Regulation of in vitro transcription by progesterone receptor. Characterization and kinetic studies

M. K. Bagchi, S. Y. Tsai, N. L. Weigel, M. J. Tsai, B. W. O'Malley

Research output: Contribution to journalArticlepeer-review

Abstract

We have devised an in vitro assay system to study the transcriptional activity of native chicken progesterone receptor (cPR). Purified cPR added to cell-free extracts from HeLa cell nuclei stimulates accurate transcription from a promoter driven by two progesterone response elements. The transcriptional enhancement is entirely progesterone response element dependent and promoter specific. We have defined the appropriate conditions of template, nuclear extract, salt, and magnesium ion requirements for efficient transcriptional enhancement by cPR. Under optimized conditions, a synthesis rate of one transcript/20 promoters is achieved in the presence of saturating amounts of cPR. Kinetic studies suggest that the progesterone receptor can form a functional preinitiation complex with RNA polymerase II and other transcription factors present in unfractionated HeLa nuclear extract. Following formation of this complex, transcription can commence rapidly upon addition of nucleotides.

Original languageEnglish (US)
Pages (from-to)5129-5134
Number of pages6
JournalJournal of Biological Chemistry
Volume265
Issue number9
StatePublished - 1990
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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