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The hok/sok, srnB and pnd systems of plasmids RI, F and R438 mediate plasmid maintenance by killing plasmid-free segregants. The systems encode exceptionally stable full-length mRNAs that code for potent cell toxins that kill the cells from within. The systems also produce truncated mRNAs whose appearance is correlated with killing activity. The truncated mRNAs are shortened by 35 to 70 nucleotides in the 3′ ends, but have the same 5′ ends as the full-length transcripts. Translation of the stable killer mRNAs is regulated by unstable antisense RNAs that are complementary to the leader regions of the full-length and truncated mRNAs. We show here, that both the presence of the antisense RNA and of the host enzyme RNase III is required for rapid cleavage of the truncated mRNAs, and we map the cleavage point in the Hok mRNA in vitro and in vivo to be located between nucleotides +245 and +246. The …