Inducible control of tissue-specific transgene expression in Xenopus tropicalis transgenic lines.
Analysis of gene function in vertebrates is facilitated by gain-of-function studies, such as injection of synthetic mRNA in amphibian embryos. This approach is hampered by lack of spatial and temporal control of expression of the introduced gene product. An additional level of control is obtained by nuclear-transfer-mediated transgenesis, but functional analyses are complicated by variability and background abnormalities in primary transgenic embryos. The GAL4/UAS system permits establishment of stable lines and elimination of nuclear-transfer-associated abnormalities, through generation of separate UAS-'effector' and GAL4 'transactivator' transgenic lines. When the GAL4 DNA-binding domain is combined with a steroid hormone ligand-binding domain, this system allows full temporal regulation of transgene expression by introduction of an exogenous steroid analogue, the progesterone antagonist RU486. We show here that by crossing stable Xenopus tropicalis transgenic lines, one bearing a UAS-enhanced cyan fluorescent protein (ECFP) reporter construct, and the other with a GAL4-progesterone receptor fusion driven by a retina-specific promoter, reporter expression in the resulting embryos can be induced with RU486 in a tissue-specific manner. These results suggest that the inducible binary system, in which the target gene expression can be controlled in a stage- and tissue-specific pattern, should be readily applicable for gene function studies at all stages of development.
Department of Biology, University of Virginia, Charlottesville, VA 22903, USA.
Jeiwook
J
Chae
Lyle B
LB
Zimmerman
Robert M
RM
Grainger
D000818:Animals; D030801:Animals, Genetically Modified; D004268:DNA-Binding Proteins; D005260:Female; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics; D008297:Male; D008845:Microinjections; D015735:Mifepristone / Q000494:pharmacology; D010641:Phenotype; D012333:RNA, Messenger / Q000008:administration & dosage / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D014018:Tissue Distribution; D014157:Transcription Factors / Q000235:genetics; D015533:Transcriptional Activation / Q000187:drug effects; D014981:Xenopus / Q000196:embryology* / Q000235:genetics*; D029867:Xenopus Proteins / Q000235:genetics
D004268:DNA-Binding Proteins; C052130:GAL4 protein, S cerevisiae; D008164:Luminescent Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D029867:Xenopus Proteins; D049452:Green Fluorescent Proteins; D015735:Mifepristone