mol biol lec 7 dna mutation and repair atea 091204

7/28/2019 Mol Biol Lec 7 DNA Mutation and Repair Atea 091204

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DNA Mutation and Repair


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DNA

Primary function permanent storage ofinformation

Does not normally change

Mutations do occur


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Mutation Heritable change in the genetic

material

Permanent structural change of

DNA

Alteration can be passed on to

daughter cells (somatic cell)

Mutations in reproductive cells

can be passed to offspring (germ

line)


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SOMATIC VS. GERM-LINE

The timing of mutations in multicellular

organisms plays an important role

Mutations may occur in gametes or a fertilized egg

Mutations may occur later in life

Embryonic or adult stages

Timing can affect

The severity of the genetic effect

The ability to be passed from parent to offspring


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Animals possess germ-line and somatic cells

Germ-line cells

Cells giving rise to gametes

Somatic cells

All cells of the body

excluding the germ-line cells

e.g., Muscle cells, nerve cells,

etc.


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Germ-line cells

Germ-line mutations can occur in

gametes Germ-line mutations can occur in a

precursor cell that produces

gametes

All cells in the resulting offspring

will contain the mutation


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Somatic cells

Somatic mutations in embryonic

cells can result in patches of tissues

containing the mutation

Size of the patch depends on the

timing of the mutation

Individual is a genetic mosaic


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Mutations

Provide allelic variation

Ultimate source of genetic variation

Foundation for evolutionary change

Various phenotypic effects

Neutral

Harmful

Beneficial


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TYPES OF MUTATIONS

Types of mutations

Chromosome mutations

Changes in chromosome structure

Genome mutations Changes in chromosome number

Single-gene mutations

Relatively small changes in DNA

structure Occur within a particular gene

Focus of study in this chapter


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TYPES OF MUTATIONS

Mutations involve the permanent alteration of

a DNA sequence

Alteration of base sequence

Removal or addition of one or more nucleotides


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MUTATIONS

Point mutations

Change in a single base pair within the DNA

Two main types of point mutations

Base substitutions

Transition

Transversion

Small deletions or insertions


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MUTATIONS

Two types ofbase substitutions

Transition

Pyrimidine changed to another pyrimidine

e.g., C

T Purine changed to another purine

e.g., A G

Transversion

Purines and pyrimidines areinterchanged

e.g., A C

More rare than transitions


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EFFECTS OF MUTATIONS

What can happen when a mutation occurs in the DNA

(e.g. sickle cell anemia)

Figure 1. Concept of a mutation in the protein-coding region of a gene. (Note that not

all mutations lead to altered proteins and that not all mutations are in protein-coding

regions.)


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Silent mutations Amino acid sequence is not altered

e.g., CCC CCG (pro pro)

Genetic code is degenerate

Alterations of the third base of a codonoften do not alter the encoded amino

acid

Phenotype is not affected


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Missense mutations

Amino acid sequence is altered

e.g., GAAGTA (glu val)

Phenotype may be affected


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Neutral mutations Type of missense mutation

Amino acid sequence is altered

e.g., CTTATT (leu ile)

e.g., GAAGAC (glu asp)

No detectable effect on protein

function

Missense mutations substitutingan amino acid with a similar

chemistry to the original is likely

to be neutral


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Nonsense mutations

Normal codon is changed into a stop

codon

e.g., AAA AAG (lys stop)

Translation is prematurely terminated

Truncated polypeptide is formed

Protein function is generally affected


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A nonsense mutation and its effect on translation


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Mutations occasionally produce a polypeptidewith an enhanced ability to function

Relatively rare

May result in an organismwith a greater likelihoodto survive and reproduce

Natural selection may

increase the frequency ofthis mutation in thepopulation


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MUTATION TYPES

Genetic terms to describe mutations

Wild-type

Relatively common genotype

Generally the most common allele

Variant

Mutant allele altering an organisms phenotype

Forward mutation

Changes wild-type allele into something else

Reverse mutation

Reversion

Restores wild-type allele


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MUTATION TYPES

Genetic terms to describe mutations Deleterious mutation

Decreases an organisms chance ofsurvival

Lethal mutation Results in the death of an organism

Extreme example of a deleteriousmutation

Conditional mutants Affect the phenotype only under a

defined set of conditions

e.g., Temperature-sensitive (ts) mutants


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MUTATION TYPES

Genetic terms to describe mutations

Suppressor mutation

Second mutation that restores the wild-type phenotype

Intragenic suppressor Secondary mutation in the same gene as

the first mutation

Differs from a reversion

Second mutation is at a different site

than the first Intergenic suppressor

Secondary mutation in a different genethan the first mutation


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MUTATION TYPES

Intergenic suppressors involving a mutantstructural gene Usually involve altered expression of one gene that

