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Chapter
14
Recombination
and repair
14.1 Introduction
14.2 Breakage and reunion involves heteroduplex DNA
14.3 Double-strand breaks initiate recombination
14.4 Double-strand breaks initiate snapsis
14.5 Bacterial recombination involves single-strand assimilation
14.6 Gene conversion accounts for interallelic recombination
14.7 Topological manipulation of DNA
14.8 Specialized recombination involves breakage and reunion
at
specific sites
14.9 Repair systems correct damage to DNA
14.10 Excision repair systems in E. coli
14.11 Controlling the direction of mismatch repair
14.12 Retrieval systems in E. coli
14.13 RecA triggers the SOS system
14.14 Eukaryotic repair systems
14.1 Introduction
Bivalent is the structure containing all four chromatids (two representing
each homologue) at the start of meiosis.
Breakage and reunion describes the mode of genetic recombination, in
which two DNA duplex molecules are broken at corresponding points and
then rejoined crosswise (involving formation of a length of heteroduplex
DNA around the site of joining).
Site-specific recombination occurs between two specific (not necessarily
homologous) sequences, as in phage integration/excision or resolution of
cointegrate structures during transposition.
Synapsis describes the association of the two pairs of sister chromatids
representing homologous chromosomes that occurs at the start of meiosis;
resulting structure is called a bivalent.
Synaptonemal complex describes the morphological structure of synapsed
chromosomes.
Transposition refers to the movement of a transposon to a new site in the
genome.
14.1
Introduction
Figure 14.1 Recombination
occurs during the first
meiotic prophase. The stages
of prophase are defined by
the appearance of the
chromosomes, each of which
consists of two replicas
(sister chromatids), although
the duplicated state becomes
visible only at the end. The
molecular interactions of any
individual crossing-over
event involve two of the four
duplex DNAs.
14.1
Introduction
Figure 1.22 The ABO
blood group locus codes
for a galactosyltransferase
whose specificity
determines the blood
group.
14.2 Breakage and reunion involves
heteroduplex DNA
Branch migration describes the ability of a DNA strand
partially paired with its complement in a duplex to extend
its pairing by displacing the resident strand with which it
is homologous.
Hybrid DNA is another term for heteroduplex DNA.
Recombinant joint is the point at which two recombining
molecules of duplex DNA are connected (the edge of the
heteroduplex region).
14.2 Breakage and
reunion involves
heteroduplex DNA
Figure 14.1 Recombination occurs
during the first meiotic prophase.
The stages of prophase are defined
by the appearance of the
chromosomes, each of which
consists of two replicas (sister
chromatids), although the
duplicated state becomes visible
only at the end. The molecular
interactions of any individual
crossing-over event involve two of
the four duplex DNAs.
14.2 Breakage and
reunion involves
heteroduplex DNA
Figure 14.2
Recombination between
two paired duplex DNAs
could involve reciprocal
single-strand exchange,
branch migration, and
nicking.
14.2 Breakage and
reunion involves
heteroduplex DNA
Figure 14.3 Branch
migration can occur
in either direction
when an unpaired
single strand
displaces a paired
strand.
14.2 Breakage and
reunion involves
heteroduplex DNA
Figure 14.4 Resolution
of a Holliday junction
can generate parental or
recombinant duplexes,
depending on which
strands are nicked. Both
types of product have a
region of heteroduplex
DNA.
14.3 Double-strand
breaks initiate
recombination
Figure 14.5
Recombination is
initiated by a doublestrand break, followed
by formation of singlestranded 3￠ ends, one
of which migrates to a
homologous duplex.
14.4 Double-strand breaks initiate synapsis
Figure 14.6 The synaptonemal complex brings
chromosomes into juxtaposition. This example of Neotellia
was kindly provided by M. Westergaard and D. Von
Wettstein.
14.4 Double-strand breaks initiate synapsis
Figure 14.7 Double-strand breaks appear when axial
elements form, and disappear during the extension of
synaptonemal complexes. Joint molecules appear and
persist until DNA recombinants are detected at the end of
pachytene.
14.4 Double-strand
breaks initiate
synapsis
Figure 14.8 Spo11 is
covalently joined to
the 5￠ ends of
double-strand breaks.
14.5 The bacterial
RecBCD system is
stimulated by chi
sequences
Figure 14.9 RecBCD
nuclease approaches a chi
sequence from one side,
degrading DNA as it
proceeds; at the chi site, it
makes an endonucleolytic
cut, loses RecD, and
retains only the helicase
activity.
