Unit 12 DNA Damage and Repair

UNIT 12
DNA DAMAGE AND REPAIR

Structure
12.1 Introduction Base Excision Repair

Expected Learning Nucleotide Excision Repair

Outcomes
Mismatch Repair
12.2 Replication Error and DNA
Recombinant Repair
Damage
Translesion DNA Synthesis
Genomic DNA Damage
12.4 Summary
Mitochondrial DNA Damage
12.5 Terminal Questions
12.3 DNA Repair
12.6 Answers
Proof Reading
12.7 Further Readings
Direct Repair

12.1 INTRODUCTION
In the previous unit we discussed about mutations, mutagenesis and how it
adds to the diversity of organisms and population. In this unit we shall discus
about DNA damage and repair. The DNA of genome is considered stable.
However, several endogenous factors generated through metabolism and
exogenous factors like radiation and environmental stressors have been
identified, which can induce changes in DNA.

An estimate suggests that human cells suffer with more than seventy
thousand DNA damages per day. The lesions may either be in the form of
single-strand break (SSB) or double-strand breaks (DSB) within the DNA.
Some changes occur during duplication of genome due to the errors in DNA
replication but most of them are induced later due to occupational hazards or
life style. The ultraviolet radiations, environmental chemicals, thermal
fluctuations, reactive metabolites or reactive oxygen species cause either 221
Block 4 Mutations and DNA Repair

deletion or substitution of one or more base pairs. The pyrimidine dimers are
formed as a result of exposure to UV light. Ionising radiations from radioactive
elements such as uranium, radium, plutonium and X-rays also damage DNA.
The superoxide and hydroxyl radicals damage DNA by either oxidation of
sugar and base or breaking of the strands.

Most of the changes that occur in DNA are instantly corrected by DNA repair
processes. Accuracy in DNA replication along with repairing accidental lesions
is essential for maintaining the genetic stability. The Mismatch repair fixes
mispaired bases just after DNA replication. The DNA damage repair pathways
detect and correct damage throughout the cell cycle. The excision repair
mechanism targets the removal of bulky DNA adducts, UV-induced
photoproducts, base-pair alterations, purine loss, DNA mismatches, single-
and double-strand DNA breaks. Several proteins and enzymes are required
during DNA damage response (DDR) and repair signalling pathways. We shall
deal about these aspects in this unit in detail.

Expected Learning Outcomes

After studying this unit, you should be able to:

 describe factors responsible for DNA damage and repair;

 draw and represent processes involved in Genomic and Mitochondrial

DNA damage; and

 explain mechanisms of different types of DNA repair.

12.2 REPLICATION ERROR AND DNA

DAMAGE
You studied about Replication in Block II of this course. You would have
certainly realized that the mechanisms involved in DNA replication have been
found extremely accurate. The process of proofreading also corrects error
during DNA replication. In addition, induced alterations are either spontaneous
or caused due to energy-rich radiations. However, some errors are
incorporated in DNA during its replication, Replication errors. The errors may
also occur during strand slippage that may induce insertions or deletions of
nucleotide bases. The shift in the position of nucleotides causes a wobble
between a normal thymine and normal guanine. The changes in sequences of
DNA get transferred into subsequent generation. If they are not repaired, the
accumulation of changes in DNA sequences turns lethal to cells, tissues or
organisms.

Loss of genomic integrity may lead to immunodeficiency, neurodegeneration

and cancer. The damage of DNA may be in the form of change in single
nucleotide by alkylation, de-amination, methylation or mismatch produced by
replication error. It may cause structural distortion due to nick in single strand
or double strand break or un-usual covalent bond formation like thymidine
dimers. The damages may be either in the genomic DNA or mitochondrial
DNA or in both of them. Let’s look at them one by one.
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Unit 12 DNA Damage and Repair

12.2.1 Genomic DNA Damage

Genomic DNA that stores the genetic information is highly susceptible to
chemicals and physical insults. It gets damaged by environmental and internal
hazards, such as irradiation, various types of chemicals, reactive oxygen
species and replication errors. These factors and stresses can generate
numerous lesions (Fig. 12.1) on base pairs and ribose sugar-phosphate
backbone of DNA. Among DNA lesions, DNA double-strand break (DSB) is
the most deleterious type of lesion. If it is not repaired, DSB can cause
genomic instability and several other genetic diseases. Loss of DNA damage
repair leads to the accumulation of DNA lesions and damage response.
Several proteins like cohesin and BRCA participate in DNA damage and
repair response. The cohesin, a multi protein complex, serves as critical
component during separation of sister chromatids and activation of DNA
damage check point. The Breast Cancer type 1 susceptibility protein, BRCA1
plays an important role in maintaining genomic stability. Mutations of BRCA1
impair DNA damage repair and also cause familial breast and ovarian tumors.

