Spatio-temporally controlled site-specific somatic mutagenesis in the mouse.
Fiche publication
Date publication
décembre 1997
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, Dr VONESCH Jean-Luc
Tous les auteurs :
Brocard J, Warot X, Wendling O, Messaddeq N, Vonesch JL, Chambon P, Metzger D
Lien Pubmed
Résumé
The efficient introduction of somatic mutations in a given gene, at a given time, in a specific cell type will facilitate studies of gene function and the generation of animal models for human diseases. We have shown previously that conditional recombination-excision between two loxP sites can be achieved in mice by using the Cre recombinase fused to a mutated ligand binding domain of the human estrogen receptor (Cre-ERT), which binds tamoxifen but not estrogens. DNA excision was induced in a number of tissues after administration of tamoxifen to transgenic mice expressing Cre-ERT under the control of the cytomegalovirus promoter. However, the efficiency of excision varied between tissues, and the highest level ( approximately 40%) was obtained in the skin. To determine the efficiency of excision mediated by Cre-ERT in a given cell type, we have now crossed Cre-ERT-expressing mice with reporter mice in which expression of Escherichia coli beta-galactosidase can be induced through Cre-mediated recombination. The efficiency and kinetics of this recombination were analyzed at the cellular level in the epidermis of 6- to 8-week-old double transgenic mice. We show that site-specific excision occurred within a few days of tamoxifen treatment in essentially all epidermis cells expressing Cre-ERT. These results indicate that cell-specific expression of Cre-ERT in transgenic mice can be used for efficient tamoxifen-dependent, Cre-mediated recombination at loci containing loxP sites to generate site-specific somatic mutations in a spatio-temporally controlled manner.
Mots clés
Animals, Epidermis, metabolism, Gene Targeting, Humans, Integrases, genetics, Mice, Mice, Transgenic, Mutagenesis, Site-Directed, Receptors, Estrogen, genetics, Recombinant Fusion Proteins, genetics, Recombination, Genetic, Tamoxifen, metabolism, Viral Proteins
Référence
Proc. Natl. Acad. Sci. U.S.A.. 1997 Dec;94(26):14559-63