The modified firefly luciferase reporter gene (luc+) but not Renilla luciferase is induced by all-trans retinoic acid in MCF-7 breast cancer cells.
Luciferase genes are widely used as reporters to analyze promoter and regulatory elements. We found that a luciferase reporter gene vector with a modified firefly luciferase gene (luc+), but not Renilla luciferase (Rluc), was induced by all-trans retinoic acid (tRA) in the MCF-7 breast cancer cell line. tRA (5 x 10(-6) M) increased luciferase activity of the pGL3 promoter vector (containing luc+) up to approximately 3.8-fold in MCF-7 cells, but not in LNCaP prostate cancer cells or JEG-3 choriocarcinoma cells. Chimeric plasmids were constructed and showed that tRA-induction required the luc+ gene, but not any specific promoter or vector sequence. Time course and dose-response studies of tRA-induction indicated that longer treatment (> 24h) and higher tRA dose (> 10(-6) M) were required for luc+ induction compared with those for a positive retinoic acid response element (maximum induction at 6 h and 10(-8) M tRA). Studies with the translation inhibitor, cycloheximide, indicated the half-life of the luc+ protein was increased from 9.7 +/- 1.5 to 22.1 +/- 3.1 h with tRA treatment. Other retinoids, TTNPB, a retinoic acid receptor beta/gamma-specific ligand, and a retinoid X receptor ligand, did not significantly increase luc+ expression. Caution is needed in analysis of retinoid responsive gene regulation with the luciferase reporter system in MCF-7 cells, especially at high retinoid concentrations.
Kogai
Takahiko
T
Molecular Endocrinology Laboratory, VA Greater Los Angeles Healthcare System, Department of Medicine, UCLA School of Medicine, Los Angeles, CA 90073, USA.
Kanamoto
Yoko
Y
Brent
Gregory A
GA
D000818:Animals; D001943:Breast Neoplasms / Q000235:genetics*; D001517:Coleoptera; D004305:Dose-Response Relationship, Drug; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D017930:Genes, Reporter / Q000187:drug effects* / Q000235:genetics*; D005822:Genetic Vectors; D006801:Humans; D008156:Luciferases / Q000187:drug effects* / Q000235:genetics*; D014212:Tretinoin / Q000494:pharmacology* / Q000502:physiology; D014407:Tumor Cells, Cultured
D014212:Tretinoin; D008156:Luciferases