1. 1
1
!
! Since many mutations are deleterious, DNA repair systems
Since many mutations are deleterious, DNA repair systems
are vital to the survival of all organisms
are vital to the survival of all organisms
–
– Living cells contain several DNA repair systems that can fix different
Living cells contain several DNA repair systems that can fix different
type of DNA alterations
type of DNA alterations
!
! DNA repair mechanisms fall into 2 categories
DNA repair mechanisms fall into 2 categories
–
– Repair of damaged bases
Repair of damaged bases
–
– Repair of incorrectly
Repair of incorrectly basepaired
basepaired bases during replication
bases during replication
!
! In most cases, DNA repair is a multi-step process
In most cases, DNA repair is a multi-step process
–
– 1. An irregularity in DNA structure is detected
1. An irregularity in DNA structure is detected
–
– 2. The abnormal DNA is removed
2. The abnormal DNA is removed
–
– 3. Normal DNA is synthesized
3. Normal DNA is synthesized
DNA Repair
DNA Repair
2. 2
2
!
! Called DIRECT REPAIR
Called DIRECT REPAIR
!
! In a few cases, the covalent modifications of
In a few cases, the covalent modifications of
nucleotides can be reversed by specific enzymes
nucleotides can be reversed by specific enzymes
–
– Photolyase
Photolyase can repair thymine
can repair thymine dimers
dimers induced by UV light
induced by UV light
"
" It splits the
It splits the dimers
dimers restoring the DNA to its original condition
restoring the DNA to its original condition
–
– O
O6
6-alkylguanine
-alkylguanine alkyltransferase
alkyltransferase repairs
repairs alkylated
alkylated bases
bases
"
" It transfers the methyl or ethyl group from the base to a
It transfers the methyl or ethyl group from the base to a cysteine
cysteine
side chain within the
side chain within the alkyltransferase
alkyltransferase protein
protein
Damaged Bases Can Be Directly Repaired
Damaged Bases Can Be Directly Repaired
4. 4
4
!
! Base excision repair (BER)
Base excision repair (BER) involves a category of enzymes
involves a category of enzymes
known as
known as DNA-N-glycosylases
DNA-N-glycosylases
–
– These enzymes can recognize a
These enzymes can recognize a single damaged base
single damaged base and cleave
and cleave
the bond between it and the sugar in the DNA
the bond between it and the sugar in the DNA
–
– Removes
Removes one base, excises several around it,
one base, excises several around it, and
and replaces with
replaces with
several new bases
several new bases using
using Pol
Pol adding to 3
adding to 3’
’ ends then
ends then ligase
ligase attaching
attaching
to 5
to 5’
’ end
end
!
! Depending on the species, this repair system can eliminate
Depending on the species, this repair system can eliminate
abnormal bases such as
abnormal bases such as
–
– Uracil
Uracil; Thymine
; Thymine dimers
dimers
–
– 3-methyladenine; 7-methylguanine
3-methyladenine; 7-methylguanine
Base Excision Repair System
Base Excision Repair System
5. 5
5
Depending on whether a purine
or pyrimidine is removed, this
creates an apurinic and an
apyrimidinic site, respectively
Nick replication would be
a more accurate term
Base Excision
Base Excision
Repair System
Repair System
6. 6
6
!
! An important general process for DNA repair is
An important general process for DNA repair is nucleotide
nucleotide
excision repair (NER)
excision repair (NER)
–
– Nicks DNA around damaged base and removes region
Nicks DNA around damaged base and removes region
–
– Then fills in with
Then fills in with Pol
Pol on 3
on 3’
’ends, and attaches 5
ends, and attaches 5’
’ end with
end with ligase
ligase
!
! This type of system can repair many types of DNA damage,
This type of system can repair many types of DNA damage,
including
including
–
– Thymine
Thymine dimers
dimers and chemically modified bases
and chemically modified bases
!
