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. Author manuscript; available in PMC: 2007 Apr 10.
Published in final edited form as: Steroids. 2006 Dec 14;72(2):188–201. doi: 10.1016/j.steroids.2006.11.009

Figure 6.

Figure 6

Phosphorylation of Ser294 PR-B is required for the regulation of a subset of endogenous PR target genes. A. RT-PCR showing differential upregulation of selected EGFR ligand transcript levels in response to phosphorylation of PR Ser294. T47D cells stably expressing either wt or S294A PR-B and treated with or without R5020 for 12 or 18 hours, and total cellular RNA was isolated with Trizol (Invitrogen) according to the manufacturer’s protocol. RNA was DNAse treated and cDNA was synthesized form 2ug of total RNA using MMLV-RT and random primers (BD Biosciences) in a total volume of 20ul. cDNA was used for PCR with EGF, TNFα and Actin specific primers and run on agarose gels. B. T47D-Y cells (PR null), and wt or S294A PR-B expressing cells were treated with vehicle or EGF for 30 min then vehicle or R5020 for 1 hour. cDNA was generated as above and PCR performed using HB-EGF or Actin specific primers.