Zusammenfassung der Ressource
Repair of DNA double
strand breaks by protein
repair machines
- DNA
damage
- DNA is composed of
phosphate backbone and
sugar moiety attached to a
phosphate
- Moieties are
prone to damage
- Most
damage
occurs
intracelluarly
- Replication errors
- Double strand
breaks are the
most lethal form of
damage
- Replication cannot
proceed if DNA is
broken
- DNA damage
can be a result of
- Cancer
- Ageing
- Neurological
dysfunction
- Neuronal cells are particularly
sensitive to DNA damage as they
can't be replaced
- Causes of DNA breaks
- Exogenous
- Radiation
- Chemicals
- Endogenous
- Oxygen: free radicals
- Can produce
SSB and DSB
- DNA replication
- Specialised
- V(D)J
recombination
- Cell actively produces
breaks and sticks them back
together to produce
antibodies
- Class switching
- Changing of
one antibody to
another
- Meiosis
- Most of the time this
damage is repaired
- If there is lots of damage the
kinase signalling pathway is
activated
- Apoptosis
- If the break is
incorrectly repaired this
causes genome
instability resulting in
cancer
- Unrepaired DNA
breaks cause
translocations
- The arm of one
chromosome is
transferred to
another
- Karyotype of an
advanced tumour
will look very
mixed up
- Accumulation of broken chromosomes is
an early marker of uncontrolled cell
growth
- You can grade cells on
appearance of DSBs
- As cells go from normal
to cancer cells, major
rearrangement occurs
- Transition from
normal to cancer cells
is mediated by
genome instability
- Double strand break repair pathways
- Homologous recombination
- 2 homologous DNA molecules aligned
- Despite the high degree of similarity
there are small differences such as
sequence variants for different alleles
- Formation of initial short regions of base
pairing between the 2 recombining DNA
molecules
- Strand invasion
- Single strand region of DNA from parental
molecule pairs with complementary strand
on homologous duplex DNA molecule
- Process regions of new duplex DNA are
generated- heteroduplex DNA
- Holliday junction formation
- 2 DNA molecules become connected by
crossing DNA strands
- Branch migration
- Holliday junction can move along DNA by
repeated melting and formation of base pairs
- Identical base pairs are formed in the
recombination intermediate
- Cleavage of Holliday junction: resolution
- Cutting DNA strands in Holliday junction
regenerates 2 separate duplex DNA
molecules and finishes genetic exchange
- The DNA strands cut has a large impact on
the extent of DNA exchange that occurs
between the 2 recombining molecules
- Non-homologous end joining (NHEJ)
- Sequence
information is lost
from broken ends
- Original sequence across
the break is not faithfully
restored during NHEJ
- 2 ends of broken DNA
are joined to each other
by misalignment
between single strands
protruding from both
ends
- Misalignment occurs by
pairing between tiny
stretches of complementary
bases
- Ku70 and Ku80 form a heterodimer that binds to DNA ends
- DNA-PKcs are recruited
- DNA PKcs form a complex with Artemis
- Artemis is a 5'-3' exonuclease and
latent endonuclease that is activated
by phosphorylation of DNA-PKcs
- These nucleolytic activities process broken ends and
prepare them for ligation
- Ligation is carried out by ligase
IV/XRCC4/Cernunnos-XLF
- Bacteria
- NHEJ occurs less
frequently in
bacteria
- Bacillus subtilis produces a
Ku-like protein and DNA ligase
when it sporulates and packages
proteins into a mature spore
- Mutant spores lacking
these proteins are
susceptible to dry heat
- Known to cause DNA breaks
- Upon germination, heated mutant spores unable
to resume growth as they are unable to rejoin
heat induced breaks
- Spores only have
1 chromosome
- Cannot rely on
sister chromatids
- Spore chromosome is
tightly coiled like a doughnut
- Holds ends of DNA
in close proximity
- Close juxtaposition
could facilitate correct
rejoining of ends even if
chromosome sustained
multiple breaks
- Ku is a
homodimer