SINDBIS VECTORS SUPPRESS SECRETION OF SUBVIRAL PARTICLES OF JAPANESE ENCEPHALITIS-VIRUS FROM MAMMALIAN-CELLS INFECTED WITH SIN-JEV RECOMBINANTS

Double-subgenomic Sindbis virus (dsSIN) recombinants that express cass ettes encoding prM-E or a C-terminally truncated form of E of Japanese encephalitis virus (JEV) were constructed. The products were efficien tly expressed in both mammalian and mosquito cell lines infected with the dsSIN recombinants. However, suppression of prM-E secretion from m ammalian cells infected with dsSIN-prM-E recombinants was observed. Th is suppression was more pronounced late in infection (< 5% of total pr oduct was secreted during an 8-hr chase) than early in infection (15% secretion during a 6-hr chase). In comparison, a vaccinia virus-prM-E recombinant (vP829) described previously (E. Konishi at al. (1991) Vir ology 185, 401-410) was shown to secrete 35-50% of total product durin g a 6- to 8-hr chase both early and late in infection. In contrast, se cretion of prM-E from dsSIN-prM-E-infected mosquito (C6/36) cells was found to be efficient (> 50% during an 8-hr chase). The prM-E secreted from both mammalian and mosquito cells was in the form of subviral pa rticles as determined by velocity gradient centrifugation, sensitivity to nonionic detergent, and analysis of processing of N-linked glycans . The truncated E protein expressed by the dsSIN recombinants was secr eted efficiently from both mammalian and mosquito cells. Coinfection e xperiments with the dsSIN-JEV recombinants + wild-type vaccinia virus and vP829 + SIN demonstrated that the reduced level of secretion of su bviral particles exhibited by the dsSIN-JEV recombinants was due to an inhibitory effect of the dsSIN vectors. Furthermore, this inhibitory effect was accounted for by the SIN nonstructural proteins since SIN r eplicons that express prM-E cassette in place of the SIN structural pr otein open reading frame exhibited a low level of subviral particle se cretion. No self-propagating infectious particles were produced in cel ls transfected with SIN replicons that encode the JEV prM-E cassette. The suppression of subviral particle secretion was apparently correlat ed with the inhibition of cell protein synthesis which is mediated in SIN-infected vertebrate cells by expression of the SIN nonstructural p roteins. (C) 1995 Academic Press, Inc.