Reactive oxygen
species (ROS) are
oxygen-containing
radicals that are
capable of
independent
existence with one or
more unpaired
electrons. The term
Fig. 12.1: Factors cause either single or double-strand break in Genomic DNA.
Replication error, X-ray, UV light, alkylating agents, spontaneous reactions and ROS is most often
reactive oxygen species cause DNA break. expanded to include
reactive oxygen-
12.2.2 Mitochondrial DNA Damage containing
compounds without
While studying extranuclear inheritance you learnt about the mitochondrial unpaired electrons,
DNA, Just to recall, mitochondria also contain their own DNA called such as hydrogen
mitochondrial DNA (mtDNA) which is about one percent of cellular DNA. The peroxide (H2O2) and
1
singlet oxygen ( O2)
mtDNA is circular and double stranded. Here what is important to note is, it is
five to ten times more prone to mutations than genomic DNA. The damages in Buildup of ROS in
mtDNA have been found responsible for several diseases like stroke, cancer, cells may cause
damage to DNA,
diabetes and neuro degeneration. The mtDNA also gets damaged by
RNA, and proteins,
radiation, genotoxic chemicals and reactive oxygen species (ROS). The and may cause cell
impact of damage on mtDNA is comparatively more because of errors during death.
DNA replication and lack of conventional histone proteins in mitochondria. The
ROS can lead to various types of oxidative damage including base
modification or removal, DNA strand breaks and cross linking (Fig. 12.2). Also,
the mtDNA DNA polymerase gamma (pol γ) has low frame shift fidelity. 223
Block 4 Mutations and DNA Repair

Fig. 12.2: Representation of mitochondrial DNA damage due to toxicants.

SAQ 1
i) Define replication error.

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ii) What are the factors involved in DNA damage?

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iii) Why is mitochondrial DNA more susceptible to damage?

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12.3 DNA REPAIR

Now you are aware what could be the probable reasons for DNA damage. Let
us see how these damages can be corrected. The alterations in the
composition of DNA are corrected by specific proteins and DNA polymerases.
The process is step-wise highly subjective based on types of damages.

Repair mechanisms can be broadly grouped into three categories: direct

reversal of the damage, excision repair and post-replication repair mechanism.
However we shall consider each repair mechanism separately.

It could be either direct repair with the help of methyl transferases or by

photolyases using Ultraviolet rays. The DNA polymerases synthesize new
DNA strand. Both prokaryotic and eukaryotic cells contain multiple DNA
polymerases. The exonuclease activity of DNA polymerases excise bases
that have been added to DNA incorrectly. In prokaryotes, out of five DNA
polymerases DNA polymerase III serves as major replicase. However, DNA
polymerase-I, DNA polymerase-II, DNA polymerase-IV and DNA polymerase-
V participate in repair of DNA damage (Table 12.1).
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Unit 12 DNA Damage and Repair

Table 12.1: DNA polymeraseswith their associated functions

Enzymes Functions

DNA polymerase I Major repair enzyme

DNA polymerase II Minor repair enzyme

DNA polymerase III DNA replication

DNA polymerase IV SOS repair

DNA polymerase V SOS repair

Under certain circumstances like ultraviolet irradiation the error prone DNA
synthesis occurs in E. Coli by DNA polymerase IV and DNA polymerase V.
Since DNA polymerase IV and DNA polymerase V are devoid of proofreading
activities they mostly induce replication error.

Under cellular stress, when there are chances of multiple DNA damages,
extensive and multi-factorial repair system including recombination mediated
repair and error prone repair participate as SOS repair system. In eukaryotic
cells, several polymerases like DNA polymerase β, DNA polymerase δ , DNA
polymerase ε and DNA polymerase κ have been observed responsible for
repairing damaged DNA and summarized in Table 12.2.