compensates for a loss-of-function mutation affecting

another gene Second gene may take over the functional role of the first

May involve proteins participating in a common cellularfunction

Sometimes involve mutations in genetic regulatory

proteins e.g., Transcription factors activating other genes that can

compensate for the mutation in the first gene


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TRINUCLEOTIDE REPEATS

DNA trinucleotide repeats

Three nucleotide sequences repeated in tandem

e.g., CAGCAGCAGCAGCAGCAG

Generally transmitted normally from parent tooffspring without mutation


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Trinucleotide repeat expansion (TNRE)

Number of repeats can readily increase from one

generation to the next

Cause of several human genetic

diseases

Length of a repeat has increased

above a certain critical size

Becomes prone to frequent

expansion


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TNRE disorders

Fragile X syndrome (FRAXA)

FRAXE mental retardation

Myotonic muscular dystrophy (DM)

Spinal and bulbar muscular atrophy (SBMA)

Huntington disease (HD)

Spinocerebellar ataxia (SCA1)


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TNRE disorders

Expansion may be within a coding sequence of a

gene

Most expansions are of a CAG repeat

Encoded proteins possess long tracts of glutamine

CAG encodes a glutamine codon

Presence of glutamine tracts causes aggregation of the

proteins

Aggregation is correlated with the progression of the

disease


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TNRE disorders

Expansion may be in a noncoding region of a gene

Two fragile X syndromes

Repeat produces CpG islands that become methylated

Methylation can lead to chromosome compaction

Can silence gene transcription

Myotonic muscular dystrophy

Expansions may cause abnormal changes in RNA structure


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TNRE disorders

Severity of the disease tends to worsen in future

generations

Anticipation

Severity of the disease depends on the parent

from whom it was inherited

e.g., In Huntingdon disease, TNRE likely to occur if

mutation gene is inherited from the father

e.g., In myotonic muscular dystrophy, TNRE likely to

occur if mutation gene is inherited from the mother


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TNRE disorders


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TNRE disorders

Cause of TNRE is not well understood

Trinucleotide repeat may produce alterations in

DNA structure

e.g., Stem-loop formation

May lead to errors in DNA replication

TNRE within certain genes alters gene expression Disease symptoms are produced


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CHROMOSOME STRUCTURE

Altered chromosome structure can alter gene

expression

Inversions and translocations commonly have no

obvious phenotypic effects

Phenotypic effects sometimes occur

Position effect


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CHROMOSOME STRUCTURE

Altered chromosome structure can alter geneexpression and phenotype

Breakpoint may occur within a gene

Expression of the gene is altered

Breakpoint may occur near a gene

Expression is altered when moved to a new location

May be moved next to regulatory elements influencing theexpression of the relocated gene

i.e., Silencers or enhancers

May reposition a gene from a euchromatic region to a highlycondensed (heterochromatic) region

Expression may be turned off


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CAUSES OF MUTATIONS

Two causes of mutations

Spontaneous mutations

Result from abnormalities in biological

processes Underlying cause lies within the cell

Induced mutations

Caused by environmental agents

Cause originates outside of the cell


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Spontaneous

Chemical Changes

Figure 7.9 (a) Deamination of cytosine to uracil. (b) Deamination of 5-methylcytosine

(5mC) to thymine.


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CAUSES OF MUTATIONS

Induced mutations are caused by mutagens

Chemical substances or physical agents originating

outside of the cell

Enter the cell and then alter the DNA structure


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CAUSES OF MUTATIONS

Spontaneous mutations are random events

Not purposeful

Mutations occur as a matter of chance

Some individuals possess beneficial mutations

Better adapted to their environment

Increased chance of surviving and reproducing

Natural selection results in differential reproductive

success The frequency of such alleles increases in the population


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Spontaneous mutations: Depurination

Most common type of naturally occurring

chemical change Reaction with water removes a purine (A or G)

from the DNA

Apurinic site


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Spontaneous mutations: Depurination

