Physicochemical Characterization of Festuca Leaf Streak Virus

Abstract

SUMMARYParticles of festuca leaf streak virus (FLSV) contain three major proteins. One of these [mol. wt. 49000 (49K)] is the main constituent of the nucleocapsid, whereas the other two (mol. wt. 58K and 20K) were released from the nucleocapsid when particles were treated with non-ionic detergent. The 58K protein is glycosylated. The 58K, 49K and 20K proteins correspond to the G, N and M proteins of rhabdoviruses, respectively. Four minor proteins associated with the virus particles have mol. wt. of 189K, l17K, 101K and 41K. The 189K and 101K proteins are associated with the nucleocapsid, whereas the 117K protein was found in the soluble fraction after detergent treatment. Nucleic acid isolated from virus particles is probably RNA with an estimated mol. wt. of 4.3 x 106. The buoyant density of virus particles in sucrose was estimated to be 1.194 g/ml and the s20.~ to be 704S. The present results, together with previous information, make FLSV a definitive member of subgroup A of the plant rhabdovirus group of the family Rhabdoviridae.Festuca gigantea plants with leaf streaking symptoms contain rhabdovirus-like particles, 330 × 61 nm, that accumulate inside membrane-bound cisternae in the cytoplasm (Lundsgaard & Albrechtsen, 1979). The virus, tentatively named festuca leaf streak virus (FLSV), multiplies in cowpea protoplasts (Van Beek et al., 1985) inoculated with isolated virions and is regarded as a possible member of the family Rhabdoviridae (Matthews, 1982). In its morphology and cytopathology, FLSV resembles northern cereal mosaic virus (NCMV) and barley yellow striate mosaic virus (BYSMV), but no serological cross-reactions were detected when the G protein and the nucleocapsid of FLSV were compared with those of NCMV and BYSMV (Lundsgaard, 1984).In this paper, some of the physicochemical properties of FLSV are compared with those of well-characterized rhabdoviruses.FLSV was purified from infected Festuca gigantea plants grown in a glasshouse. Infected plants were propagated by division. About 50 g of infected leaves were triturated in 5 parts (w/v) TMN buffer pH 8.4 (0.1 M-Tris-HCI, 0.01 M-MgCI> 0.04 M-NazSO3) in a Waring blender. The homogenate was filtered through cheesecloth and then three times through a Celite pad. The virus particles were sedimented by centrifugation at 26000 g for 1 h, resuspended in TMN buffer pH 7.5, and separated by rate zonal centrifugation (linear 10 to 30% sucrose gradient in TMN buffer pH 7.5). The layer with virus particles was removed and centrifuged at 36000 g for 1 h. The pellet was resuspended in water and kept at -20 °C until used. After thawing, precipitated green material was removed by low-speed centrifugation. Electron microscopy of purified virus showed that most of the FLSV particles were in two to four pieces which were linked together.The structural proteins of FLSV were analysed by SDS-PAGE in 5% or 10% horizontal slab gels. Gels and samples were prepared according to Weber & Osborn (1969) as outlined in Sigma Technical Bulletin No. MWS-877. The electrophoresed ...

Routing of the G Protein During Maturation of Festuca Leaf Streak Rhabdovirus in its Plant Host

“…Glycosylation of VSV-G has been shown to be essential for routing of this protein to the plasma membrane (Machamer et al, 1985), However, G protein of purified FLSV is glycosylated (Lundsgaard, 1987), excluding that lack of glycosylation is the reason for the retention of FLSV-G in ER.…”