Polyadenylation can regulate ColE1 type plasmid copy number independently of any effect on RNAI decay by decreasing the interaction of antisense RNAI with its RNAII target.
Replication of ColE1-type plasmids is regulated by RNAI, an antisense RNA that interacts with the replication pre-primer, RNAII. Exonucleolytic attack at the 3' end of RNAI is impeded in pcnB mutant bacteria, which lack poly(A) polymerase I-the principal RNA polyadenylase of E. coli; this leads to accumulation of an RNAI decay intermediate (RNAI(-5)) and dramatic reduction of the plasmid copy number. Here, we report that polyadenylation can also affect RNAI-mediated control of plasmid DNA replication by inhibiting interaction of RNAI(-5) with RNAII. We show that mutation of the host pcnB gene profoundly affects the plasmid copy number, even under experimental conditions that limit the effects of polyadenylation on RNAI(-5) decay. Moreover, poly(A) tails interfere with RNAI/RNAII interaction in vitro without producing any detectable alteration of RNAI secondary structure. Our results establish the existence of a previously undetected mechanism by which RNA polyadenylation can control plasmid copy number.
Department of Genetics, Stanford University, Stanford, CA 94305-5120, USA.
Feng-Feng
FF
Xu
Carina
C
Gaggero
Stanley N
SN
Cohen
D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000235:genetics; D011061:Poly A / Q000235:genetics*; D016133:Polymerase Chain Reaction; D016372:RNA, Antisense / Q000235:genetics* / Q000378:metabolism; D012329:RNA, Bacterial / Q000235:genetics; D034741:RNA, Small Interfering
D001426:Bacterial Proteins; D017931:DNA Primers; D016372:RNA, Antisense; D012329:RNA, Bacterial; D034741:RNA, Small Interfering; D011061:Poly A