Table 16.1 describes all of the above mutations 16-7 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

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Gene Mutations and Their Effects on Genotype and Phenotype In a natural population, the wild-type is the most common genotype A forward mutation changes the wild-type genotype into some new variation If it is beneficial, it may move evolution forward Otherwise, it will be probably eliminated from a population A reverse mutation has the opposite effect It is also termed a reversion 16-9 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Some mutations are called conditional mutants Mutations can also be described based on their effects on the wild-type phenotype When a mutation alters an organism’s phenotypic characteristics, it is said to be a variant Variants are often characterized by their differential ability to survive Deleterious mutations decrease the chances of survival The most extreme are lethal mutations Beneficial mutations enhance the survival or reproductive success of an organism Some mutations are called conditional mutants They affect the phenotype only under a defined set of conditions 16-10 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Suppressor mutations are classified into two types A second mutation will sometimes affect the phenotypic expression of another These second-site mutations are called suppressor mutations or simply suppressors Suppressor mutations are classified into two types Intragenic suppressors The second mutant site is within the same gene as the first mutation Intergenic suppressors The second mutant site is in a different gene from the first mutation 16-11 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Gene Mutations in Noncoding Sequences These mutations can still affect gene expression A mutation, may alter the sequence within a promoter Up promoter mutations make the promoter more like the consensus sequence They may increase the rate of transcription Down promoter mutations make the promoter less like the consensus sequence They may decrease the rate of transcription A mutation can also alter splice junctions in eukaryotes Refer to Table 16.2 for other examples 16-12 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

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Changes in Chromosome Structure Can Affect Gene Expression A chromosomal rearrangement may affect a gene because the break occurred in the gene itself A gene may be left intact, but its expression may be altered because of its new location This is termed a position effect There are two common reasons for position effects: 1. Movement to a position next to regulatory sequences Refer to Figure 16.2a 2. Movement to a position in a heterochromatic region Refer to Figure 16.2b AND 16.3 16-19 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Regulatory sequences are often bidirectional Figure 16.2 16-20 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Mutations Can Occur in Germ-Line or Somatic Cells Geneticists classify the animal cells into two types Germ-line cells Cells that give rise to gametes such as eggs and sperm Somatic cells All other cells Germ-line mutations are those that occur directly in a sperm or egg cell, or in one of their precursor cells Refer to Figure 16.4a Somatic mutations are those that occur directly in a body cell, or in one of its precursor cells Refer to Figure 16.4b AND 16.5 16-21 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

The size of the patch will depend on the timing of the mutation The earlier the mutation, the larger the patch An individual who has somatic regions that are genotypically different from each other is called a genetic mosaic Therefore, the mutation can be passed on to future generations Therefore, the mutation cannot be passed on to future generations Figure 16.4 16-22

16.2 OCCURRENCE AND CAUSES OF MUTATION Mutations can occur spontaneously or be induced Spontaneous mutations Result from abnormalities in cellular/biological processes Errors in DNA replication, for example Induced mutations Caused by environmental agents Agents that are known to alter DNA structure are termed mutagens These can be chemical or physical agents Refer to Table 16.4 16-23 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

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Spontaneous Mutations Are Random Events Are mutations spontaneous occurrences or causally related to environmental conditions? This is a question that biologists have asked themselves for a long time  Jean Baptiste Lamarck Proposed that physiological events (e.g. use and disuse) determine whether traits are passed along to offspring Charles Darwin Proposed that genetic variation occurs by chance Natural selection results in better-adapted organisms 16-25 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Contain many mutations at exactly the same site within the gene Figure 6.20 16-32 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Mutation Rates and Frequencies The mutation frequency for a gene is the number of mutant genes divided by the total number of genes in a population If 1 million bacteria were plated and 10 were mutant The mutation frequency would be 1 in 100,000 or 10-5 The mutation frequency depends not only on the mutation rate, but also on the Timing of the mutation Likelihood that the mutation will be passed on to future generations 16-33 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Causes of Spontaneous Mutations Spontaneous mutations can arise by three types of chemical changes 1. Depurination 2. Deamination 3. Tautomeric shift The most common 16-34 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Causes of Spontaneous Mutations Depurination involves the removal of a purine (guanine or adenine) from the DNA The covalent bond between deoxyribose and a purine base is somewhat unstable It occasionally undergoes a spontaneous reaction with water that releases the base from the sugar This is termed an apurinic site Fortunately, apurinic sites can be repaired However, if the repair system fails, a mutation may result during subsequent rounds of DNA replication 16-35 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

