REGULATION OF PLASMID R1 REPLICATION - PCNB AND RNASE-E EXPEDITE THE DECAY OF THE ANTISENSE RNA, COPA

The replication frequency of plasmid R1 is controlled by an unstable a ntisense RNA, CopA, which, by binding to its complementary target, blo cks translation of the replication rate-limiting protein RepA. Since t he degree of inhibition is directly correlated with the intracellular concentration of CopA, factors affecting CopA turnover can also alter plasmid copy number, We show here that PcnB (PAPI - a poly(A)polymeras e of Escherichia coil) is such a factor. Previous studies have shown t hat the copy number of ColE1 is decreased in pcnB mutant strains becau se the stability of the RNase E processed form of RNAI, the antisense RNA regulator of ColE1 replication, is increased. We find that, analog ously, the twofold reduction in RI copy number caused by a pcnB lesion is associated with a corresponding increase in the stability of the R Nase E-generated 3' cleavage product of CopA. These results suggest th at CopA decay is initiated by RNase E cleavage and that PcnB is involv ed in the subsequent rapid decay of the 3' CopA stem-loop segment, We also find that, as predicted, under conditions in which CopA synthesis is unaffected, pcnB mutation reduces RepA translation and increases C opA stability to the same extent.