Table 12.2: EukaryoticDNA polymerases and their associated functions

Enzymes Functions

DNA polymerase α Nuclear replication

DNA polymerase δ Nuclear replication

DNA polymerase ε Nuclear replication

DNA polymerase γ Mitochondrial replication

DNA polymerase β Base excision repair

DNA polymerase δ Thymine dimer bypass

DNA polymerase ε Base damage repair

DNA polymerase ι Required in meiosis

DNA polymerase κ Deletion and base substitution

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 Block 4 Mutations and DNA Repair

Let us consider these repair mechanisms in detail.

12.3.1 Proofreading
The processivity of DNA polymerase slows and exonuclease property gets
activated if any wrong or incorrectly paired nucleotide gets added. During DNA
replication, the DNA polymerases remove incorrect nucleotide added during
replication. The process is described as proofreading activity of DNA
polymerase. The incorrect nucleotide is removed and replaced with the
correct nucleotide before continuing with DNA synthesis.

12.3.2 Direct Repair

Direct repair is rare and involves the reversal or simple removal of the
damage. There are some lesions in DNA that can be repaired by direct
reversal of the damage, without the removal of any base or a nucleotide. For
example, Photo-reactivation of pyrimidine-dimers, in which dimers are
reversed by a light-dependent enzyme, DNA photolyase. This enzyme is
present in almost all cells from bacteria to animals (Fig.12.3).

UV-induced thymine
dimers are repaired
by photoreactivation
enzyme. The energy
from visible light splits
bonds and forms
cyclobutane ring. The
enzyme gets released
to restore native DNA
having a normal state
of thymine.

Fig. 12.3: Diagrammatic representation of direct repair mechanism using

226 photolyase enzyme.
Unit 12 DNA Damage and Repair

Exposure to UV light causes damage to DNA by formation of pyrimidine dimer

in which adjacent pyrimidines on the same strand of DNA are joined by the
formation of a cyclobutane ring. They distort the structure of the DNA chain
and block transcription or replication beyond the site of damage. These UV-
induced pyrimidine dimers are repaired by direct reversal of the dimerization
reaction. The energy derived from visible light is used to break the cyclobutane
ring and restore the pyrimidine bases in their normal state. This process is
called photo-reactivation. The repair of pyrimidine dimers by photo-
reactivation is very common in cells of prokaryotes and eukaryotes including
plants and animals. Direct repair mechanism is not universal to humans.

SAQ 2
i) List any three enzyme involved in DNA repair in prokaryotes.

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ii) What is the importance of proofreading on replication of DNA?

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iii) What is meant by SOS repair?

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iv) Define photo-reactivation.

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12.3.3 Base Excision Repair

The lesions produced by mutagenic agents are recognized by a group of
enzymes called DNA glycosylases. The enzyme detects and removes a
specific kind of damaged base by cleaving N-glycosyl bond and plays a key
role in base excision repair. For example, de-amination converts a cytosine
base into uracil, a base typically found only in RNA. During DNA replication,
uracil pairs with adenine because of uncorrected cytosine-to-uracil change.
This process induces a mutation. To prevent such mutations, a glycosylase
from the base excision repair pathway detects and removes de-aminated
cytosine. Once the base has been removed, the "empty" piece of DNA
backbone is also removed, and the gap is filled and sealed by enzyme DNA
ligase (Fig. 12.4).

The DNA glycosylases also recognize and remove other abnormal bases,
including hypoxanthine formed by the de-amination of adenine, pyrimidine 227
Block 4 Mutations and DNA Repair

dimers, alkylated purines other than O6-alkylguanine, and bases damaged by

oxidation or ionizing radiation.

Fig. 12.4: Diagrammatic representation of base excision repair after DNA

damage. De-amination of cytosine (C) leads to formation of uracil (U).

DNA glycosylase cleaves the bond between uracil and the deoxyribose,
leaving a sugar without base (an AP site). AP endonuclease recognizes this
site and cleaves the DNA chain. The remaining deoxyribose is removed by
deoxyribose phosphodiesterase. The cytosine gets incorporated instead of
uracil during activity of DNA polymerase.
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Unit 12 DNA Damage and Repair

12.3.4 Nucleotide Excision Repair

Nucleotide excision repair is another pathway to remove and replace damaged
nucleotides. It is the major repair system which operates on the bulky structure
and large distortion of DNA. It is different from base-excision repair
because DNA glycolyase in base excision repair only recognizes specific
forms of damaged bases whereas nucleotide excision repair detects
DNA damage that distorts the DNA molecule. It gets activated when huge
deformation is created in the helical structure of DNA by the DNA lesions. It
detects bases that have been modified with bulky chemical groups and
corrects types of damage that distort the DNA double helix, such as DNA
methylation induced due to chemicals in cigarette smokers.

