A new approach for the control of Aujeszky's disease by using germ-line transformation

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Intracellular immunization using dominant-negative mutants of viral proteins is prorposed as a new approach to antiviral therapy in humans and to germ-line transformation in animals to confer resistance to viral infections. To obtain effective repressors of Aujeszky's disease virus (ADV) immediate-early (IE) gene expression, mutant genes encoding dominant-negative mutants of ADV IE protein (IE180) and early protein 0 (EPO), and a chimeric gene encoding a fusion protein consisting of the DNA-binding domain of IE180 and a tail-trucated herpes simplex virus 1 VP16 lacking the transcription activation domain were constructed. These gene products inhibited transcription from the ADV IE promoter in transient expression assays. HeLa cell lines stably transformed with the genes showed resistance to ADV infection. Among them, resistance of a cell line transformed with the chimeric transgene was remarkable.In order to assess the antiviral potential of the fusion protein in vivo, the chimeric gene under the control of polypepetide chain elongation factor 1alpha or the mouse Mx1 promoter was used for generation of transgenic mice. C57BL/6 zygotes were microinjected with about 1000 copies of the DNA fragment encoding the chimeric gene. Of the resulting 41 births, five animals had the transgene as determined by Southern blot analysis of tail DNA.Four of the founders failed to transmit the transgene to their progeny. A remaining line transmitted the transgene to F1 progeny. In the transgenic mice, the fusion protein is expressed under the control of the Mx1 promoter which is inducible by double stranded RNA,interferon, or virus infection. To assess the resistance of the established transgenic mice to ADV challenge, breeding of the F1 mice is now in progress.