14.6 RecA catalyzes singlestrand assimilation
Paranemic joint describes a region in which two
complementary sequences of DNA are associated
side by side instead of being intertwined in a
double helical structure.
Single-strand assimilation describes the ability of
RecA protein to cause a single strand of DNA to
displace its homologous strand in a duplex; that is,
the single strand is assimilated into the duplex.
14.6 RecA catalyzes single-strand assimilation
Figure 14.10 RecA promotes the assimilation of invading single strands
into duplex DNA so long as one of the reacting strands has a free end.
14.6 RecA
catalyzes singlestrand
assimilation
Figure 14.2
Recombination
between two paired
duplex DNAs could
involve reciprocal
single-strand exchange,
branch migration, and
nicking.
14.6 RecA
catalyzes singlestrand
assimilation
Figure 14.11 RecAmediated strand
exchange between
partially duplex and
entirely duplex DNA
generates a joint
molecule with the same
structure as a
recombination
intermediate.
14.6 RecA catalyzes single-strand assimilation
Figure 14.12
RuvAB is an
asymmetric
complex
that
promotes
branch
migration of
a Holliday
junction.
14.6 RecA catalyzes
single-strand
assimilation
Figure 14.13 Bacterial enzymes
can catalyze all stages of
recombination in the repair
pathway following the
production of suitable
substrate DNA molecules.
14.8 Gene conversion accounts for
interallelic recombination
Gene conversion is the alteration of one strand of a
heteroduplex DNA to make it complementary with
the other strand at any position(s) where there were
mispaired bases.
Postmeiotic segregation describes the segregation
of two strands of a duplex DNA that bear different
information (created by heteroduplex formation
during meiosis) when a subsequent replication
allows the strands to separate.
14.8 Gene conversion accounts for
interallelic recombination
Figure 14.14 Spore
formation in the
Ascomycetes
allows
determination of
the genetic
constitution of
each of the DNA
strands involved
in meiosis.
14.9 Topological manipulation of DNA
Supercoiling describes the coiling of a closed
duplex DNA in space so that it crosses over its own
axis.
Topological isomers are molecules of DNA that
are identical except for a difference in linking
number.
Twisting number of a DNA is the number of base
pairs divided by the number of base pairs per turn
of the double helix.
Writhing number is the number of times a duplex
axis crosses over itself in space.
14.9 Topological manipulation of DNA
Figure 14.14 Spore
formation in the
Ascomycetes allows
determination of the
genetic constitution
of each of the DNA
strands involved in
meiosis.
14.9 Topological
manipulation of DNA
Figure 14.15 Separation
of the strands of a DNA
double helix could be
achieved by several
means.
14.9 Topological manipulation of DNA
Figure 9.18 E. coli sigma factors recognize
promoters with different consensus sequences.
(Numbers in the name of a factor indicate its
mass.)
14.9 Topological
manipulation of DNA
Figure 14.16 Bacterial type I
topoisomerases recognize
partially unwound
segments of DNA and pass
one strand through a break
made in the other.
14.9 Topological
manipulation of DNA
Figure 14.17 Type II
topoisomerases can pass a
duplex DNA through a
double-strand break in
another duplex.
14.9 Topological
manipulation of DNA
Figure 14.18 DNA
gyrase may introduce
negative supercoils in
duplex DNA by
inverting a positive
supercoil.
14.10 Specialized recombination involves
breakage and reunion at specific sites
att sites are the loci on a phage and
the bacterial chromosome at which
recombination integrates the phage
into, or excises it from, the bacterial
chromosome.
14.10 Specialized
recombination involves
breakage and reunion at
specific sites
Figure 14.19 Circular phage DNA
is converted to an integrated
prophage by a reciprocal
recombination between attP and
attB; the prophage is excised by
reciprocal recombination
between attL and attR.
14.10 Specialized
recombination
involves breakage
and reunion at
specific sites
Figure 14.20 Does
recombination
between attP and
attB proceed by
sequential
exchange or
concerted cutting?
14.10 Specialized recombination involves
breakage and reunion at specific sites
Figure 14.21 Staggered cleavages in the common core
sequence of attP and attB allow crosswise reunion to
generate reciprocal recombinant junctions.
14.10 Specialized
recombination
involves breakage
and reunion at
specific sites
Figure 14.2
Recombination
between two paired
duplex DNAs could
involve reciprocal
single-strand
exchange, branch
migration, and
nicking.
14.10 Specialized recombination involves
breakage and reunion at specific sites
Figure 14.22 Int and IHF bind to different
sites in attP. The Int recognition sequences in
the core region include the sites of cutting.
