Download Brooker Chapter 16

LECTURE 5
Gene Mutation
(Chapter 16.1-16.2)
Slides 1-37
On your own: Slides 38-45
1
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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 needed for a species to adapt to
changes in the environment
– On the negative side, new mutations are much more likely to
be harmful than beneficial to the individual and often are the
cause of diseases
• Understanding the molecular nature of mutations is a
deeply compelling area of research.
• Since mutations can be quite harmful, organisms have
developed ways to repair damaged DNA
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2
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. Gene mutations
• Relatively small change in DNA structure that affects a single
gene
– Type 3 will be discussed in this chapter
3
Gene Mutations Change the
Sequence

DNA
A point mutation is a change in a single base pair

It can involve 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
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4
Gene Mutations Change the
Sequence

DNA
Mutations may also involve the addition or deletion
of short sequences of DNA
5’ AACGCTAGATC 3’
3’ TTGCGATCTAG 5’
5’ AACGCTC 3’
3’ TTGCGAG 5’
Deletion of four base pairs
5’ AACGCTAGATC 3’
3’ TTGCGATCTAG 5’
5’ AACAGTCGCTAGATC 3’
3’ TTGTCAGCGATCTAG 5’
Addition of four base pairs
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5
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 acid has no detectable effect on protein
function, the mutation is said to be neutral. This can occur if the
new amino acid has similar chemistry to the amino acid it replaced
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© Phototake/Alamy
Normal red blood cells
© Phototake/Alamy
10 μm
Sickled red blood cells
10 μm
(a) Micrographs of red blood cells
NORMAL : NH2 – VALINE – HISTIDINE – LEUCINE – THREONINE – PROLINE – GLUTAMIC ACID – GLUTAMIC ACID...
SICKLE
: NH2 – VALINE – HISTIDINE – LEUCINE – THREONINE – PROLINE – VALINE– GLUTAMIC ACID...
CELL
(b) A comparison of the amino acid sequence between normal b-globin and sickle-cell b-globin
Figure 16.1
7
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 stop codon

Frameshift mutations involve the addition or deletion of a
number of nucleotides that is not divisible by three


This shifts the reading frame so that translation of the mRNA
results in a completely different amino acid sequence downstream
of the mutation
Table 16.1 describes all of the above mutations
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8
9
Gene Mutations outside of coding
sequences can still affect phenotype


Mutations in the core promoter can change levels of gene
expression
 Up mutations increase expression. Down mutations
decrease expression
Other important non-coding mutations are in Table 16.2
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10
Gene Mutations and Their Effects on
Genotype and Phenotype

In a natural population, the wild-type is the relatively
prevalent genotype. Genes with multiple alleles may
have two or more wild-types.

A forward mutation changes the wild-type genotype
into some new variation

A reverse mutation changes a mutant allele back to
the wild-type

It is also termed a reversion
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11


Mutations can also be described based on their
effects on the wild-type phenotype
They 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
The environment can affect whether a given mutation is
deleterious or beneficial
Some mutations are conditional


They affect the phenotype only under a defined set of
conditions
An example is a temperature-sensitive mutation
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12
13
Changes in Chromosome Structure
Can Affect Gene Expression


A chromosomal rearrangement may affect a gene
because the chromosomal breakpoint occurs within
the gene
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 heterochromatic region

Refer to Figure 16.2b AND 16.3
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14
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B
A
Coding
sequence
Core
promoter
B
Gene B
A
Regulatory
sequence
Coding
sequence
Core
promoter
Regulatory sequences
are often bidirectional
Inversion
Core promoter
for gene A is
moved next to
regulatory
sequence of
gene B.
Gene A
(a) Position effect due to regulatory sequences
Active
gene
Gene
is now
inactive.
Translocation
Heterochromatic
chromosome
(more compacted)
Euchromatic
chromosome
Translocated
heterochromatic
chromosome
Shortened euchromatic
chromosome
(b) Position effect due to translocation to a heterochromatic
chromosome
Figure 16.2
15
Mutations Can Occur in
Germ-Line or Somatic Cells

Geneticists classify animal cells into two types

Germ-line cells


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


Cells that give rise to gametes such as eggs and sperm
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
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16
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Germ-line
mutation
Gametes
Embryo
Somatic
mutation
The size of the patch
will depend on the
timing of the mutation
The earlier the mutation,
the larger the patch
Therefore, the
mutation can be
passed on to future
generations
Mutation is
found
throughout
the entire
body.
Mature
individual
An individual who has
somatic regions that are
genotypically different
from each other is called
a genetic mosaic
Therefore, the mutation cannot be
passed on to future generations
Half of
the gametes
carry the
mutation.
Figure 16.4
Patch of
affected
area
(a) Germ-line mutation
None of
the gametes
carry the
mutation.
(b) Somatic cell mutation
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18
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
– Underlying cause originates within the cell
• 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
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20
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: Physiological adaptation theory


