Figure 2

SENP1 and SENP2 deSUMOylate PR-B and enhance their transcriptional activity. A) DeSUMOylation of PR-B by wt SENP1 and SENP2. HeLa cells were cotransfected with pSG5-PR-B, GFP-SUMO-1 and SENP1 or SENP2 as indicated. Cells were grown in the presence (+) or absence (-) of R5020. PR in cell extracts separated on SDS-PAGE, were detected with anti-PR 1294 monoclonal antibody. β-actin served as a loading control. B) HeLa cells were transfected with the PRE2-Luc reporter plasmid in the presence of pSV40-Renilla as internal control along with PR-B and increasing amounts (50-1000 ng) of SENP1, SENP1 mutant, or SENP2 expression vectors, or an empty vector control (-). Cells were treated without (-) or with (+) 10 nM R5020 for 24 hrs before being assayed for luciferase activity. The relative luciferase activity of wt PR-B in the presence of 10 nM R5020 is set as 100%.