14.10 Specialized recombination
involves breakage and reunion at
specific sites
Figure 14.23 The Int binding
sites in the core lie on one face
of DNA. The large circles
indicate positions at which
methylation is influenced by Int
binding; the large arrows
indicate the sites of cutting.
Photograph kindly provided by
A. Landy.
14.10 Specialized
recombination involves
breakage and reunion at
specific sites
Figure 14.24 Multiple
copies of Int protein may
organize attP into an
intasome, which initiates
site-specific
recombination by
recognizing attB on free
DNA.
14.11 Repair systems correct damage to DNA
Figure 14.25
Substitutions of
individual bases
create mismatched
pairs that may be
corrected by
replacing one base;
if uncorrected they
cause a mutation in
one daughter
duplex.
14.11 Repair
systems correct
damage to DNA
Figure 14.26
Modifications or
removal of bases may
cause structural
defects that prevent
replication or induce
mutations in each
replication cycle until
they are removed.
14.11 Repair systems correct damage to DNA
Figure 14.14 Spore formation in the
Ascomycetes allows determination of the
genetic constitution of each of the DNA
strands involved in meiosis.
14.12 Excision repair systems in E. coli
Excision of phage or episome or
other sequence describes its
release from the host
chromosome as an autonomous
DNA molecule.
14.12 Excision repair
systems in E. coli
Figure 14.27 Excisionrepair removes and
replaces a stretch of
DNA that includes the
damaged base(s).
14.12 Excision repair
systems in E. coli
Figure 14.28 The Uvr
system operates in stages
in which UvrAB
recognizes damage,
UvrBC nicks the DNA,
and UvrD unwinds the
marked region.
14.12 Excision repair
systems in E. coli
Figure 14.28 The Uvr
system operates in stages
in which UvrAB
recognizes damage,
UvrBC nicks the DNA,
and UvrD unwinds the
marked region.
14.12 Excision repair
systems in E. coli
Figure 14.37 A helicase
unwinds DNA at a
damaged site,
endonucleases cut on
either side of the lesion,
and new DNA is
synthesized to replace the
excised stretch.
14.13 Base flipping is
used by methylases
and glycosylases
Figure 14.29 A
glycosylase
removes a base
from DNA by
cleaving the bond
to the deoxyribose.
14.13 Base flipping is
used by methylases
and glycosylases
Figure 14.30 A
glycosylase hydrolyzes
the bond between base
and deoxyribose (using
H20), but a lyase takes
the reaction further by
pening the sugar ring
(using NH2).
14.13 Base flipping is
used by methylases and
glycosylases
Figure 14.31 A
methylase "flips" the
target cytosine out of
the double helix in
order to modify it.
Photograph kindly
provided by Rich
Roberts.
14.15 Controlling the direction of mismatch repair
Figure 14.32
Preferential removal
of bases in pairs that
have oxidized
guanine is designed
to minimize
mutations.
14.15 Controlling the direction of mismatch repair
Figure 13.30
Replication of
methylated DNA gives
hemimethylated DNA,
which maintains its
state at GATC sites
until the Dam
methylase restores the
fully methylated
condition.
14.15 Controlling the
direction of mismatch repair
Figure 14.33 GATC sequences
are targets for the Dam
methylase after replication.
During the period before this
methylation occurs, the
nonmethylated strand is the
target for repair of
mismatched bases.
14.15 Controlling the
direction of mismatch
repair
Figure 14.34 MutS
recognizes a mismatch
and translocates to a
GATC site. MutH cleaves
the unmethylated strand
at the GATC.
Endonucleases degrade
the strand from the GATC
to the mismatch site.
14.16
Retrival
systems in
E. coli
Figure 14.31 An E. coli
retrieval system uses a
normal strand of DNA
replace the gap left in a
newly synthesized strand
opposite a site of unrepaired
damage.
14.17 RecA
triggers the
SOS system
Figure 14.32 The
LexA protein
represses many genes,
including repair
functions, recA and
lexA,. Activation of
RecA leads to
proteolytic cleavage
of LexA and induces
all of these genes.
14.17 RecA
triggers the
SOS system
Figure 14.32 The
LexA protein
represses many genes,
including repair
functions, recA and
lexA,. Activation of
RecA leads to
proteolytic cleavage
of LexA and induces
all of these genes.
14.18 Eukaryotic
repair systems
Figure 14.37 A helicase
unwinds DNA at a
damaged site,
endonucleases cut on
either side of the lesion,
and new DNA is
synthesized to replace the
excised stretch.
Summary