! NER is found in all eukaryotes and prokaryotes
NER is found in all eukaryotes and prokaryotes
–
– However, its molecular mechanism is better understood in prokaryotes
However, its molecular mechanism is better understood in prokaryotes
Nucleotide Excision Repair System
Nucleotide Excision Repair System
7. 7
7
DNA REPAIR of damaged base:
DNA REPAIR of damaged base:
Nucleotide Excision Repair fixes errors created by mutagens
Nucleotide Excision Repair fixes errors created by mutagens
!
! Excision repair
Excision repair
enzymes release
enzymes release
damaged regions of
damaged regions of
DNA.
DNA.
!
! Single strand
Single strand
released
released
!
! Repair is then
Repair is then
completed by DNA
completed by DNA
polymerase and DNA
polymerase and DNA
ligase
ligase
8. 8
8
!
! Several human diseases have been shown to involve
Several human diseases have been shown to involve
inherited defects in genes involved in NER
inherited defects in genes involved in NER
–
– These include
These include xeroderma
xeroderma pigmentosum
pigmentosum (XP) and
(XP) and Cockayne
Cockayne
syndrome (CS)
syndrome (CS)
"
" A common characteristic of both syndromes is an increased sensitivity
A common characteristic of both syndromes is an increased sensitivity
to sunlight
to sunlight
–
– Xeroderma
Xeroderma pigmentosum
pigmentosum can be caused by defects in seven
can be caused by defects in seven
different NER genes
different NER genes
Nucleotide Excision Repair Removes
Nucleotide Excision Repair Removes
Damaged DNA Segments
Damaged DNA Segments
10. 10
10
Mistakes during replication alter genetic
Mistakes during replication alter genetic
information
information
!
! Errors during replication are exceedingly rare, less
Errors during replication are exceedingly rare, less
than once in 10
than once in 109
9 base pairs
base pairs
!
! Proofreading enzymes correct errors made during
Proofreading enzymes correct errors made during
replication
replication
–
– DNA polymerase has 3
DNA polymerase has 3’
’ –
– 5
5’
’ exonuclease
exonuclease activity which
activity which
recognizes mismatched bases and excises
recognizes mismatched bases and excises them
them
–
– If errors slip through proofreading:
If errors slip through proofreading:
"
" In bacteria, methyl-directed mismatch repair finds these errors on
In bacteria, methyl-directed mismatch repair finds these errors on
newly synthesized strands and corrects them
newly synthesized strands and corrects them
"
" In
In euks
euks, mismatch repair
, mismatch repair finds these errors on newly synthesized
finds these errors on newly synthesized
strands and corrects them
strands and corrects them
12. 12
12
Mismatch Repair System
Mismatch Repair System
!
! If proofreading fails, the
If proofreading fails, the methyl-directed
methyl-directed
mismatch repair
mismatch repair system comes to the rescue
system comes to the rescue
!
! This repair system is found in all species
This repair system is found in all species
!
! In humans, mutations in the system are
In humans, mutations in the system are
associated with particular types of cancer
associated with particular types of cancer
!
! Methyl-directed mismatch repair
Methyl-directed mismatch repair
recognizes mismatched base pairs, excises
recognizes mismatched base pairs, excises
the incorrect bases, and then carries out
the incorrect bases, and then carries out
repair synthesis.
repair synthesis.
14. 14
14
Mismatch Repair in Eukaryotes
Mismatch Repair in Eukaryotes
!
! Eukaryotes also have mismatch repair, but it is
Eukaryotes also have mismatch repair, but it is
not clear how old and new DNA strands are
not clear how old and new DNA strands are
identified.
identified.
–
– Four genes are involved in humans,
Four genes are involved in humans, hMSH2
hMSH2 and
and
hMLH1, hPMS1,
hMLH1, hPMS1, and
and hPMS2
hPMS2
–
– All of these are
All of these are mutator
mutator genes
genes
–
– mutation in any one of them confers hereditary
mutation in any one of them confers hereditary
predisposition to hereditary
predisposition to hereditary nonpolyposis
nonpolyposis colon cancer
colon cancer
(HNPCC: OMIM 120435).
(HNPCC: OMIM 120435).
