Functional and comparative analysis of two distinct ecdysteroid-responsive gene expression constructs in Drosophila S2 cells.
Inducible expression systems have proven to be of major interest when analysing the function of specific genes or when expressing cytotoxic proteins. In an effort to develop inducible switches allowing for flexible fine-tuning of gene expression levels in insect cells, we have compared the induction capacities of two Drosophila minimal promoters when linked to four consecutive ecdysone response elements. These minimal promoters, either containing a TATA-box or a downstream promoter element, drove the expression of a luciferase reporter gene. Potent induction capacities were observed with the insect moulting hormone, 20-hydroxyecdysone, and with ponasterone A, a plant ecdysteroid. The developed inducible switches further expand the repertoire of molecular tools for functional expression of proteins of interest in insect cells. In addition, the combination of an ecdysone switch with promoters that possess different structural elements can provide novel insights into ecdysteroid-induced transcription in an insect cell line.
Poels
Jeroen
J
Laboratory for Developmental Physiology, Genomics and Proteomics, Zoological Institute, Naamsestraat 59, B-3000 Leuven, Belgium.
Martinez
Alberto
A
Suner
Marie-Marthe
MM
De Loof
Arnold
A
Dunbar
Stuart J
SJ
Vanden Broeck
Jozef
J
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D004330:Drosophila / Q000166:cytology / Q000235:genetics*; D029721:Drosophila Proteins; D004441:Ecdysterone / Q000031:analogs & derivatives* / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics*; D017930:Genes, Reporter; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics; D019476:Insect Proteins / Q000096:biosynthesis / Q000235:genetics; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000235:genetics*; D011987:Receptors, Steroid / Q000096:biosynthesis / Q000235:genetics; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection
D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D018840:HSP70 Heat-Shock Proteins; D019476:Insect Proteins; D011987:Receptors, Steroid; D014157:Transcription Factors; C066030:USP protein, Drosophila; C055469:ecdysone receptor; D004441:Ecdysterone; C008999:ponasterone A; D008156:Luciferases