Further studies on the translation products of tobacco rattle virus RNA In vitro

Abstract

Translation of RNA-1 (MW, 2.5 × 106) of tobacco rattle virus (TRV) strain PRN in wheat germ extracts containing spermidine resulted in the production of many polypeptides with a maximum MW of 170,000 and a clear band of 140,000. Translation of RNA-1 in lysates of rabbit reticulocytes resulted in these two products but little else. PRN-RNA-2 (MW, 1.0 x 106) was translated in both in vitro systems as two polypeptides, coat protein and a protein of MW 31,000 (band 1). Three bands of possible premature termination products were also produced in wheat germ extracts. The identity of coat protein was shown by coelectrophoresis in 10 and 3% polyacrylamide gels, specific aggregation with PRN protein, and coincidence between tryptic peptide fingerprints. Tryptic peptide mapping showed that band 1 was not a precursor for coat protein. There were, however, some peptides in digests of band 1 which coincided with coat protein peptides, which suggests a partial overlap of amino acid sequences. The relative abundance of coat protein and band 1 protein in translation products of PRN-RNA-2 in wheat germ extracts was dependent on the concentration of magnesium ion in the extracts. At concentrations suboptimal for total incorporation, translation resulted mainly in coat protein, whereas, at supraoptimal concentrations of magnesium, band 1 protein predominated in the translation products. We suggest that PRN-RNA-2 contains two open initiation sites. © 1977.