Skip to main content
. 2008 Jan 17;22(4):823–837. doi: 10.1210/me.2007-0437

Figure 1.

Figure 1

Progestins Induce Rapid Activation of EGFR Upstream of Erk1/2 MAPK

A, Serum-starved T47D cells stably expressing wt PR-B (T47D-YB) were treated with R5020 (10 min). EGFR in whole-cell lysates was immunoprecipitated (IP). Immune complexes were subjected to Western blotting with phospho-Tyr 1173 Ab. Membranes were stripped and reprobed to detect total EGFR in each IP and lysates (10% input). IgG was included as a specificity control for IP-specific Ab. B, T47D-YB cells were pretreated with DMSO vehicle control, U0126, PP2 (10 μm each), or AG1478 (1 μm) for 30 min before addition of R5020 or EtOH for 10 min. Western blotting of whole-cell lysates was performed to detect phospho (p-) and total (t-) Erk1/2 MAPK. C, Serum-starved T47D-YB cells were incubated with DMSO or AG1478 for 30 min, followed by EGF (10 min) or PMA (30 min). Cell lysates were subjected to Western blotting as above; actin blots indicate equal protein loading. D, Triplicate cultures of PR-null (T47D-Y) or wt PR-B (T47D-YB) expressing cells were grown in the absence of steroid hormones for 48 h. After 18 h treatment with EtOH vehicle control, R5020, or EGF, cells were fixed and stained with propidium iodide. DNA content was measured by flow cytometry. Bars indicate percentage of cells in the indicated phase of the cell cycle: G1 (dark gray), S (black), G2/M (light gray); error bars represent sd (n = 3). E, Steroid-starved T47D-YB cells were pretreated for 30 min with DMSO vehicle control or AG1478 (10 μm), followed by 18 h of treatment with R5020 or EtOH vehicle control and analyzed as above. F, Steroid-starved T47D-YB cells were pretreated with vehicle control and the ER antagonist ICI 182,780 (0.10 μm) or PP2 or U0126 (10 μm each), treated with or without R5020 for 18 h, fixed with EtOH, and stained with propidium iodide. Bars indicate percentage of cells in the indicated phase of the cell cycle: G1 (dark gray), S (black), G2/M (light gray): error bars represent sd (n =3). All experiments were repeated at least three times with similar results.