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Molecular Immunogenetics Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104
The autoantigen Ku, composed of subunits Ku70 and Ku86, is
necessary for repair of DNA double-strand breaks by nonhomologous end
joining. Similarly, Ku participates in repair of DNA double-strand
breaks that occur during V(D)J recombination, and it is therefore
required for the development of B and T lymphocytes. Although previous
studies have identified the DNA-binding activities of Ku, little is
known concerning its dynamics, such as the mobility of Ku in the
nucleus and its rate of association with substrates. To address this
question, fluorescence photobleaching experiments were performed using
HeLa cells and B cells expressing a green fluorescent protein (GFP)
fusion construct of either Ku70 or Ku86. The results show that Ku moves
rapidly throughout the nucleus even following irradiation of the cells.
However, the rate of diffusion of Ku was 
100-fold slower than that
predicted from its size. Association of Ku-GFP with a filamentous
nuclear structure was also evident, and nuclear extraction experiments
suggest that this represents nuclear matrix. A central domain of Ku70
containing its DNA-binding and heterodimerization regions and its
nuclear localization signal shows that this alone is sufficient for the
observed mobility of Ku70-GFP and its association with nuclear matrix.
These data suggest the mobility of Ku is characterized by a transient,
high flux association with nuclear substrates that includes both DNA
and the nuclear matrix and may represent a mechanism for repair of
double-strand breaks using the nuclear matrix as a
scaffold.
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