MGMT (O6-
Methylguanine-DNA
methyltransferase)
enzyme transfers a
methyl group from the O6
position of guanine to the
active-site cysteine
residue of the MGMT
polypeptide chain. This
restores the normal base
and forms the stable S-
methylcysteine adduct,
which inactivates the
enzyme. This is why the
reaction catalyzed is said
to be "suicide
mechanism” by MGMT.

Fig.12.5: Diagrammatic representation of nucleotide excision repair after DNA

damage.
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Block 4 Mutations and DNA Repair

Nucleotide excision repair system also fixes damages caused by UV radiation.

The UV radiation leads to formation of the most common type of linkage, a
thymine dimer. It consists of two thymine bases that react with each other and
become chemically linked. In nucleotide excision repair, the damaged
nucleotides are removed along with a surrounding patch of DNA. It replaces
the missing DNA and seal the gap in the backbone of the strand (Fig.12.5).

The thymine dimer causing DNA damage is recognised and cleaved at both
sides by nucleases. The helicase unwind the DNA, DNA polymerases
synthesize nucleotides and the gap is sealed by ligase.

In E. coli, the products of three genes (UvrA, B and C) catalyze nucleotide-

excision repair where protein UvrA recognizes damaged DNA and recruits
UvrB and UvrC to the site of the lesion. The protein UvrB and UvrC remove
the damaged site by excising an oligonucleotide consisting of 12 or 13 bases.
The helicase removes the damaged oligonucleotides from the double-stranded
DNA. The DNA polymerase-I adds corresponding nucleotides and gaps are
sealed by DNA ligase. In eukaryotes, XP protein (subunit of TFIIH) functions
like Uvr system of prokaryotes.

Individuals suffer with inherited diseases namely, xerodermapigmentosum

(XP), Cockayne's syndrome and trichothiodystrophydue to deficiencies in the
ability to repair DNA damage.

12.3.5 Mismatch Repair

During DNA replication sometimes mismatched bases get incorporated.

Generally, proofreading activity of DNA polymerase removes such
mismatched bases. If escaped by proofreading, mismatch repair system
remove and replace mispaired bases. In mismatch repair mechanism, a
protein complex (MutS, MutL, and MutH) recognizes and binds to the
mispaired base. A second complex composed of Helicases and Exonucleases
cuts the DNA near the mismatch, removes incorrect nucleotide and a
surrounding patch of DNA. A DNA polymerase then replaces the missing
section with correct nucleotides, and an enzyme called a DNA ligase seals the
gap (Fig.12.6).

The eukaryotic homologs of MutS and MutL bind to the mismatched bases
and correct them like E.coli. In mammalian cells, presence of single strand
breaks in the newly replicated DNA provides strand-specificity of mismatch
repair. Mutations in homologs of MutS, and MutL are responsible for inherited
colon cancer.

MutS binds to the mismatched base, followed by MutL which activates MutH.
The MutH then cleaves the unmodified strand. MutS, MutL, helicase and an
exonuclease remove part of the unmodified strand that contains the mismatch.
The DNA polymerase and ligase complete the process of repair.
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Unit 12 DNA Damage and Repair

Fig. 12.6: Diagrammatic representation of mismatch repair mechanism after

DNA damage.

12.3.6 Recombination Repair

Some types of environmental factors or high-energy radiation causes double-
stranded breaks in DNA that may lead to loss of large segments of
chromosomes and the hundreds of genes (Fig.12.7). In principle, a
recombination event occurs on either side of the damaged site. It allows
undamaged single strand to pair with the damaged strand which facilitated
reconstruction and continuation of the replication fork bypassing the damaged
site. The recombination-repair pathways help to restore the fork after
completion of the damage has been repaired. This is post replication repair
system. The pathway for handling a stalled replication fork requires RecA and
the RecBC system in E. coli. In one possible pathway, RecA binds to single-
stranded DNA at the stalled replication fork, stabilizes it, and possibly acts as
the sensor that detects the stalling event. The RecBC is involved in excision- 231
Block 4 Mutations and DNA Repair

repair of the damage. After the damage has been repaired, replication can
resume. Stalled replication forks can be rescued by recombination-repair.