Proposed that physiological events (e.g. use and disuse)
determine whether traits are passed along to offspring
Charles Darwin: Random mutation theory

Proposed that genetic variation occurs by chance

Natural selection results in better-adapted organisms
21
Random Mutations Can Give an
Organism a Survival Advantage


Joshua and Ester Lederberg(1950s) devised an
ingenious way to test these alterative theories
experimentally
Studied the resistance of E. coli to infection by
bacteriophage T1



tonr (T one resistance)
Hypothesis: E. coli cells that survive T1 infection were
already resistant to the phage prior to exposure
 Due to random mutations
"Replica plating"
22





The Lederbergs'
experiment:
A few tonr colonies were
observed at the same
location on both plates!!!
This indicates that mutations
conferring tonr occurred
randomly on the primary
(nonselective plate)
The presence of T1 in the
secondary plates simply
selected for previously
occurring tonr mutants
This supports the random
mutation theory
Figure 16.7 Replica plating
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Master plate containing
many colonies that were
grown in the absence of
T1 phage
A velvet cloth (wrapped over a
cylinder) is pressed gently onto the
master plate and then lifted. A little
bit of each bacterial colony adheres
to the velvet cloth, thereby creating
a replica of the arrangement of
colonies on the master plate.
Velvet cloth
The replica is then gently pressed
onto 2 secondary plates that
contain T1 phage.
Petri plate
with T1 phage
Petri plate
with T1 phage
Incubate overnight to
allow bacterial growth.
23
Mutation Rate

The term mutation rate is the likelihood that a gene
will be altered by a new mutation



The mutation rate for a given gene is not constant


It is commonly expressed as the number of new mutations
in a given gene per cell generation
It is in the range of 10-5 to 10-9 per generation
It can be increased by the presence of mutagens
Mutation rates vary substantially between species
and even within different strains of the same species
24
Mutation Rates and Frequencies

Within the same individual, some genes mutate at a
much higher rate than other genes

Some genes are larger than others


Some genes have locations within the chromosome that
make them more susceptible to mutation


This provides a greater chance for mutation
These are termed hot spots
Note: Hot spots can be also found within a single gene

Specific bases or regions that are more likely to be the site of a
mutation within a gene
25
Causes of
Spontaneous Mutations

Spontaneous mutations can arise by three types of
chemical changes

1. Depurination

2. Deamination

3. Tautomeric shift
The most common; We will focus here
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26
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
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27
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5′
3′
5′
C G
A T
T A
C G
G C
C G
A T
T A
C
G C
Depurination
5′
3′
3′
Apurinic site
3′
5′
5′
3′
(a) Depurination
3′
5′
C G
A T
T A
C
G C
3′
DNA replication
3′
5′
5
3′
C G
A T
T A
X
G C
5′
(b) Replication over an apurinic site
Figure 16.8
Three out of four (A, T and G)
are the incorrect nucleotide
C G
A T
T A
C G
G C
3′
Spontaneous depurination
There’s a 75% chance
of a mutation
X could be
A, T, G, or C
5′
28
Mutations Due to Trinucleotide
Repeats

Several human genetic diseases are caused by an
unusual form of mutation called trinucleotide repeat
expansion (TNRE)


These diseases include



The term refers to the phenomenon that a sequence of 3
nucleotides can increase from one generation to the next
Huntington disease (HD)
Fragile X syndrome (FRAXA)
Refer to Table 16.5 for these and other examples
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29
30

Certain regions of the chromosome contain
trinucleotide sequences repeated in tandem


In normal individuals, these sequences are transmitted from
parent to offspring without mutation
However, in persons with TNRE disorders, the length of a
trinucleotide repeat has increased above a certain critical size



Disease symptoms occur
In some diseases, it also becomes prone to expansion
This phenomenon is shown here with the trinucleotide repeat CAG
CAGCAGCAGCAGCAGCAGCAGCAGCAGCAGCAG
n = 11
CAGCAGCAGCAGCAGCAGCAGCAGCAGCAGCAGCAGCAGCAGCAGCAGCAGCAG
n = 18
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
In some cases, the expansion is within the coding
sequence of the gene


Typically the trinucleotide expansion is CAG (glutamine)
Therefore, the encoded protein will contain long tracks of
glutamine



This causes the proteins to aggregate with each other
This aggregation is correlated with the progression of the disease
In other cases, the expansions are located in
noncoding regions of genes