16-36 Figure 16.8 Spontaneous depurination Three out of four (A, T and G) are the incorrect nucleotide There’s a 75% chance of a mutation Spontaneous depurination Figure 16.8 16-36

Deamination involves the removal of an amino group from the cytosine base The other bases are not readily deaminated Figure 16.9 DNA repair enzymes can recognize uracil as an inappropriate base in DNA and remove it However, if the repair system fails, a C-G to A-T mutation will result during subsequent rounds of DNA replication 16-37 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Deamination of 5-methyl cytosine can also occur Figure 16.9 Thymine is a normal constituent of DNA This poses a problem for repair enzymes They cannot determine which of the two bases on the two DNA strands is the incorrect base For this reason, methylated cytosine bases tend to create hot spots for mutation 16-38 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

These rare forms promote AC and GT base pairs A tautomeric shift involves a temporary change in base structure (Figure 16.10a) The common, stable form of thymine and guanine is the keto form At a low rate, T and G can interconvert to an enol form The common, stable form of adenine and cytosine is the amino form At a low rate, A and C can interconvert to an imino form These rare forms promote AC and GT base pairs Refer to Figure 16.10b For a tautomeric shift to cause a mutation it must occur immediately prior to DNA replication Refer to Figure 16.10c 16-39 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Common Rare Figure 16.10 16-40

Figure 16.10 16-41

Shifted back to its normal fom Temporary tautomeric shift Figure 16.10 16-42

Types of Mutagens An enormous array of agent can act as mutagens to permanently alter the structure of DNA The public is concerned about mutagens for two main reasons: 1. Mutagens are often involved in the development of human cancers 2. Mutagens can cause gene mutations that may have harmful effects in future generations Mutagenic agents are usually classified as chemical or physical mutagens Refer to Table 16.5 16-52 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Mutagens Alter DNA Structure in Different Ways Chemical mutagens come into three main types 1. Base modifiers 2. Base analogues 3. Intercalating agents 16-54 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Base modifiers covalently modify the structure of a nucleotide For example, nitrous acid, replaces amino groups with keto groups (–NH2 to =O) This can change cytosine to uracil and adenine to hypoxanthine These modified bases do not pair with the appropriate nucleotides in the daughter strand during DNA replication Refer to Figure 16.13 Some chemical mutagens disrupt the appropriate pairing between nucleotides by alkylating bases within the DNA Examples: Nitrogen mustards and ethyl methanesulfonate (EMS) 16-55 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

These mispairings create mutations in the newly replicated strand Mispairing of modified bases Figure 16.13 16-56 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Intercalating agents contain flat planar structures that intercalate themselves into the double helix This distorts the helical structure When DNA containing these mutagens is replicated, the daughter strands may contain single-nucleotide additions and/or deletions Examples: Acridine dyes Proflavin 16-57 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

This tautomeric shift occurs at a relatively high rate Base analogues become incorporated into daughter strands during DNA replication For example, 5-bromouracil is a thymine analogue It can be incorporated into DNA instead of thymine Normal pairing Mispairing This tautomeric shift occurs at a relatively high rate Figure 16.14 16-58 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

In this way, 5-bromouracil can promote a change of an AT base pair into a GC base pair Figure 16.14 16-59 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Physical mutagens come into two main types 1. Ionizing radiation 2. Nonionizing radiation Ionizing radiation Includes X rays and gamma rays Has short wavelength and high energy Can penetrate deeply into biological molecules Creates chemically reactive molecules termed free radicals Can cause Base deletions Single nicks in DNA strands Cross-linking Chromosomal breaks 16-60 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Nonionizing radiation Includes UV light Has less energy Cannot penetrate deeply into biological molecules Causes the formation of cross-linked thymine dimers Thymine dimers may cause mutations when that DNA strand is replicated Figure 16.15 16-61 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Testing Methods Can Determine If an Agent Is a Mutagen Many different kinds of tests have been used to evaluate mutagenicity One commonly used test is the Ames test Developed by Bruce Ames The test uses a strain of Salmonella typhimurium that cannot synthesize the amino acid histidine It has a point mutation in a gene involved in histidine biosynthesis A second mutation (i.e., a reversion) may occur restoring the ability to synthesize histidine The Ames test monitors the rate at which this second mutation occurs 16-62 Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display

Provides a mixture of enzymes that may activate a mutagen The control plate indicates that there is a low level of spontaneous mutation The Ames test for mutagenicity Figure 16.16 16-63