Fig. 2.

ESCO2 interacts with PCNA outside of replication foci. (A) Tethered ESCO2 recruits PCNA. An mCherry-lacI-ESCO2 fusion protein, expressed in U2OS cells containing a stably integrated tandem array of lac operator sequences, is recruited to nuclear loci where it can be seen as 1 or 2 red foci in confocal micrographs. Cotransfected GFP-PCNA was recruited to the ESCO2 focus (Top), and this was unaffected by deletion of the PIP box in ESCO2 (Middle). PCNA was not enriched in the foci in cells containing mCherry-lacI (without ESCO2) (Bottom). (Scale bars, 10 μm.) (B) Normalized fluorescence intensity. The fluorescence intensity of lines drawn across the lac operator arrays from a number of cells treated as in A was averaged. Fluorescence of mCherry-lacI fusion is indicated by a red line, DAPI in blue, and GFP in green. n > 25 cells in each sample. Shown is a representative example of 3 independent experiments.