The inducible lac operator-repressor system is functional for control of expression of injected DNA in Xenopus oocytes.
We have investigated the use of the Escherichia coli lac operator-repressor system to regulate the expression of genes introduced by microinjection into Xenopus laevis oocytes. We observe that expression of an MSV-cat fusion gene, in which the lac operator was inserted between the TATA box and the transcription start point (tsp), or between the tsp and the start codon (ATG), is completely repressed when the lac repressor protein is added to the plasmid suspension prior to injection. The lac repressor had no detectable effect on the expression of a coinjected HSV-1 tk gene that had no operator insertion (or on an MSV-cat gene without an operator), indicating that the nonspecific DNA-binding properties of the repressor do not inhibit transcription. CAT activity expressed from the operator-containing MSV-cat genes transcribed in the oocyte nucleus was also inhibited by repressor injected into the oocyte cytoplasm, showing that biologically active repressor proteins can enter the nucleus from the cytoplasm. Injection of the inducer IPTG into the oocyte cytoplasm markedly derepressed the repressed cat genes but not the HSV-1 tk gene coinjected as an internal control. Overall, our results show that the lac operator-repressor system can be useful as a genetic switch in the regulation of gene expression of injected DNA in frog oocytes. Finally, our observations on the vectors used in this work show that the MSV enhancer significantly activates transcription from the SV40 early promoter in frog oocytes, although previous studies have indicated that the MSV enhancer is not necessary for the activity of the MSV promoter in oocytes [Graves et al., Mol. Cell. Biol. 5 (1985) 1945-1958].
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Church Chemical Laboratories, California Institute of Technology, Pasadena 91125.
Davidson
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D000818:Animals; D004742:Enhancer Elements, Genetic; D004926:Escherichia coli / Q000235:genetics*; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D005798:Genes, Bacterial; D005814:Genes, Viral; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D008845:Microinjections; D008980:Moloney murine sarcoma virus / Q000235:genetics; D009865:Oocytes / Q000378:metabolism; D009875:Operator Regions, Genetic; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis*; D012097:Repressor Proteins / Q000235:genetics / Q000502:physiology*; D013539:Simian virus 40 / Q000235:genetics; D014157:Transcription Factors / Q000502:physiology*; D014982:Xenopus laevis
D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D007544:Isopropyl Thiogalactoside