Conditional site-specific recombination in mammalian cells using a ligand-dependent chimeric Cre recombinase.
Fiche publication
Date publication
juillet 1995
Journal
Proceedings of the National Academy of Sciences of the United States of America
Auteurs
Membres identifiés du Cancéropôle Est :
Pr CHAMBON Pierre, Dr METZGER Daniel
Tous les auteurs :
Metzger D, Clifford J, Chiba H, Chambon P
Lien Pubmed
Résumé
We have developed a strategy to generate mutant genes in mammalian cells in a conditional manner by employing a fusion protein, Cre-ER, consisting of the loxP site-specific Cre recombinase linked to the ligand-binding domain of the human estrogen receptor. We have established homozygous retinoid X receptor alpha-negative (RXR alpha-/-) F9 embryonal carcinoma cells constitutively expressing Cre-ER and have shown that estradiol or the estrogen agonist/antagonist 4-hydroxytamoxifen efficiently induced the recombinase activity, whereas no activity was detected in the absence of ligand or in the presence of the antiestrogen ICI 164,384. Furthermore, using a targeting vector containing a selection marker flanked by loxP sites, we have inactivated one retinoic acid receptor alpha allele in such a line, demonstrating that the presence of the recombinase does not inhibit homologous recombination. Combining this conditional site-specific recombination system with tissue-specific expression of Cre-ER may allow modification of the mammalian genome in vivo in a spatiotemporally regulated manner.
Mots clés
DNA Nucleotidyltransferases, genetics, Enzyme Induction, Estradiol, analogs & derivatives, Estrogen Antagonists, pharmacology, Gene Targeting, Genetic Vectors, Integrases, Mutagenesis, Site-Directed, Polyunsaturated Alkamides, Receptors, Estrogen, agonists, Receptors, Retinoic Acid, genetics, Recombinant Fusion Proteins, metabolism, Recombination, Genetic, Repetitive Sequences, Nucleic Acid, genetics, Tamoxifen, analogs & derivatives, Tumor Cells, Cultured, Viral Proteins
Référence
Proc. Natl. Acad. Sci. U.S.A.. 1995 Jul;92(15):6991-5