Expression of immunogenic Puumala virus nucleocapsid protein in transgenic tobacco and potato plants.

Abstract

Transgenic plants, expressing recombinant proteins, are suitable alternatives for the production of relevant immunogens. In the present study, the expression of Puumala virus nucleocapsid protein in tobacco and potato plants (Nicotiana tabacum and Solanum tuberosum) and its immunogenicity was investigated. After infection of leaf discs of SR1 tobacco and tuber discs of potato cv. "Desiree" with the Agrobacterium strain LBA4404 (pAL4404, pBinAR-PUU-S) containing the 1302 bp cDNA sequence of S-RNA segment of a Puumala virus, transgenic tobacco and potato plants expressed the Puumala virus nucleocapsid protein under control of the cauliflower 35S promoter. The recombinant proteins were found to be identical to the authentic Puumala virus nucleocapsid protein as analyzed by immunoblotting. Expression of the nucleocapsid protein was investigated over four plant generations (P to F4) and found to be stable (1 ng/3 microg dried leaf tissue). Transgenic tobacco plants were smaller compared to controls. The transformed potato plants were morphologically similar to control plants and produced tubers as the control potatoes. The S-antigen was expressed at a level of 1 ng protein/5 microg and 1 ng protein/4 microg dried leaf and root tissues, respectively, and remained stable in the first generation of vegetatively propagated potato plants. The immunogenicity of the Puumala virus nucleocapsid protein expressed in Nicotiana tabacum and Solanum tuberosum was investigated in New Zealand white rabbits. They were immunized with leaf extracts from transgenic tobacco and potato plants, and the serum recognized Puumala virus nucleocapsid protein. Transgenic plants expressing hantaviral proteins can thus be used for the development of cost-effective diagnostic systems and for alternative vaccination strategies.