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. Author manuscript; available in PMC: 2017 Feb 1.
Published in final edited form as: Mol Cancer Res. 2015 Nov 17;14(2):141–162. doi: 10.1158/1541-7786.MCR-15-0431

Figure 1. Stable expression of PR isoforms in ES-2 cells.

Figure 1

(A) Inset, Western blot analysis showing total PR expression in ES-2 cell pools expressing GFP-tagged empty vector control (EV), GFP-tagged PR-A (PR-A), or GFP-tagged PR-B (PR-B) and treated without or with R5020 (10 nM) for 24 hr. ES-2 cells expressing EV, PR-A, or PR-B were transiently transfected with a progesterone response element (2X-PRE) containing luciferase reporter gene and treated for 18 hr with R5020 (10 nM). Relative luciferase units (RLU) were normalized to the mean result ± standard deviation (SD) for Renilla luciferase expression (n=3, **p≤0.01). (B) RT-qPCR analysis of HEF1 and BIRC3 mRNA expression after 24 hr R5020 (10 nM) treatment in ES-2 cell pools expressing EV, PR-A or PR-B (n=3, *p≤0.05 **p≤0.01). (C) Western blot analysis of total PR expression in ES-2 cells stably expressing GFP-tagged EV control (clone #3), GFP-tagged PR-A (GFP-PR-A clone #1, #5, #4, #7), or GFP-tagged PR-B (GFP-PR-B clone #1, #3) relative to T47D breast cancer cells stably expressing PR-A-only (YA), PR-B-only (YB), and both endogenous PR isoforms (CO). Actin served as a loading control. (D) Western blot analysis of PR-A and PR-B phosphorylation at Ser294 and Ser190, and total PR protein expression in ES-2 PR-expressing cells treated with R5020 (10 nM) for 1 hr. ◆◆denotes a non-specific band present in the phospho-PR Ser294 blot. Actin served as a loading control.