Figure 4.

Binding to Ub is required for polι to relocalize into replication foci. (A) Wild-type pEGFP polι (upper panels) or pEGFP polι P692R (lower panels) were transfected into MRC5 cells and either mock treated (left hand panels) or irradiated with 7 J/m² UV (right-hand panels). Nuclei from two adjacent cells are shown in each panel for comparison. (B) Histogram showing the percentage of nuclei with foci after exposure to various concentrations of the 26S proteasome inhibitor, NP-L₃-VS before (closed bars) and after UV-irradiation (open bars). Numbers are based upon 200 nuclei analyzed per data point. (C) Western blot of Ub or PCNA in whole-cell extracts (50 μg) separated by SDS–PAGE on a 15% gel, after treatment with NP-L₃-VS with and without UV-irradiation (7 J/m²). Note that the levels of Ub and Ub-PCNA decrease as the dose of NP-L₃-VS is increased.