Structural analysis of late intermediate complex formed between plasmid ColIb-P9 Inc RNA and its target RNA - How does a single antisense RNA represstranslation of two genes at different rates?

The antisense Inc RNA encoded by the IncI alpha Colib-P9 plasmid replicon c ontrols the translation of repZ encoding the replication initiator and its leader peptide repY at different rates with different mechanisms. The initi al loop-loop base pairing between Inc RNA and the target in the repZ mRNA l eader inhibits formation of a pseudoknot required for repZ translation. A s ubsequent base pairing at the 5' leader of Inc RNA blocks repY translation. To delineate the molecular basis for the differential control, we analyzed the intermediate complexes formed between RepZ mRNA and Inc RNA(54), a 5'- truncated Inc RNA derivative. We found that the initial base pairing at the loops transforms into a more stable intermediate complex by its propagatio n in both directions. The resulting extensive base pairing indicates that t he inhibition of the pseudoknot formation is established at this stage. Fur thermore, the region of extensive base pairing includes bases different in related plasmids showing different incompatibility. Thus, the observed exte nsive base pairing is important for determining the incompatibility of the low-copy-number plasmids. We discuss the evolution of replication control s ystems found in IncI alpha, IncB, and IncFII group plasmids.