Progestin inhibition of progesterone receptor gene expression in human breast cancer cells.
The present study was designed to investigate whether inhibition of progesterone receptor (PR) gene transcription and/or regulation of PR mRNA half-life were involved in the progestin-mediated decrease of PR in T-47D human breast cancer cells. Cells were treated with the progestin ORG 2058 and PR mRNA measured by Northern blot analysis of total RNA. A major PR mRNA around 13.5 kilobases and minor species around the 28S ribosomal RNA subunit were decreased upon ORG 2058 treatment. The decrease was not detectable until 2-3 h after treatment and was the same at all ORG 2058 concentrations (1-100 nM) tested. The decrease in PR mRNA was unaffected by actinomycin D in the first 3 h but was inhibited thereafter. There was a partial recovery of PR mRNA levels 24 h after ORG 2058 exposure. Immunoblot analysis showed that immunoreactive PR decreased in parallel with PR mRNA. The rate of protein loss in the first 12 h after progestin treatment was related to the ORG 2058 concentration used. Nuclear run-on experiments showed that ORG 2058 caused a decrease of up to 70% in the transcription rate of the PR gene. The half-life of PR mRNA was shown to be 2-2.5 h by [3H]uridine incorporation, which was much shorter than estimates obtained using actinomycin D, and was unaffected by ORG 2058. In summary, these data have shown that the mechanism by which progestins decrease the concentration of PR includes inhibition of transcription of the PR gene.
Alexander
I E
IE
Garvan Institute of Medical Research, St. Vincent's Hospital, Sydney, New South Wales, Australia.
Clarke
C L
CL
Shine
J
J
Sutherland
R L
RL
D015152:Blotting, Northern; D001943:Breast Neoplasms / Q000503:physiopathology*; D002460:Cell Line; D003609:Dactinomycin / Q000494:pharmacology; D015536:Down-Regulation; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression / Q000187:drug effects*; D006801:Humans; D011282:Pregnenediones / Q000493:pharmacokinetics*; D045167:Progesterone Congeners / Q000493:pharmacokinetics; D012333:RNA, Messenger / Q000187:drug effects; D011980:Receptors, Progesterone / Q000235:genetics*; D014158:Transcription, Genetic / Q000187:drug effects
D011282:Pregnenediones; D045167:Progesterone Congeners; D012333:RNA, Messenger; D011980:Receptors, Progesterone; D003609:Dactinomycin; C020175:16 alpha-ethyl-21-hydroxy-19-nor-4-pregnene-3,20-dione