Amber mutants and polarity in vitro

Abstract

Polarity resulting from the translation of amber codons as nonsense in vitro has been studied. The amber codons are located in the coat protein gene of the RNA-containing bacteriophage f2. Extensive translational polarity is observed when an amber codon is located on the RNA at the site coding for the sixth amino acid from the NH 2-terminal of the coat protein ( sus-3), while little or no polarity is observed when an amber codon is located at the site coding for the seventieth amino acid from the NH 2-terminal of the coat protein ( sus-11). When polar coat mutant RNA is translated in Su − extracts, only monosomes are formed, while when wild-type RNA or RNA containing the non-polar coat mutation is translated under these conditions, polysomes as well as monosomes are formed. Since the mutant which shows the polarity effect forms only monosomes in Su − extracts, we conclude that there is only one major starting point for the translation of f2 RNA. These data support the hypothesis that the genes on this RNA are translated sequentially in vitro, and that the translation of the non-coat genes is contingent upon the translation of at least the codon for site 6 of the coat protein gene.