Biology Reference
In-Depth Information
sensitive for IR.
88,261
Similar to
Rad51
and
Rad52
mutants,
Rad54
mutants
were hypersensitive to IR as well as DNA cross-linking and alkylating agents
that eventually cause DSBs.
41
Rad54
mutants display only minor defects in
meiotic recombination in yeast due to the presence of the meiotic homolog
Rdh54/Tid1.
262
RAD54
knockdown in mice causes hypersensitivity to IR at
embryonic stages but not in adult stages due to rescue by NHEJ repair.
263
However, all developmental stages of
RAD54
-deficient mice are hypersensitive
to DNA cross-linking agents.
263
Mutational analysis of Walker A box-conserved
lysine indicated that in both mice and yeast, ATP hydrolysis of Rad54 is
essential for its function
in vivo
.
264-266
Rad54 expression levels increase during the late G1 phase of the cell
cycle,
267,268
presumably to connect HR repair of DSBs during the late S and
G2 phases.
269
Rad54
expression levels are upregulated during DSB formation
and Rad54 foci formation is dependent on Rad51.
64,270
However, Rad51 foci
formation is not dependent on Rad54, indicating that Rad54 acts downstream
of Rad51.
64
Similarly, RAD54 colocalizes with RAD51 foci following IR in
mammalian cells.
271,272
RAD54 belongs to the Swi2/Snf2 SF2 (superfamily 2) of proteins.
250-252
The
Snf2/Swi2 proteins are commonly known for dsDNA-dependent ATPase, ATP-
dependent chromatin remodeling, DNA translocase, and DNA-supercoiling activ-
ities.
250-252
Like other members of SF1 and SF2, members of RAD54 possess
several signature helicase motifs I, Ia, II, III, IV, V, and VI that constitute the two
tandem RecA-like lobes that utilize the energy of ATP binding and hydrolysis for
functioning.
273,274
However, unlike helicases, the SF2 family of proteins does not
unwind but translocates on dsDNA.
250,251
Also unlike helicases, the ATPase activity
of RAD54 is not stimulated by ssDNA, nor does it translocate on ssDNA.
274,275
Both yeast and human RAD54 are strictly dsDNA-dependent ATPases, with a
catalytic turnover rate ranging from 3000 to 6000 min
1
.
250
The binding affinity for
branched DNA structures such as PX junctions (partial Holliday junctions) is
approximately 200 times higher than for ssDNA or dsDNA.
276
RAD54 function has been implicated in all three stages of recombination:
presynapsis, synapsis, and postsynapsis.
251,252
These include interaction with
RAD51 to stabilize the ssDNA NPF, stimulation of homology search and strand
exchange catalyzed by RAD51, chromatin remodeling during the homology
search, disruption of RAD51-dsDNA filaments, branch migration of Holliday
junctions, and interaction with specific endonucleases to stimulate resolution of
Holliday junctions.
RAD54 interacts with RAD51 in a species-specific manner through its
N-terminal domain.
277-280
This interaction is seen with both free RAD51 and
the RAD51 ssDNA NPF.
281
The ATPase activity of RAD54 is not required for
RAD51 NPF stability. This was shown
in vivo
and
in vitro
using a RAD54
ATPase-deficient mutation where a Walker A box lysine to arginine substitution
allows only ATP binding.
281,282
