Basic models of HR (based on how the HR is initiated)

• Holliday model: 1 single strand break on each paired DNA double

helix
• Meselson-Radding model: a single strand break on only one of the
paired DNA double helix, then anther single strand break on the other
strand
• Double strand break model (more plausible!!!)

• HR can occur in germ cells during meiosis as a controlled process to

produce diversified off-springs of an organism
• HR can be used to switch mating type in yeast
• HR can occur in somatic cells as a DNA repair mechanism
• HR is used to resolve stalled replication forks
Initiation of Homologous Recombination by Single Strand Break

Holliday Model Meselson-Radding Model

Genomes, Figure 14.29

Double strand break induced recombination.

Genomes, 2nd ed. Fig14.33

 Proteins may be required for involved in HR
• Initiation of DNA strand breaks: nuclease in some cases

• Helicase and end processing enzymes (nuclease)

• Single strand binding proteins (SSB or RPA)

• Strand invasion protein: Rec A in E. Coli and its family members

in eukaryotes

• Proteins helping RecA family to load on single strand DNA and

displace RPA

• Ligase to form Holliday junctions

• Helicase in branch migration

• Nuclease to resolve holiday junctions

• Others: DNA polymerase, topoisomerase, chromatin remodeling

 Proteins involved in DSB-induced HR
E. Coli Yeast Mammalian
DSB Damage HO, Spo11, Damage Nuclease, Damage,
replication Stalled replication Stalled replication forks,
forks, etc etc
End processing RecBCD MRE11/RAD50/XRS2 MRE11/RAD50/NSB1
? ??
Single strand binding SSB RPA RPA
protein
Strand invasion RecA RAD51 RAD51, RAD55, RAD57,
DCM1 BRCA2?

Helper for RAD51’s & RAD52 RAD52 ?, RAD54?,

facilitators for strand RAD54 BRCA2?
annealing, etc ????? ?????
Branch migration RuvAB ? ?

Resolution of Holliday RuvC Mus8? Mus8?

J.
Proteins involved in mammalian HR

End processing:
MRN nuclease complex: MRE11-RAD50-
NBS1
Other nucleases
Recombination:
RAD52
RAD51
RAD54 (helicase)
XRCC3
BRCA2
RPA
DNA polymerase, PCNA,
Other Helicase:
WRN: Werner’s syndrome gene
BLM: bloom syndrome gene
Accessory proteins (??):
Fanconi Anemia Proteins
BRCA1
ATM
ATR
Ratio and dynamics of these proteins are
important !!!

Valerie K & Povirk LF, Oncogene, 2003.

 Rec A protein and strand invasion

RAD51 protein family

• E Coli: RecA
• Yeast (Saccharomyces cerevisiae): RAD51, RAD55,RAD56, DCM1
• Mammalian cells: RAD51, RAD51B, RAD51C, RAD51D, RAD55,
RAD57, XRCC3
Other proteins that may facilitate the strand invasion or annealing:
RAD52, BRCA2(?), etc.
 Double-Strand Break Repair
Homologous Recombination (HR)
Binding and processing
RAD51 of the ends to reveal
RAD52 single stranded 3’-ends
RAD54
XRCC3 Single stranded DNA
invades the homologous
Helicases template
RPA
BRCA1 DNA synthesis and
BRCA2 branch migration to form
etc. a complicated structure
named Holliday Junction
(HJ)

Resolving of HJ to
complete the repair
(minior) process
(major)
Introduction: DNA Repair and Carcinogenesis
 Some interesting observation but full
mechanisms are not so clear

• Mismatches often slow down or inhibit HR

• In mismatch repair deficient cells, higher HR is often seen

• HR confers hypersensitivity to cross-links

• Activities of HR is often higher in S and G2 phase

• Resolution of replication blockage is often coupled with HR

• Homologous recombination between two similar (but not

completely homologous region is sometimes called
homeologous recombination.
Single Strand Annealing
Single-strand annealing - a repair
pathway shared between NHEJ and
HRR. SSA could occur when, at a DSB,
the DNA strand is resected by a
nuclease, for example, MRE11 to leave
ssDNA overhangs. The length of the
overhangs and the extent of homology
ranging from microhomologies to several
hundred bases or longer most likely
determine how SSA is executed. In the
case of long homologies between direct
repeats on the overhangs, RPA and
RAD52 are necessary for facilitating
DNA pairing followed by removal of the
tails by ERCC1/XPF nuclease and gap
filling by DNA polymerase as shown in
the figure. Sensors, such as ATM, may
signal and attract nucleases to the DSB.
Microhomology-based SSA, although
similar in principle, probably has quite a
different enzymology

Valerie K & Povirk LF, Oncogene, 2003. 22:5792

 Double-Strand Break Repair
Non-homologous End-Joining (NHEJ)
5'
3'
Steps:
(Exonuclease)

2. DNA ends processed

5'
3'
Reveal “micro-
homologies” at ends
(Anneal)
3. Micro-homologies
5'
3'
alignment
4. Ligase seals
(Synthesis and ligation)

5'
3'

•Imprecise (deletions and small insertions)

•Key role in chromosomal translocations
•Important in V(D)J recombination
Outline of the two principal pathways of DNA double-strand break repair. NHEJ involves six
known proteins, in a coordinated sequence to ensure ligation of the broken ends. Loss of
DNA sequence occurs at the site of repair. HR involves multiple proteins in a complex to
allow strand exchange and resolution. The process of HR can be an error-free process.
Simon N Powell and Lisa A Kachnic. 22: 5784-5791