Role of the coat Protein-RNA interaction in the life cycle of bacteriophage MS2

Abstract

The coat protein of the RNA bacteriophage MS2 interacts with viral RNA to translationally repress replicase synthesis. This protein-RNA interaction is also thought to play a role in genome encapsidation. In this study the strength of the interaction was perturbed by constructing a recombinant genome containing a super-repressing coat mutation. Because replicase synthesis is prematurely repressed, the mutant produces plaques about five orders of magnitude less efficiently than wild-type. The few plaques obtained are second-site revertants of the original coat mutation and fall into two categories. Those of the first type contain nucleotide substitutions within the translational operator that reduce or destroy its ability to bind coat protein, showing that this interaction is not necessary for genome encapsidation. Revertants of the second type are double mutants in which one substitution converts the coat initiator AUG to AUA and the other substitutes an A for the G normally present two nucleotides upstream of the coat start codon. The mutation of the coat protein gene AUG to AUA, by itself, reduces coat protein synthesis to a few percent of the wild-type level. The second substitution destabilizes the coat initiator stem-loop and restores coat protein synthesis to within a few fold of wild-type levels.