The FinOP repressor system of plasmid R1: analysis of the antisense RNA control of traJ expression and conjugative DNA transfer.
A key determinant of the frequency of IncF plasmid-mediated DNA transfer between enterobacterial cells is the FinOP system. traJ, a positive regulator of the transfer (tra) genes is controlled at the post-transcriptional level by two negative elements, finP and finO. FinP is a plasmid-specific antisense RNA, whereas finO encodes a proteic co-repressor which is not plasmid specific but exchangeable among F-like plasmids. We designed a traJ-lacZ test system that allowed us to monitor the effects of FinP and various FinP mutants on traJ expression. Furthermore, the introduction of finO into the test system enabled us to assess the function of FinO in the interaction of FinP with its target, the traJ mRNA. In this test system, FinP, expressed from a single-copy plasmid, in the absence of FinO, repressed traJ expression six-fold. When expressed from a pBR322-derived multicopy plasmid FinP repressed traJ expression approx. 2000-fold. This result unambiguously demonstrated that FinP is sufficient to repress traJ expression in a gene dosage-dependent manner. Mutations of finP creating base exchanges either in loop I or loop II of the two stem-loop structures of the antisense RNA led to a dramatic decrease in the repressor activity. In a combined loop I-loop II mutation the repressor activity was almost completely lost, supporting the model that the first critical interaction between the two RNA molecules occurs via 'kissing' of both loops of the RNAs. Addition of finO to the test system enhanced the repression of traJ expression by FinP by up to two orders of magnitude. This effect of FinO on FinP activity in vivo might indicate that FinO, in addition to its function as an RNA stabilizer, promotes complex formation between the target mRNA and the antisense RNA. Such a function of FinO has recently been shown to exist in vitro (van Biesen and Frost (1994) Mol Microbiol 14: 427-436).
Koraimann
G
G
Institut für Mikrobiologie, Karl-Franzens-Universität Graz, Austria.
Teferle
K
K
Markolin
G
G
Woger
W
W
Högenauer
G
G
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics*; D003227:Conjugation, Genetic / Q000235:genetics; D004274:DNA, Recombinant; D029968:Escherichia coli Proteins; D018628:Gene Dosage; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics*; D009690:Nucleic Acid Conformation; D017354:Point Mutation; D011815:R Factors / Q000235:genetics*; D016372:RNA, Antisense / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D016601:RNA-Binding Proteins; D012097:Repressor Proteins / Q000235:genetics; D017384:Sequence Deletion
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004274:DNA, Recombinant; D029968:Escherichia coli Proteins; D016372:RNA, Antisense; D012329:RNA, Bacterial; D016601:RNA-Binding Proteins; D012097:Repressor Proteins; C488999:TraJ protein, E coli; C041427:TraJ protein, bacteria; C074275:FinO protein, E coli