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. 2012 Jul 19;8(7):e1002826. doi: 10.1371/journal.pgen.1002826

Figure 3. PCNA interacts in vivo with Ubp10.

Figure 3

(A) The sliding clamp PCNA and Ubp10 specific-ubiquitin protease interact physically in vivo. Co-immunoprecipitation assay showing physical interaction between Ubp10-myc and FLAG tagged PCNA. PCNA-FLAG was immunoprecipitated from formaldehyde-crosslinked protein extracts (see methods) both from untreated or 0.020% MMS-treated cells, blots were incubated with α-myc (to detect Ubp10) or α-FLAG (to detect PCNA). The immunoblots shown are those from untreated cells (a similar result was obtained with MMS-treated cells). As indicated the strains used in this assays were UBP10-myc POL30-FLAG and UBP10-myc POL30-FLAG rad18Δ. Immunoprecipitated Ubp10-myc was quantitated, normalized and plotted. Each immunoprecipitation experiment was repeated three times to gain an estimate of error. (B) Co-immunoprecipitation assay showing physical interaction between Ubp10-myc and PCNA. PCNA was immunoprecipitated both from untreated or 0.020% MMS-treated cells, blots were incubated with α-myc (to detect Ubp10) or α-PCNA. The immunoblots shown are those from MMS-treated cells (a similar result was obtained with untreated cells). As indicated the strains used in this assays were UBP10-myc and UBP10-myc rad18Δ. Immunoprecipitated Ubp10-myc was quantitated, normalized and plotted. Note that in our experiments we detect Ubp10 interacting with unmodified PCNA (or unmodified PCNA-FLAG).