Genes involved in production of plasmidlike forms by a Bacteroides conjugal chromosomal element share amino acid homology with two-component regulatory systems

Stevens, Ann M.; Sanders, John M.; Shoemaker, Nadja B.; Salyers, Abigail A.

doi:10.1128/jb.174.9.2935-2942.1992

Cited by 80 publications

(98 citation statements)

References 32 publications

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“…eutrophus AcoR (Kruger and Steinbuchel, 1992); Pseudomonas syringae pv. phaseolicola (Psy) HrpS (Grimm and Panopoulos, 1989); Xanthobacter autotrophicus GJ10 (Xau) DhlR (van der Ploeg and Janssen, 1995); Vibrio parahaemolyticus (Vpa) FlaK (Stewart and McCarter, 1996); Bacillus subtilis (Bsu) LevR (Debarbouille et al, 1991); B. subtilis RocR (Calogero et al, 1994) and Bacteroides thetaiotaomicon (Bth) RteB (Stevens et al, 1992). Two NtrC and NifA proteins are included in this alignment.…”

Section: Introductionmentioning

confidence: 99%

A conserved region in the σ⁵⁴‐dependent activator DctD is involved in both binding to RNA polymerase and coupling ATP hydrolysis to activation

Wang

Lee

Brewer

et al. 1997

Molecular Microbiology

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SummaryRhizobium melioti DctD activates transcription from the dctA promoter by catalysing the isomerization of closed complexes between 54 -RNA polymerase holoenzyme and the promoter to open complexes. DctD must make productive contact with 54 -holoenzyme and hydrolyse ATP to catalyse this isomerization. To define further the activation process, we sought to isolate mutants of DctD that had reduced affinities for 54 -holoenzyme. Mutagenesis was confined to the well-conserved C3 region of the protein, which is required for coupling ATP hydrolysis to open complex formation in 54 -dependent activators. Mutant forms of DctD that failed to activate transcription and had substitutions in the C-terminal half of the C3 region were efficiently cross-linked to 54 and the ␤-subunit of RNA polymerase, suggesting that they bound normally to 54 -holoenzyme. In contrast, some mutant forms of DctD with amino acid substitutions in the N-terminal half of the C3 region had reduced affinities for 54 and the ␤-subunit in the cross-linking assay. These data suggest that the N-terminal half of the C3 region of DctD contains a site that may contact 54-holoenzyme during open complex formation.

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Section: Introductionmentioning

confidence: 99%

A conserved region in the σ⁵⁴‐dependent activator DctD is involved in both binding to RNA polymerase and coupling ATP hydrolysis to activation

Wang

Lee

Brewer

et al. 1997

Molecular Microbiology

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“…Previously, we had shown that functions provided by the Tcr elements were needed for the excision and circularization of the NBUs (12,14,15). One model that followed from these observations was that the excised circular forms were transfer intermediates.…”

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“…First, the copy number of the NBUs was estimated to be less than one per cell. Second, tetracycline induction of functions on a coresident Tcr element was needed to produce the circle forms, and the circle forms disappeared when tetracycline was removed (14,15). However, it is still possible that the apparent low copy number was due to an artifact of the procedure used to isolate the DNA (12,15).…”

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confidence: 99%

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Excision, transfer, and integration of NBU1, a mobilizable site-selective insertion element

et al. 1993

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The Bacteroides species harbor a family of conjugative transposons called tetracycline resistance elements (Tcr elements) that transfer themselves from the chromosome of a donor to the chromosome of a recipient, mobilize coresident plasmids, and also mediate the excision and circularization of members of a family of 10-to 12-kbp insertion elements which share a small region of DNA homology and are called NBUs (for nonreplicating Bacteroides units). The NBUs are sometimes cotransferred with Tcr elements, and it was postulated previously that the excised circular forms of the NBUs were plasmidlike forms and were transferred like plasmids and then integrated into the recipient chromosome. We used chimeric plasmids containing one of the NBUs, NBU1, and a Bacteroides-Escherichia coli shuttle vector to show that this hypothesis is probably correct. NBU1 contained a region that allowed mobilization by both the Tcr elements and IncP plasmids, and we used these conjugal elements to allow us to estimate the frequencies of excision, mobilization, and integration of NBU1 in Bacteroides hosts to be approximately 10-2, 10-5 to 10-4, and 10-2, respectively. Although functions on the Tcr elements were required for the excision-circularization and mobilization of NBU1, no Tcr element functions were required for integration into the recipient chromosome. Analysis of the DNA sequences at the integration region of the circular form of NBU1, the primary insertion site in the Bacteroides thetaiotaomicron 5482 chromosome, and the resultant NBUl-chromosome junctions showed that NBU1 appeared to integrate into the primary insertion site by recombining within an identical 14-bp sequence present on both NBU1 and the target, thus leaving a copy of the 14-bp sequence at both junctions. The apparent integration mechanism and the target selection of NBU1 were different from those of both XBU4422, the only member of the conjugal Tcr elements for which these sequences are known, and Tn4399, a mobilizable Bacteroides transposon. The NBUs appear to be a distinct type of mobilizable insertion element.Large (>70-kbp) conjugative transposons have been found in many clinical isolates of Bacteroides species from the human colon. These conjugative transposons have been called Tcr elements because most of them carry a tetracycline resistance (Tcr) gene, tetQ, but some members of this family of conjugative transposons are cryptic (TcS), e.g., XBU4422 (13). NBUs were originally detected as 10-to 12-kbp covalently closed circles (plasmidlike forms) in a Bacteroides uniformis 0061 derivative (BU1004) which carried a Tcr element. The circular forms of the NBUs were seen only when the strain was grown in medium containing tetracycline. Subsequently, we found that the NBUs are normally integrated in the chromosome and that trans action by a Tcr element is required to excise and circularize them (12, 15).The circular forms of the NBUs appeared not to replicate and were designated nonreplicating Bacteroides units to distinguish them from plasmids. Two lines of e...

