Proteasome activator 200 kDa (PA200) forms nuclear foci after exposure of cells to ionizing radiation and enhances proteasome activity in vitro. Within cells, it is unclear whether PA200 responds to radiation alone or in association with proteasomes. In the present study, we identified three forms of cellular PA200 (termed PA200i, ii and iii) at the mRNA and protein levels. Neither PA200ii nor PA200iii appears to associate with proteasomes. All detectable PA200i is associated with proteasomes, which indicates that PA200i and proteasomes function together within the cell. Consistent with this idea, we find that exposure of cells to radiation leads to an equivalent accumulation of both PA200i and core proteasomes on chromatin. This increase in PA200 and proteasomes on chromatin is not specific to the stage of cell cycle arrest since it occurs in cells that arrest in G(2)/M and cells that arrest in G(1)/S after exposure to radiation. These data provide evidence that PA200 and proteasomes function together within cells and respond to a specific radiation-induced damage independent of the stage of cell cycle arrest.