Rescue of synthetic RNAs into Thogoto and influenza A virus particles using core proteins purified from Thogoto virus

The ribonucleoprotein (RNP) complexes of Thogoto virus (THOV), a tick-borne orthomyxovirus, have been purified from detergent-lysed virions. The purif ied RNPs were then disrupted by centrifugation through a CsC1-glycerol grad ient to obtain fractions highly enriched in nucleoprotein (NP) and virtuall y devoid of viral genomic RNA. When these NP-enriched fractions were incuba ted with a synthetic THOV-like RNA, and the mixtures were transfected into THOV-infected cells, the synthetic RNA was expressed and packaged into THOV particles. Similarly, hybrid mixtures containing purified THOV NP and infl uenza A virus synthetic RNAs (either a model CAT RNA or a gene encoding the viral neuraminidase), were prepared and transfected into influenza A virus -infected cells. The synthetic CAT RNA, was shown to be expressed and packa ged into virus particles, and the neuraminidase gene was rescued into influ enza virions. These data are discussed in terms of the similarities observe d between THOV and influenza A virus and the potential application of the T HOV purified proteins for rescuing synthetic genes into infectious viruses. (C) 2000 Elsevier Science B.V. All rights reserved.