Derepression of conjugal transfer of the antibiotic resistance plasmid R100 by antisense RNA

Dempsey, Walter B.

doi:10.1128/jb.171.5.2886-2888.1989

Cited by 8 publications

(8 citation statements)

References 15 publications

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“…Antisense RNA also regulates replication of the IncFII plasmids (Light and Molin, 1983), of plasmid RI 162 (Kim and Meyer, 1986) and of plasmid pT181 (Kumar and Novick, 1985), but in these cases regulation is exerted on the mRNAs of the replication initiation proteins. In addition, antisense RNA is involved in regulation of the hoklsok killing function of plasmid RI (Gerdes et al, 1988) and in control of the conjugal transfer of plasmids of the IncFI and IncFII groups (Dempsey, 1989).…”

Section: Introductionmentioning

confidence: 99%

Control of replication of plasmid R1: structures and sequences of the antisense RNA, CopA, required for its binding to the target RNA, CopT.

1990

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The replication frequency of plasmid R1 is determined by the availability of the RepA protein, which acts at the origin of replication to promote initiation. Synthesis of RepA is negatively regulated both at the transcriptional and post‐transcriptional levels. Post‐transcriptional control is exerted through the action of an antisense RNA, CopA RNA. The target of CopA RNA, CopT RNA, is located in the leader region of the RepA mRNA. Binding between CopA and CopT inhibits repA expression. We have previously presented an in vitro analysis of the binding reaction between CopA and CopT RNAs. In this communication, we extend the in vitro analysis by determining the regions of CopA required for binding, and also demonstrate that binding occurs in at least two steps. The first step is the formation of an initial, transient complex; stem‐loop II is the structure in CopA necessary and sufficient for this step. The subsequent step(s), resulting in the formation of a complete duplex, requires a stretch of single‐stranded nucleotides located 5′ to stem‐loop II in CopA, and its counterpart in CopT. We show that the single‐stranded region can be positioned on either side of stem‐loop II provided that there is a complementary stretch of nucleotides in CopT, indicating that the second step(s) is not sequence‐specific. Furthermore, the effects of salt concentration and temperature on the binding reaction indicate that duplex formation occurs through a mechanism of gradual intra‐strand breaking and inter‐strand formation of hydrogen bonds.

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Section: Introductionmentioning

confidence: 99%

Control of replication of plasmid R1: structures and sequences of the antisense RNA, CopA, required for its binding to the target RNA, CopT.

1990

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“…Previous studies showed that the ratio of sense (traJ ) to antisense (finP ) transcripts was important in conjugal regulation (Dempsey, 1989a;1994b). Another study showed that traM transcripts had the potential to affect that ratio (Dempsey, 1994a).…”

Section: Discussionmentioning

confidence: 99%

“…The normal donor ability of R100 is 0.01-0.02 transconjugants per 100 donors (Dempsey, 1989a;1994b). Co-resident multicopy plasmids carrying finP lower that rate to 1-7 × 10 ¹4 (Dempsey, 1989a;1994b;Fee and Dempsey, 1986). Co-resident multicopy plasmids containing the traJ promoter and the traJ leader region raise the donor ability to 200 transconjugants per 100 donors (Dempsey, 1989a;1994b).…”

Section: R100 (Nr1 R222mentioning

confidence: 99%

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The finM promoter and the traM promoter are the principal promoters of the traM gene of the antibiotic resistance plasmid R100

Stockwell

Dempsey

1997

Molecular Microbiology

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SummaryfinP multicopy repression and traJ multicopy derepression indicate that the ratio of sense to antisense transcripts is important in the regulation of R100 conjugation. The extension of R100 traM transcripts into traJ shows that promoters in traM can affect this ratio, making the regulation of traM transcription important in the regulation of R100 conjugation. Since R100 traM, traY and traI proteins bind to the traM promoter region, we examined traM transcription in R100-1 traM, traY and traI mutants and compared it with traM transcription in both R100-1 and R100. We verified that the traM and finM promoters provide virtually all the transcripts originating in the R100-1 traM gene. When either is deleted, as in VAR22 or VAR30, the remaining promoter is highly active. We show here that traY positively regulates R100-1 traM transcription, as has been found for F. We found that traI did not regulate R100-1 traM transcription. The measured activity of the native R100 traM promoter was 12% of that in R100-1, whereas the native R100 finM promoter was 45% of that in R100-1. These data and data from the R100-1 traY and traI mutants show that the activities of the two promoters varied independently.

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“…Natural antisense RNAs have been shown to exert regulatory effects in bacteriophages T4 (Belin et aI., 1987), P1 and P7 (Citron and Schuster, 1990) and P22 (Liao et al, 1987), to control plasmid replication (Polisky, 1988;Persson et al, 1988;Nordstrom et al, 1988;Blomberg et al, 1990) and conjugation (Dempsey, 1989), and to regulate osmoregulatory genes (Aiba et al, 1987;'Cole and Honore, 1989) and the transposase of IS10 (Case et al, 1990;Ma and Simons, 1990).…”

Section: Antisense Control In Naturementioning

confidence: 98%

Towards Gene-Inhibition Therapy: A Review of Progress and Prospects in the Field of Antiviral Antisense Nucleic Acids and Ribozymes

James

1991

Antivir Chem Chemother

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Antisense RNA and its derivatives may provide the basis for highly selective gene inhibition therapies of virus infections. In this review, I concentrate on advances made in the study of antisense RNA and ribozymes during the last five years and their implications for the development of such therapies. It appears that antisense RNAs synthesized at realistic levels within the cell can be much more effective inhibitors than originally supposed. Looking at those experiments that enable comparisons to be made, it seems that inhibitory antisense RNAs are not those that are complementary to particular sites within mRNAs but those that are able to make stable duplexes with their targets, perhaps by virtue of their secondary structure and length. The inclusion of ribozyme sequences within antisense RNAs confers RNA-cleaving activity upon them in vitro and possibly in cells, thereby offering the possibility of markedly increasing their therapeutic potential. The varieties of natural ribozyme and their adaptation as artificial catalysts are reviewed. The implications of these developments for antiviral therapy are discussed.

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Derepression of conjugal transfer of the antibiotic resistance plasmid R100 by antisense RNA

Abstract: Conjugal transfer of the normally repressed antibiotic resistance plasmid R100 was derepressed by fragments of R100 that carried the traj promoter and the traJ leader but lacked the finP promoter.

Cited by 8 publications

References 15 publications

Control of replication of plasmid R1: structures and sequences of the antisense RNA, CopA, required for its binding to the target RNA, CopT.

Control of replication of plasmid R1: structures and sequences of the antisense RNA, CopA, required for its binding to the target RNA, CopT.

The finM promoter and the traM promoter are the principal promoters of the traM gene of the antibiotic resistance plasmid R100

Towards Gene-Inhibition Therapy: A Review of Progress and Prospects in the Field of Antiviral Antisense Nucleic Acids and Ribozymes

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