Translation products of RNA from Indian peanut clump virus

SUMMARY When RNA from particles of Indian peanut clump virus (IPCV) was translated using messenger-dependent reticulocyte lysate the largest product had a mol. wt. of about 195000 and the other main products had mol. wt. of 143000 and 24500. The 24500 mol. wt. polypeptide co-migrated with IPCV coat protein and was the only product that reacted with antiserum to IPCV particles. Translation of separated RNA species showed that the 195000 mol. wt. and 143000 mol. wt. , as well as some smaller less abundant products, arose from translation of RNA-1 (mol. wt. 1.9 x 106) whereas RNA-2 (mol. wt. 1.6 x 106) was translated to give only the 24500 mol. wt. product Translation ofRNA ofmol, wt. 0.5 x 106 to 0.9 × 106 gave a 50000 mol. wt. product that was present and sometimes prominent in translation products of unfractionated RNA. A product of about 20000 mol. wt. was made by translation of RNA of about 0.2 x 106 tool. wt. IPCV resembles the viruses with bipartite genomes whose larger RNA species is translated into large polypeptides and whose coat protein gene is on the smaller RNA species. The other translation products may be those of subgenomic messenger, or of satellite, RNA species. Indian peanut clump virus (IPCV) has rod-shaped particles mainly of two lengths (approx. 249 and 184 nm) (Reddy et al., 1983) which contain RNA ofmol, wt. 1.9 x 106 (RNA-1) and 1.6 x 106 (RNA-2) respectively (D. V. R. Reddy, D. J. Robinson & B. D. Harrison, unpublished). Its particles are similar in size, but serologically unrelated, to those of peanut clump virus (PCV) from West Africa (Reddy et al., 1983). Moreover, there is evidence that both viruses are transmitted by Polymyxa graminis (Thouvenel & Fauquet, 1981 a; D. V. R. Reddy, unpublished observations) and that their particle morphology resembles that of soil-borne wheat mosaic (SBWMV) and beet necrotic yellow vein (BNYVV) viruses, both of which are transmitted by Polymyxa spp. (Estes & Brakke, 1966; Thouvenel & Fauquet, 1981 b). In this paper we describe the polypeptides made when IPCV RNA is translated in vitro and compare these results with those of similar studies with other Polymyxa-transmitted viruses. The Ludhiana isolate of IPCV (Reddy et al., 1983) was used throughout. Particles were precipitated from chloroform-clarified sap of systemically infected Nicotiana clevelandii plants by adding NaC1 to 0.2 M and polyethylene glycol 6000 to 4~, resuspended and further purified by ultracentrifugation through a sucrose cushion. In most experiments particles were also centrifuged in 10 to 40~ (w/v) sucrose density gradients prepared in 0.2 M-urea, 30 mMKH2PO,, 20 mM-NazB4OT. RNA was extracted from particles suspended in 0.06 M-NaC1, 0.01 i-Tris-HC1, 3 mM-EDTA, pH 8.6, by adding SDS to 0.5~, incubating for 15 min at about 20 °C and extracting protein with phenol :m-cresol (9:1, v/v) containing 0.1 ~ 8-hydroxyquinoline. RNA was then recovered by ethanol precipitation and re-precipitated from 0.3 M-potassium acetate. Translation mixtures contained 66 ~ (v/v) reticulocyte lysate (P & S Biochemicals, Liverpool, U.K.), 82 mM-KC1, 0.4 mi-MgC12, 8 mM-creatine phosphate, 0.4 mM-dithiothreitol, 40 ~M each essential amino acid except leucine, 410 ~tCi/ml L-[4,5-3H]leucine (124 Ci/mmol; Amersham)