A chimeric mammalian transactivator based on the lac repressor that is regulated by temperature and isopropyl β-D-thiogalactopyranoside

S. B. Baim, M. A. Labow, A. J. Levine, T. Shenk

Research output: Contribution to journalArticle

87 Scopus citations

Abstract

LAP267 is a lacI activator protein (LAP) containing an insertion of the transcriptional activation domain of the herpes simplex virus virion protein 16 within the inducer-binding and dimerization domain of the lac repressor protein. LAP267 strongly induces expression in a conditional manner from a minimal simian virus 40 early promoter linked to lac operator sequences. LAP267 is temperature-sensitive, activating expression at 32°C but not at 39.5°C. It is allosterically regulated in a manner opposite that of wild-type lac repressor, in that LAP267 activity is rescued at the nonpermissive temperature by isopropyl β-D-thiogalactopyranoside (IPTG). Stable mouse cell lines containing both the LAP267 gene and a LAP-inducible chloramphenicol acetyltransferase (CAT) reporter gene were readily established and exhibited up to a 1200-fold increase in CAT activity within 24 hr upon addition of IPTG. Thus, LAP267 is a powerful inducible switch in mammalian cells, imparting a regulatory stringency similar to that observed with lac repressor in Escherichia coli.

Original languageEnglish (US)
Pages (from-to)5072-5076
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume88
Issue number12
DOIs
StatePublished - 1991

All Science Journal Classification (ASJC) codes

  • General

Keywords

  • reverse regulation
  • temperature-sensitive activation
  • transcriptional regulation

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