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Epigenetic silencing and tissue independent expression of a novel tetracycline inducible system in double-transgenic pigs

Wilfried A. Kues^*, Reinhard Schwinzer, Dagmar Wirth, Els Verhoeyen^,1, Erika Lemme^*, Doris Herrmann^*, Brigitte Barg-Kues^*, Hansjörg Hauser, Kurt Wonigeit^,2 and Heiner Niemann^*,2

^* Department of Biotechnology, Institute for Animal Breeding (FAL), Mariensee, Germany;

Klinik für Viszeral- und Transplantationschirurgie, Medizinische Hochschule Hannover, Hannover, Germany; and

Molecular Biotechnology, Gesellschaft für Biotechnologische Forschung mbH, Braunschweig, Germany

²Correspondence: K. W., Klinik f. Viszeral-und Transplantations Clinique, Medizinische Hochschule Hannover, Carl-Neuberg Str. 1, Hannover D-30623, Germany. E-mail: wonigeit.kurt{at}mh-hannover.de H. N., Institut for Animal Breeding (FAL) Department of Biotechnology Höltystr. 10, Neustadt D-31535, Germany. E-mail: niemann{at}tzv.fal.de

ABSTRACT

The applicability of tightly regulated transgenesis in domesticated animals is severely hampered by the present lack of knowledge of regulatory mechanisms and the long generation intervals. To capitalize on the tightly controlled expression of mammalian genes made possible by using prokaryotic control elements, we have used a single-step transduction to introduce an autoregulative tetracycline-responsive bicistronic expression cassette (NTA) into transgenic pigs. Transgenic pigs carrying one NTA cassette showed a mosaic transgene expression restricted to single muscle fibers. In contrast, crossbred animals carrying two NTA cassettes with different transgenes, revealed a broad tissue-independent and tightly regulated expression of one cassette, but not of the other one. The expression pattern correlated inversely with the methylation status of the NTA transcription start sites indicating epigenetic silencing of one NTA cassette. This first approach on tetracycline regulated transgene expression in farm animals will be valuable for developing precisely controlled expression systems for transgenes in large animals relevant for biomedical and agricultural biotechnology.—Kues, W.A., Schwinzer, R., Wirth, D., Verhoeyen, E., Lemme, E., Hermann, D., Barg-Kues, B., Hauser, H., Wonigeit, K., and Niemann, H. Epigenetic silencing and tissue independent expression of a novel tetracycline inducible system in double-transgenic pigs.

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