Another pathway i.e. Non-homologous end joining (NHEJ) is a repair

mechanism which does not involve homologous recombination. It facilitates
joining of two ends of the broken DNA double strand using a heterodimeric
enzyme complex consisting of proteins Ku. The Ku, a member of highly
conserved family protein found in bacteria, yeast and humans.

Fig. 12.7: Diagrammatic representation of recombination repair mechanism of

Double stranded break in DNA.
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Unit 12 DNA Damage and Repair

Replication forks collapse when the advancing fork encounter nick on the
template strand. The broken arm is processed by RecBCD to create 3’
overhang. The 3’ overhang invades the donor DNA and forms a D-loop.
Invasion is followed by formation of Holliday junction. Resolution of holiday
junction restores the replication fork.

12.3.7 Translesion DNA Synthesis

Translesion DNA synthesis (TLS) is the process by which cells replicate DNA
containing unrepaired damage that prevents progression of the replication
fork. Translesion synthesis can bypass a replication block caused by various
DNA damage. It replicates DNA over such lesions but the enzymes are error
prone. After that these lesions are repaired by the excision repair process.
Translesion synthesis is catalysed by a specialised class of DNA polymerases
that synthesize DNA directly across the site of the damage. When any lesion
is found in the template during replication, DNA polymerase III with its sliding
clamp dissociates from the DNA. It is replaced by the translesion DNA
polymerase, which extends DNA synthesis across such lesions like the
thymine dimer on the template strand (Fig. 12.8). Once, it has replicated the
lesion, translesion polymerase is then replaced by the DNA polymerase III.
The translesion polymerase is a complex of the proteins UmuC and UMuD. It
belongs to a distinct family of DNA polymerases found in many organisms
known as the Y-family of DNA polymerases. These polymerases are template
dependent. They incorporate nucleotides in a manner that is independent of
base pairing.

Fig.12.8: Diagrammatic representation of Translesion DNA synthesis. 233

Block 4 Mutations and DNA Repair

DNA polymerase III dissociates from the DNA and is replaced by the
translesion DNA polymerase if encounters a lesion in the template during
replication. The translesion DNA polymerase extends DNA synthesis across
the thymine dimer on the template strand.

SAQ 3
Choose the correct option:

a) Which enzyme is responsible for photo-reactivation of DNA:

i) Photoligase

ii) Photoreductase

iii) Photo-oxidase

iv) Photolyase

b) The process involved in replication of damaged DNA is

i) DNA replication

ii) Truncated DNA synthesis

iii) Translesion DNA synthesis

iv) Mitochondrial DNA synthesis

c) Translesion DNA polymerase is composed of

i) DNA pol I, II and III

ii) UmuC and UmuD protein complex

iii) MutS, MutH and MutL protein complex

iv) Nucleases

d) Which of the enzyme(s) is involved in base excision repair?

i) DNA glycosylase

ii) AP endonuclease

iii) AP exonuclease

iv) DNA polymerase

e) Which enzymes remove damaged/mismatched DNA ahead of the

replication fork?

i) Helicases

ii) DNA polymerases

iii) Primases

iv) Topoisomerases
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Unit 12 DNA Damage and Repair

f) The enzyme complex MutS, MutL and MutH is involved in

i) Direct repair mechanism

ii) DNA double strand repair mechanism

iii) Mismatch repair mechanism

iv) Nucleotide excision repair mechanism

12.4 SUMMARY
 DNA repair can be described as any of the several mechanisms by
which a cell maintains the integrity of its genetic code. DNA repair
ensures the survival of a species by enabling parental DNA to be
inherited as faithfully as possible to theoffspring’s. It also preserves the
health of an individual.

 Mutations in the genetic code can lead to cancer and other genetic
disease. Spontaneous mutations occur when DNA bases react with their
environment and changes its structure, causing it to pair with an
incorrect base. Replication errors are minimized when the DNA
replication machinery “proofreads” its own synthesis, but sometimes
mismatched base pairs escape proofreading.