~10,000 purines lost per 20 hours at 37oC in atypical mammalian cell

Rate of loss increased by agents causing certain basemodification

e.g., Attachment of alkyl(methyl, ethyl, etc.) groups

Generally recognized by

DNA repair enzymes Mutation may result if

repair system fails


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Spontaneous mutations: Deamination of

cytosines

Methylation of cytosine occurs in many eukaryotic

species as well as prokaryotes

Removal of an amino group from the 5-methyl

cytosine produces thymine DNA repair enzymes cannot determine which is

the incorrect base

Hot spots for mutations are produced


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Spontaneous mutations: Tautomeric shifts

Common, stable form of T and G is the keto form

Interconvert to an enol form at a low rate

Common, stable form of A and C is the amino

form

Interconvert to an imino form at a low rate


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Spontaneous mutations: Tautomeric shifts

Enol and imino forms do not conform to normal

base-pairing rules

AC and GT base pairs are formed


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Certain non-mutagenic chemicals can be

altered to a mutagenically active form after

ingestion

Cellular enzymes such as oxidases can activatesome mutagens

Certain foods contain chemicals acting as

antioxidants Antioxidants may be able to counteract the effects

of mutagens and lower cancer rates


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CAUSES OF MUTATIONS

Mutagens alter DNA structure in various ways

Nitrous acid (HNO3) replaces amino groups with

keto groups

-NH2 =O

Can change cytosine

to uracil

Pairs with A, not G

Can change adenineto hypoxanthine

Pairs with C, not T


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Alkylating agents covalently attach methyl or ethyl

groups to bases

e.g., Nitrogen mustards, ethylmethanesulfonate (EMS)

Appropriate base pairing is

disrupted


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CAUSES OF MUTATIONS


Some mutagens directly interfere with the DNA

replication process

e.g., Acridine dyes such as proflavin Flat, planar structures interchelate into the double helix

Sandwich between adjacent base pairs

Helical structure is distorted

Single-nucleotide additions and deletions can result


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CAUSES OF MUTATIONS


Some mutagens are base analogs

e.g., 2-aminopurine

e.g., 5-bromouracil (5BU)

Become incorporated into

daughter strands during

DNA replication


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CAUSES OF MUTATIONS

Mutagens alter DNA structure in

various ways

Some mutagens are base analogs

5-bromouracil (5BU) is a thymineanalog

Incorporated in place of thymine

5BU can base-pair with adenine

Can tautomerize and base-pair withguanine at a relatively high rate

AT A5BU G5BU GC

Transition mutations occur


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CAUSES OF MUTATIONS


DNA molecules are sensitive to physical agents

such as radiation

e.g., Ionizing radiation such as X rays and gamma rays Short wavelength and high energy

Can penetrate deeply into biological materials

Creates free radicals

Chemically reactive molecules Free radicals alter DNA structure in a variety of ways

Deletions, single nicks, cross-linking, chromosomal breaks


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CAUSES OF MUTATIONS

Mutagens alter DNA structure in various ways DNA molecules are sensitive to physical agents such as

radiation

e.g., Nonionizing radiation such as

UV light Contains less energy

Penetrates only the surface of materialsuch as the skin

Causes the formation of thymine dimers

May be repaired through one of numerousrepair systems

May cause a mutation when that DNAstrand is replicated


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Effect of UV light on base DNA


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CAUSES OF MUTATIONS

Many different kinds of testes can determine

if an agent is mutagenic

Ames test is commonly used

Developed by Bruce Ames

Uses his- strains ofSalmonella typhimurium

Mutation is due to a point mutation rendering an

enzyme inactive

Reversions can restore his+ phenotype

Ames test monitors rate of reversion mutations


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CAUSES OF MUTATIONS

Ames test Suspected mutagen is mixed with rat liver extract

and his-Salmonella typhimurium

Rat liver extract provides cellular

enzymes that may be required toactivate a mutagen

Bacteria are plated on minimalmedia

his+ revertants can be detected

Mutation frequency calculated

Compared to control


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DNA REPAIR

Most mutations are deleterious DNA repair systems are vital to the survival

Bacteria possess several different DNA repairsystems

Absence of a single system greatlyincreases mutation rate

Mutator strains

Humans defective in a single DNA

repair system may manifest variousdisease symptoms

e.g., Higher risk of skin cancer


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DNA REPAIR

Living cells contain several DNA repair systems

Able to fix different types of DNA alterations


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mol biol lec 7 dna mutation and repair atea 091204

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