Translation in vitro of carnation mottle virus RNA Regulatory function of the 3′-region

Abstract

Carnation mottle virus (CarMV) RNA was translated in a wheat-germ, cell-free system into three discrete polypeptide chains of 77,000 (CP-I), 38,000 (CP-II) and 30,000 M r (CPIII). The size of the three polypeptide products represents the translation of almost the entire length of the viral RNA. The CP-II in vitro product was identified as the viral coat protein by gel electrophoresis under denaturing conditions, inununoprecipitation with antibodies directed against the disrupted viral particles, and by peptide mapping. The discrete nature of the three proteins was shown by peptide analysis of the proteolytic products, the absence of cross-reaction between antibodies against CarMV-disrupted particles and CP-I or possibly CP-III, and by the order of their appearance during in vitro translation. Limited phosphorolysis of the 3′-terminus of the viral RNA chains by Escherichia coli polynucleotide phosphorylase caused the selective disappearance of the 77,000 protein product, paralleled with a gradual loss of infectivity. These observations might indicate that the viral RNA is polycistronic, and suggest that the structure at the 3′-terminus of the carnation mottle virus RNA may have a regulatory role in the translation of the viral RNA chains.