Segmented genome and nucleocapsid of La Crosse virus

Abstract

La Crosses (LAC) virions purified by velocity and equilibrium gradient centrifugation contained 3 single-stranded RNA species. The 3 segments had sedimentation coefficients of 31S, 25S and 12S by sodium dodecyl sulfate-sucrose gradient centrifugation. By comparison with other viral and cellular RNA species, the LAC viral RNAs had MW of 2.9 .times. 106, 1.8 .times. 106 and 0.4 .times. 106. Phenol-sodium dodecyl sulfate-extracted LAC virion RNA was not infectious for BHK[baby hamster kidney]-21 cell cultures under conditions in which Sindbis viral RNA was infectious. Treatment of LAC virus with the nonionic detergent Triton X-100 and salt released 3 nucleocapsid structures, each containing 1 species of virion RNA. The nucleocapsids had sedimentation coefficients of 115S, 90S and 65S. Negative-contrast electron microscopy of the nucleocapsids indicated that they were convoluted, supercoiled and apparently circular. They had a mean diameter of 10-12 nm and modal lengths of 200, 510 and 700 nm (some were even longer). By chemical and enzymatic analysis of purified viral RNA, 1 type of 5'' nucleotide (pppAp) present in the proportion of 1/RNA segment was identified. After periodate oxidation, each virion RNA species was labeled by reduction with [3H]sodium borohydride. Although the nucleocapsids appear as closed loops, the viral RNA apparently has free 5'' and 3'' ends and is, therefore, not circular.