 Chemical agents modify bases and interfere with DNA replication. There
are three type of repair mechanism; direct reversal of the damage,
excision repair and post-replication repair.

 Direct repair reversal is specific to the damage. For direct repair reversal
of alkylation events, a DNA methyltransferases or DNA glycosylase
detects and removes the alkyl group. Excision repair can be specific or
nonspecific.

 In base excision repair DNA glycosylase specifically identify and remove

the mismatched base. In nucleotide excision repair, the repair machinery
recognizes a wide array of distortions in the double helix caused by
mismatched bases; in the form of repair, the entire distorted region is
excised. Post-replication repair occurs downstream of the lesion,
because replication is blocked at the actual site of damage. Error-prone
repair tends to be inaccurate and subject to mutation.

 Hereditary nonpolyposis colorectal cancer results from a mutation in

MSH2 and MLH1 proteins, which repair mismatches during replication.
Xerodermapigmentosum (XP) is another condition that results from
failed DNA repair.

 Therefore, an effective DNA repair mechanism is very necessary for the

survival of the organism as well as for the identity and integrity of its
hereditary molecules.
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Block 4 Mutations and DNA Repair

12.5 TERMINAL QUESTIONS

1. How does damage of Genomic and mitochondrial DNA occur?

2. What is DNA repair?

3. Describe types of DNA polymerase involved in DNA repair.

4. Why direct repair mechanism is important.

5. Differentiate between exonuclease and polymerase.

6. How do DNA lesions cause genomic instability?

7. In what condition mismatch repair is active?Illustrate the mechanism of

mismatch repair.

8. What is the role of MutS and MutH?

9. Explain the mechanism of base excision repair.List the enzymes

involved in base excision repair.

10. What are the factors responsible for nucleotide excision repair?Explain
nucleotide excision repair with the help of a suitable diagram.

11. Differentiate between Base Excision Repair and Nucleotide Excision

Repair.

12. Why is recombination repair important?How do RecA and RecBC

function?

13. Explain translesion DNA synthesis and its importance.

12.6 ANSWERS
Self-Assessment Questions
1. i) Some errors are incorporated in DNA during its replication and are
known as replication errors.The errors may also occur during
strand slippage that may induce insertions or deletions of
nucleotide bases.

ii) DNA damage may occur during duplication of genome due to the
errors in DNA replication but most of them are induced later due to
occupational hazards or life style. The ultraviolet radiations,
environmental chemicals, thermal fluctuations, reactive
metabolites or reactive oxygen species cause either deletion or
substitution of one or more base pairs. The pyrimidine dimers are
formed as a result of exposure to UV light. Ionising radiations from
radioactive elements such as uranium, radium, plutonium and X-
rays also damage DNA. The superoxide and hydroxyl radicals
damage DNA by either oxidation of sugar and base or breaking of
the strands. The damage of DNA may be in the form of change in
single nucleotide by alkylation, de-amination, methylation or
mismatch produced by replication error. It may cause structural
distortion due to nick in single strand or double strand break or un-
usual covalent bond formation like thymidine dimers.
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Unit 12 DNA Damage and Repair

iii) mtDNA is comparatively more susceptible to damage because of

errors during DNA replication and lack of conventional histone
proteins in mitochondria. The ROS can lead to various types of
oxidative damage including base modification or removal, DNA
strand breaks and cross linking. Also, the mtDNA DNA polymerase
gamma (pol γ) has low frame shift fidelity.

2. i) In prokaryotes, out of five DNA polymerases DNA polymerase III

serves as major replicase. However, DNA polymerase-I, DNA
polymerase-II, DNA polymerase-IV and DNA polymerase-V
participate in repair of DNA damage.

ii) Proofreading activity of DNA polymerases remove incorrect

nucleotide added during replication. The incorrect nucleotide is
removed and replaced with the correct nucleotide before
continuing with DNA synthesis.

iii) Under cellular stress, when there are chances of multiple DNA
damages, extensive and multi-factorial repair system including
recombination mediated repair and error prone repair participate
and is known asSOS repair system.

iv) Exposure to UV light causes damage to DNA by formation of

pyrimidine dimer in which adjacent pyrimidines on the same strand
of DNA are joined by the formation of a cyclobutane ring. These
UV-induced pyrimidine dimers are repaired by direct reversal of
the dimerization reaction. The energy derived from visible light is
used to break the cyclobutane ring and restore the pyrimidine
bases in their normal state. This process is called photo-
reactivation.