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“…They act in trans to excise and mobilize smaller unlinked DNA elements called NBUs (nonreplicating Bacteroides units) ( Fig. 1) (1,12,13,16,17). Two NBUs, NBU1 and NBU2, have been characterized (12, 13).…”

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The mobilization regions of two integrated Bacteroides elements, NBU1 and NBU2, have only a single mobilization protein and may be on a cassette

Shoemaker

Wang

et al. 1995

J Bacteriol

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Bacteroides conjugative transposons can act in trans to excise, circularize, and transfer unlinked integrated elements called NBUs (for nonreplicating Bacteroides units). Previously, we localized and sequenced the mobilization region of one NBU, NBU1, and showed that this mobilization region was recognized by the IncP plasmids RP4 and R751, as well as by the Bacteroides conjugative transposons. We report here that the single mobilization protein carried by NBU1 appears to be a bifunctional protein that binds to the oriT region and catalyzes the nicking reaction that initiates the transfer process. We have also localized and sequenced the mobilization region of a second NBU, NBU2. The NBU2 mobilization region was 86 to 90% identical at the DNA sequence to the oriT-mob region of NBU1. The high sequence similarity between NBU1 and NBU2 ended abruptly after the stop codon of the mob gene and about 1 kbp upstream of the oriT region, indicating that the oriT-mob regions of NBU1 and NBU2 may be on some sort of cassette. A region on NBU1 and NBU2 which lies immediately upstream of the oriT region had 66% sequence identity to a region upstream of the oriT region on a mobilizable transposon, Tn4399, an element that had previously appeared to be completely unrelated to the NBUs.Many Bacteroides clinical isolates carry large (Ͼ70-kbp) integrated self-transmissible elements, called conjugative transposons (2, 11). These elements have also been called Tc r elements because most of them carry a tetracycline resistance (Tc r ) gene, tetQ. Bacteroides Tc r elements have an unusual activity not reported for any other known conjugative transposon. They act in trans to excise and mobilize smaller unlinked DNA elements called NBUs (nonreplicating Bacteroides units) ( Fig. 1) (1,12,13,16,17). Two NBUs, NBU1 and NBU2, have been characterized (12, 13). These NBUs are 10 to 12 kbp in size and appear to be unrelated to the conjugative transposons that excise and mobilize them (12). During transfer, the NBUs are excised from the chromosome to form a double-stranded circle, which is the transfer intermediate ( Fig. 1) (7, 13). The circular form of the NBU then uses the transfer pore provided by the conjugative transposon to move into a recipient cell (7). The Bacteroides conjugative transposons also enhance, by at least 1000-fold, the transfer frequency of another integrated element, the mobilizable transposon Tn4399 (5, 8). Results of previous studies had indicated that Tn4399 was not related to the NBUs. For example, Tn4399 has a different mode of integration than that of the NBUs, and the mobilization region of Tn4399 had no significant sequence similarity to the mobilization region of the NBUs (6, 7, 13). In this report, we provide the first evidence that Tn4399 and the NBUs do in fact share some sequence similarity.The oriT-mob region of NBU1 allows NBU1 to be mobilized not only by Bacteroides conjugative transposons but also by the IncP plasmids R751 and RP4 (7). Thus, mobilization of the NBU1 circle form by IncP plasmids presumably ...

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Genes involved in production of plasmidlike forms by a Bacteroides conjugal chromosomal element share amino acid homology with two-component regulatory systems

Cited by 80 publications

References 32 publications

A conserved region in the σ⁵⁴‐dependent activator DctD is involved in both binding to RNA polymerase and coupling ATP hydrolysis to activation

A conserved region in the σ⁵⁴‐dependent activator DctD is involved in both binding to RNA polymerase and coupling ATP hydrolysis to activation

Excision, transfer, and integration of NBU1, a mobilizable site-selective insertion element

The mobilization regions of two integrated Bacteroides elements, NBU1 and NBU2, have only a single mobilization protein and may be on a cassette

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Genes involved in production of plasmidlike forms by a Bacteroides conjugal chromosomal element share amino acid homology with two-component regulatory systems

Cited by 80 publications

References 32 publications

A conserved region in the σ54‐dependent activator DctD is involved in both binding to RNA polymerase and coupling ATP hydrolysis to activation

A conserved region in the σ54‐dependent activator DctD is involved in both binding to RNA polymerase and coupling ATP hydrolysis to activation

Excision, transfer, and integration of NBU1, a mobilizable site-selective insertion element

The mobilization regions of two integrated Bacteroides elements, NBU1 and NBU2, have only a single mobilization protein and may be on a cassette

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A conserved region in the σ⁵⁴‐dependent activator DctD is involved in both binding to RNA polymerase and coupling ATP hydrolysis to activation

A conserved region in the σ⁵⁴‐dependent activator DctD is involved in both binding to RNA polymerase and coupling ATP hydrolysis to activation