Regulated gene expression by glucocorticoids in cultured Virginia pine (Pinus virginiana Mill.) cells.
The effects of six glucocorticoids (dexamethasone, hydrocortisone, 6-methylprednisolone, prednisolone, prednisone, and triamcinolone) on inducible gene expression, based on the chimaeric transcriptional activator GVG and carried by the binary expression vector pINDEX3-m-gfp5-ER, were evaluated in transgenic Virginia pine cell cultures. The concentration that activated GVG transcription factor activity, the level of inducible m-gfp5-ER expression, and the kinetics of inducible m-gfp5-ER expression were determined for each glucocorticoid. Transgenic cells produced green fluorescence upon blue light excitation after treatment with prednisolone, prednisone, 6-methylprednisolone, dexamethasone, triamcinolone, and hydrocortisone. Green fluorescence was observed at 6-12 h after treatment of all six glucocorticoids at concentrations of 1, 3, 5, and 10 mg l(-1). Differential expression of gfp was confirmed by northern blot analysis and by quantitative fluorescence analyses of confocal images taken by a LSM 510 Laser Scanning Microscope. Fresh and dry weight increases of transgenic cell cultures were not affected by all six glucocorticoids at concentrations of 0.1, 0.5, 1, 3, and 5 mg l(-1). It is shown that triamcinolone had the most potent effect on the GVG system. Different glucocorticoids can therefore be used to regulate the GVG transcriptional activator and to induce gene expression in transgenic plant cells, and this property could be useful in establishing an optimum system of transgene regulation.
Tang
Wei
W
Department of Biology, Howell Science Complex, East Carolina University, Greenville, NC 27858-4353, USA. tangw@mail.ecu.edu
Newton
Ronald J
RJ
D002478:Cells, Cultured; D018744:DNA, Plant / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D018506:Gene Expression Regulation, Plant / Q000187:drug effects*; D017930:Genes, Reporter; D005938:Glucocorticoids / Q000494:pharmacology*; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000032:analysis / Q000235:genetics; D028223:Pinus / Q000166:cytology / Q000187:drug effects / Q000235:genetics*; D030821:Plants, Genetically Modified; D010957:Plasmids / Q000187:drug effects / Q000235:genetics; D016133:Polymerase Chain Reaction
D018744:DNA, Plant; D005938:Glucocorticoids; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins