title: Cadmium stimulates transcription of the cytochrome p450 side chain cleavage gene in genetically modified stable porcine granulosa cells.
abstract: We investigated the effects of cadmium (Cd2+) on transcription of the cytochrome p450 side chain cleavage (p450scc) gene and on progesterone synthesis in stable granulosa cells. We used the stable porcine granulosa cell line, JC-410, genetically modified to express a luciferase genomic construct carrying 2320 base pairs (bp) of the p450scc gene promoter (p450scc-2320-LUC). A construct containing only the luciferase gene, pOLUC, was used as a promoterless control. At 1 microM, cadmium chloride (CdCl2) increased transient expression of p450scc-2320-LUC in JC-410 cells by 2.6-fold after 24-h incubation. A similar pattern of stimulation by CdCl2 was observed in cells transiently transfected with a luciferase genomic construct carrying 100 bp of the p450scc gene promoter p450scc-100-LUC, whereas no stimulation by CdCl2 was observed in cells transfected with pOLUC. At 0.6, 1, and 2 microM, CdCl2 stimulated the activity of the p450scc-2320-LUC promoter in a dose-related fashion by 1.58-, 3.19-, and 2.67-fold, respectively, after 24-h incubation. Northern blot analysis showed that CdCl2 at 0.1, 1, 2, and 3 microM increased p450scc mRNA levels by 3.13-, 1.38-, 1.61-, and 1.57-fold, respectively, after 24-h incubation. After 48-h incubation, CdCl2 at 0.6, 1, and 2 microM further increased p450scc mRNA levels by 3.43-, 2.08-, and 2.4-fold, respectively. At 1, 2, and 3 microM, CdCl2 inhibited progesterone synthesis to 0.48-, 0.38-, and 0.29-fold, respectively. After 48-h incubation, CdCl2 at 0.1 microM stimulated progesterone synthesis by 1.6-fold. We conclude that Cd2+ has a dual action in stable porcine granulosa cells: Low concentrations activate, whereas high concentrations inhibit, expression of the p450scc gene and progesterone synthesis. The stimulatory effect of Cd2+ appears to be mediated via a cis-acting element located 100 bp upstream of the p450scc gene transcription start site.
authors.affiliation: Toxicology Centre, University of Saskatchewan, Saskatchewan, Canada.
authors.forename: Andrea D
authors.initials: AD
authors.lastname: Smida
authors.forename: Ximena P
authors.initials: XP
authors.lastname: Valderrama
authors.forename: Maria C
authors.initials: MC
authors.lastname: Agostini
authors.forename: Michael A
authors.initials: MA
authors.lastname: Furlan
authors.forename: Jorge
authors.initials: J
authors.lastname: Chedrese
mesh_terms: D000818:Animals; D019256:Cadmium Chloride / Q000633:toxicity*; D002460:Cell Line; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000235:genetics*; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D006107:Granulosa Cells / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D008156:Luciferases / Q000235:genetics; D013552:Swine; D014158:Transcription, Genetic / Q000187:drug effects*
chemical_list: D008156:Luciferases; D002786:Cholesterol Side-Chain Cleavage Enzyme; D019256:Cadmium Chloride