15. 15
15
Transpositions
Transpositions
!
! Cytologically
Cytologically invisible sequence rearrangement:
invisible sequence rearrangement:
–
– movement of a segment of DNA from one location to another in the
movement of a segment of DNA from one location to another in the
genome.
genome.
–
– Not a translocation
Not a translocation…
….
.
!
! This may be a transfer of DNA or a duplication of DNA.
This may be a transfer of DNA or a duplication of DNA.
!
! The sequences that cause
The sequences that cause transpostions
transpostions
–
– are called
are called transposable elements
transposable elements,
,
–
– have specific characteristics,
have specific characteristics,
–
– notably the potential to propagate themselves.
notably the potential to propagate themselves.
!
! Transposable elements are found in virtually all organisms.
Transposable elements are found in virtually all organisms.
16. 16
16
Types of Transposable Elements
!
! Transposons
Transposons:
:
–
– Move their DNA directly without the requirement of an
Move their DNA directly without the requirement of an
RNA intermediate.
RNA intermediate.
!
! Retroposons
Retroposons:
:
–
– Copy and then
Copy and then move the copied DNA
move the copied DNA
–
– via reverse transcription of an RNA intermediate.
via reverse transcription of an RNA intermediate.
17. 17
17
Transposons
Transposons
!
! Encode an enzyme called
Encode an enzyme called Transposase
Transposase.
.
!
! Rather than converting RNA to DNA, this enzyme:
Rather than converting RNA to DNA, this enzyme:
–
– directly removes the DNA sequence and
directly removes the DNA sequence and
–
– inserts it in another location.
inserts it in another location.
!
! Transposons
Transposons usually have inverted repeats (IR) on
usually have inverted repeats (IR) on
either side upstream and downstream.
either side upstream and downstream.
19. 19
19
Transposons
Transposons
!
! Transposase
Transposase excises the sequence between the
excises the sequence between the
inverted repeats and inserts it into another region of
inverted repeats and inserts it into another region of
the genome
the genome
!
! The gap created is widened by
The gap created is widened by exonucleases
exonucleases.
.
!
! The gap is filled in by repair enzymes that use the sister
The gap is filled in by repair enzymes that use the sister
chromatid
chromatid or homologous chromosome as a template to fill
or homologous chromosome as a template to fill
the gap.
the gap.
–
– If copying from a sister
If copying from a sister chromatid
chromatid (also containing the
(also containing the transposon
transposon) it
) it
will reappear in the original location
will reappear in the original location
"
" And hence be copied in the genome
And hence be copied in the genome
–
– If the homologous chromosome is used to replace missing
If the homologous chromosome is used to replace missing transposon
transposon
(did not have
(did not have transposon
transposon) it will
) it will not be replaced
not be replaced
"
" And hence not be copied in the genome
And hence not be copied in the genome
21. 21
21
Retroposons
Retroposons
!
! The DNA sequence in a
The DNA sequence in a retroposon
retroposon codes for a
codes for a
reverse transcriptase
reverse transcriptase,
,
–
– which catalyzes the formation of DNA from an RNA
which catalyzes the formation of DNA from an RNA
template.
template.
!
! They always copy DNA and cause a duplication
They always copy DNA and cause a duplication
!
! Many
Many retroposons
retroposons also have other polypeptide
also have other polypeptide
coding sequences.
coding sequences.
!
! Many
Many retroposons
retroposons have a poly A tail.
have a poly A tail.
!
! Others have direct repeat sequences on either side,
Others have direct repeat sequences on either side,
–
– these are generated because of the way the DNA
these are generated because of the way the DNA
sequence has been inserted.
sequence has been inserted.
25. 25
25
Homework Problems
Homework Problems
!
! Chapter
Chapter 15
15
!
! # 15
# 15
!
! DON
DON’
’T forget to take the online QUIZ
T forget to take the online QUIZ
!
! DON
DON’
’T forget to submit the online
T forget to submit the online iActivity
iActivity
–
– “
“Overview B
Overview B”
”
