Biology Reference
In-Depth Information
Once HRR has been activated, there is a defined set of enzymatic reactions
that yields the final product. Within the process of homology search including
strand invasion, RAD51 is capable of the reaction
in vitro
without any histone
remodelers even in the presence of histones
32
and no histone modifications
have been associated.
In vitro
, the yeast SWI/SNF chromatin-remodeling
complex enhances the process specifically in heterochromatin.
29
This indicates
that although RAD51 is capable of performing the essential steps in the HRR
mechanism, there are steps in certain situations that require further chromatin
processing to be efficient in the nuclear environment. Otherwise, the chroma-
tin regulation associated with basic processes of HRR is still unclear to date.
Finally, the resolution of HRR requires that the chromatin structure be
restored. This process is still being elucidated. The most well-described portion
of resolution is the dephosphorylation of
g
-H2AX and this is consistent in both
HRR and NHEJ. This process is mediated by protein phosphatase 2A (PP2A)
in mammalian cells and phosphatase Pph3 in yeast or PP4C in mammalians.
33
Inactivation of these phosphatases leads to sustained
g
-H2AX and inefficient
repair. It is clear that these factors are responsible for the resolution of
g
-H2AX, but the activation of this process remains undescribed.
The removal of the acetylation events on H3 and H4 described earlier
requires multiple histone deacetylases (HDACs), which are activated in the
latter portion of the repair process and facilitate the condensation of chromatin
back into its predamage configuration. The specific pathways are reviewed by
Huertas
et al.
11
The chromatin methylation events H3K79me and H4K20me are a result of
the opening of the chromatin structure. It is reasonable to assume that this
process is reversed by reformation of the original chromatin structure and
these methylations being reburied within the tertiary structure. It is also
possible that there are other factors that facilitate the binding and reformation
of HP1 and KAP-1 structures associated with H3K9me3, but these have yet to
be described.
In the resolution of HRR, chromatin must also be replaced in its original
configuration. This is accomplished by the process of chromatin assembly,
though it is somewhat controversial because, as already described, it is not
completely clear whether the chromatin is truly evicted or not. In the case
where chromatin is evicted, it must be reestablished. Biochemically, two factors
have been implicated in this process: chromatin assembly factor 1 (CAF-1) and
antisilencing factor 1 (Asf1). CAF-1 is present at DSBs and is likely responsible
for the insertion of H3.
34
Further, CAF-1 and Asf1 interact and it is proposed
that Asf1 is recruited to sites of DNA damage in this fashion. Asf1 also stimu-
lates acetylation of free H3 on K56 via the HAT Rtt109 and this modified H3
inserted into the DNA flanking the repaired DSB is the signal for completion of
repair.
35
