PROGRAMMED CELL-DEATH BY HOK SOK OF PLASMID R1 - PROCESSING AT THE HOK MESSENGER-RNA 3'-END TRIGGERS STRUCTURAL REARRANGEMENTS THAT ALLOW TRANSLATION AND ANTISENSE RNA-BINDING/

The hok/sok locus of plasmid R1 mediates plasmid stabilization by kill ing of plasmid-free cells. The locus specifies two RNAs, hok mRNA and Sok antisense RNA. The post-segregational killing mediated by hok/sok is governed by a complicated control mechanism that involves both post -transcriptional inhibition of translation by Sok-RNA and activation o f hok translation by mRNA 3' processing. Sok-RNA inhibits translation of a reading frame (mok) that overlaps with hok, and translation of ho k is coupled to translation of mok. In the inactive full-length hok mR NA, the translational activator element at the mRNA 5'-end (tac) is se questered by the fold-back-inhibitory element located at the mRNA 3'-e nd (fbi). The 5' to 3' pairing locks the RNA in an inert configuration in which the SDmok and Sok-RNA target regions are sequestered. Here w e show that the 3' processing leads to major structural rearrangements in the mRNA 5'-end. The structure of the refolded RNA explains activa tion of translation and antisense RNA binding. The refolded RNA contai ns an antisense RNA target stem-loop that presents the target nucleoti des in a single-stranded conformation. The stem of the target hairpin contains SDmok and AUG(mok) in a paired configuration. Using toeprinti ng analysis, we show that this pairing keeps SDmok in an accessible co nfiguration. Furthermore, a mutational analysis shows that an internal loop in the target stem is prerequisite for efficient translation and antisense RNA binding. (C) 1997 Academic Press Limited.