Some of these expansions are hypothesized to cause
abnormal changes in RNA structure
Some produce methylated CpG islands which may silence
the gene
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32

There are two particularly unusual features that
some TNRE disorders have in common

1. The severity of the disease tends to worsen in future
generations


This phenomenon is called anticipation
2. Anticipation usually depends on whether the disease is
inherited from the father or mother



In Huntington disease, the TNRE is more likely to occur if inherited
from the father
In myotonic muscular dystrophy, the TNRE is more likely to occur if
inherited from the mother
This suggests that TNRE can occur more frequently during
oogenesis or spermatogenesis, depending on the gene involved.
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33

The “DNA” cause of TNRE is not fully understood

TNREs contain at least one C and one G


This allows formation of a hairpin
During DNA replication, a hairpin can lead to an increase
or decrease in the length of the DNA



Polymerase can slip off DNA
Hairpin forms and pulls strand back
DNA polymerase hops back on



See Figure 16.12 for details
These changes can occur during gamete formation


Begins synthesis from new location
offspring will have very different numbers of repeats
Can also increase repeats in somatic cells

This can increase severity of the disease with age
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34
Mechanisms of trinucleotide repeat expansion or deletion
One DNA template strand prior to DNA replication
One DNA template strand prior to DNA replication
TNRE
TNRE
DNA replication begins
and goes just past the TNRE.
Hairpin forms in template strand
prior to DNA replication.
DNA
polymerase
DNA polymerase slips off
the template strand and a
hairpin forms.
DNA replication occurs and
DNA polymerase slips over
the hairpin.
DNA polymerase resumes
DNA replication.
DNA repair occurs.
DNA repair occurs.
TNRE is longer.
TNRE is shorter.
OR
TNRE is the same length.
(b) Mechanism of trinucleotide repeat expansion
OR
TNRE is the same length.
(c) Mechanism of trinucleotide repeat deletion
Figure 16.12b and c
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35
Types of Mutagens


An enormous array of agents can act as mutagens
that 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 16.6
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37
Mutagens Alter DNA Structure in
Different Ways

Chemical mutagens come into three main types

1. Base modifiers

2. Intercalating agents

3. Base analogues
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38

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 16.15

Some chemical mutagens disrupt the appropriate pairing
between nucleotides by alkylating bases within the DNA

Examples: Nitrogen mustards and ethyl methanesulfonate (EMS)
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39
Template strand
H
After replication
H
NH2
O
N
H
N
N
H
HNO2
N
N
N
Sugar
O
Sugar
H
Cytosine
N
N
H
Sugar
N
O
H
Uracil
These mispairings
create mutations in the
newly replicated strand
Adenine
H
N
H
N
H
NH2
O
H
H
N
HNO2
N
N
Sugar
N
Sugar
N
N H
H
N
N
H
Adenine
N
H
Hypoxanthine
O
Sugar
Cytosine
Figure 16.15 Mispairing of modified bases
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40

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 resulting in frameshifts

Examples:


Acridine dyes
Proflavin
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41

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
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H
O
Br
N
H
H
N
O
5-bromouracil
(keto form)
O
N
N
Sugar
Sugar
H
H
H
N
O
O
This tautomeric shift
occurs at a relatively
high rate
H
H
N
N
N
5-bromouracil
(enol form)
Adenine
Normal pairing
Figure 16.16
Br
H
N
N
N
Sugar
N
N
Sugar
N
H
Guanine
Mispairing
(a) Base pairing of 5BU with adenine or guanine
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42
In this way, 5-bromouracil can promote a change
of an AT base pair into a GC base pair
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5′
5′
3′
A 5BU
3′
3′
A T
DNA
replication
3′
5′
5′
5′
5′
3′
G 5BU
3′
5′
3′
G C
DNA
replication
3′
5′
5′
3′
G or A 5BU
3′
5′
(b) How 5BU causes a mutation in a base pair during DNA replication
Figure 16.16
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43

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
Oxidized bases
Single nicks in DNA strands
Cross-linking
Chromosomal breaks
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44
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H
O

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
O
P
O
CH2
O–
H
H
H
N
CH3
H
H
Thymine
CH3
O
O
P
O
CH2
O–
H
H
O
O
H
H
N
N
H
H
O
Thymine
H
Ultraviolet
light
O
O
P
O
O
H
O
CH2
O–
H
H
N
O
O
O
H
H
N
H
CH3
H
H
CH3
O
O
P
O
CH2
O–
Figure 16.17
N
O
H
H
H
O
O
H
H
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N
H
N
H
O
Thymine dimer
45
BIO 184, Exam 2, Spring 2012
35
30
25
20
15
10
5
0
A
B
C
D
F
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