3. a) Photolyase

b) Translesion DNA synthesis

c) UmuC and UmuD protein complex

d) DNA glycosylase

e) Helicases

f) Mismatch repair mechanism

Terminal Questions
1. Several endogenous factors generated through metabolism and
exogenous factors like radiation and environmental stressors have been
identified, which can induce changes in DNA.The mtDNA also gets
damaged by radiation, genotoxic chemicals and reactive oxygen species
(ROS). Refer section 12.2

2. The alterations in the composition of DNA are corrected by specific

proteins and DNA polymerases. DNA repair process is step-wise, highly
subjective based on the types of damages.Repair mechanisms can be
broadly grouped into three categories: direct reversal of the damage,
excision repair and post-replication repair mechanism.
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Block 4 Mutations and DNA Repair

3. Refer table 12.1 and 12.2

4. Direct repair mechanism is important because it involves the reversal or

simple removal of the damage. There are some lesions in DNA that can
be repaired by direct reversal of the damage, without the removal of any
base or a nucleotide. For example, Photo-reactivation of pyrimidine-
dimers, in which dimers are reversed by a light-dependent enzyme, DNA
photolyase. This enzyme is present in almost all cells from bacteria to
animals. Refer section 12.3.2

5. The DNA polymerases synthesize new DNA strand. Both prokaryotic

and eukaryotic cells contain multiple DNA polymerases while the
exonuclease activity of DNA polymerases excise bases that have been
added to DNA incorrectly. Refer section 12.3.

6. Loss of DNA damage repair leads to the accumulation of DNA lesions

and damage response.Among DNA lesions, DNA double-strand break
(DSB) is the most deleterious type of lesion. If it is not repaired, DSB can
cause genomic instability and several other genetic diseases. Refer
section 12.2.1

7. During DNA replication sometimes mismatched bases get incorporated.

Generally, proofreading activity of DNA polymerase removes such
mismatched bases. If escaped by proofreading, mismatch repair system
becomes active and remove or replace mispaired bases. Refer section
12.3.5

8. MutS, and MutH protein complex recognizes and binds to the mispaired
base in mismatch repair mechanism.

9. In base excision repair mechanism the lesions produced by mutagenic

agents are recognized by a group of enzymes called DNA glycosylases.
The enzyme detects and removes a specific kind of damaged base by
cleaving N-glycosyl bond and plays a key role. Refer section 12.3.3

10. Nucleotide excision repair gets activated when huge deformation is

created in the helical structure of DNA by the DNA lesions. It detects
bases that have been modified with bulky chemical groups and corrects
types of damage that distort the DNA double helix, such as DNA
methylation induced due to chemicals in cigarette smokers. Refer
section 12.3.4

11. Refer section 12.3.3 and 12.3.4

12. Some types of environmental factors or high-energy radiation causes

double-stranded break in DNA that may lead to loss of large segments of
chromosomes and the hundreds of genes. This is post replication repair
system. The recombination-repair pathways help to restore the
replication fork after completion of the damage has been repaired. RecA
binds to single-stranded DNA at the stalled replication fork, stabilizes it,
and possibly acts as the sensor that detects the stalling event. The
RecBC is involved in excision-repair of the damage. After the damage
has been repaired, replication can resume. Stalled replication forks can
be rescued by recombination-repair. Refer section 12.3.6
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Unit 12 DNA Damage and Repair

13. Translesion DNA synthesis (TLS) is the process by which cells replicate
DNA containing unrepaired damage that prevents progression of the
replication fork. Translesion synthesis can bypass a replication block
caused by various DNA damage. Refer section 12.3.7

12.7 FURTHER READINGS

1. Lodish H et al., Molecular Biology of the Cell, 5th edition (New York,
Freeman, 2004)

2. Lewin B, Genes VIII, Pearson International Edition (New York, 2004)

3. Albert B et al., The Cell, 6th edition (Garland Science, New York, 2017)

4. Yu X (2016), A special issue on the DNA damage response and genome

instability. Acta Biochim Biophys Sin, 48(7), 593.

5. Singh, G., Pachouri, U. C., Khaidem, D. C., Kundu, A., Chopra, C., &
Singh, P. (2015). Mitochondrial DNA Damage and Diseases.
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