OTUB1 represses ERα mediated transcription. A, HeLa cells were transfected with an ERE-luciferase reporter construct, ERα, OTUB1, or vector control. ERα-dependent transcription was measured in the presence of 1 nm estradiol (E2) or ethanol vehicle. B, OTUB1 represses ERα activity in the presence of coactivator SRC-3. HeLa cells were transfected with an ERE-luciferase reporter construct and indicated vectors and analyzed for luciferase activity. Representative result of three independent experiments is shown. C, OTUB1 represses ERβ activity. HeLa cells were transected with ERE-luciferase, ERβ, and the indicated vectors, and luciferase activity was analyzed. D, OTUB1 does not affect PR-B transcriptional activity. HeLa cells were transfected with a GRE-Luciferase reporter construct, PR-B, OTUB1, or vector control. PR-B dependent transcriptional activity was measured in the presence of 100 nm progesterone (P4) or ethanol vehicle. E, OTUB1 does not repress AF-2 domain of ERα. HeLa cells were transfected with pG5-luciferase reporter, pGAL4-AF2, and OTUB1 or empty vector. F, OTUB1 represses ERα lacking the AF1 domain. HeLa cells were transfected with ERE-luciferase, pCR3.1 ERα179C, and OTUB1 or empty vector.