The presence of conjugative transposon Tn916 in the recipient strain does not impede transfer of a second copy of the element

Norgren, Mari; Scott, June R.

doi:10.1128/jb.173.1.319-324.1991

Cited by 48 publications

(25 citation statements)

References 33 publications

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“…This is consistent with Tn916 reintegration into att1 or death of cells in which Tn916 has excised. We do not favor the first hypothesis because Tn916 can integrate into multiple sites (29,80,81), and we have observed that Tn916 does not have a preference for reintegration into att1 in transconjugants (L. D. Wright and A. D. Grossman, unpublished results).…”

Section: Discussionmentioning

confidence: 91%

Autonomous Replication of the Conjugative Transposon Tn916

2016

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Integrative and conjugative elements (ICEs), also known as conjugative transposons, are self-transferable elements that are widely distributed among bacterial phyla and are important drivers of horizontal gene transfer. Many ICEs carry genes that confer antibiotic resistances to their host cells and are involved in the dissemination of these resistance genes. ICEs reside in host chromosomes but under certain conditions can excise to form a plasmid that is typically the substrate for transfer. A few ICEs are known to undergo autonomous replication following activation. However, it is not clear if autonomous replication is a general property of many ICEs. We found that Tn916, the first conjugative transposon identified, replicates autonomously via a rolling-circle mechanism. Replication of Tn916 was dependent on the relaxase encoded by orf20 of Tn916. The origin of transfer of Tn916, oriT(916), also functioned as an origin of replication. Using immunoprecipitation and mass spectrometry, we found that the relaxase (Orf20) and the two putative helicase processivity factors (Orf22 and Orf23) encoded by Tn916 likely interact in a complex and that the Tn916 relaxase contains a previously unidentified conserved helix-turn-helix domain in its N-terminal region that is required for relaxase function and replication. Lastly, we identified a functional single-strand origin of replication (sso) in Tn916 that we predict primes second-strand synthesis during rolling-circle replication. Together these results add to the emerging data that show that several ICEs replicate via a conserved, rolling-circle mechanism. IMPORTANCE Integrative and conjugative elements (ICEs) drive horizontal gene transfer and the spread of antibiotic resistances in bacteria.ICEs reside integrated in a host genome but can excise to create a plasmid that is the substrate for transfer to other cells. Here we show that Tn916, an ICE with broad host range, undergoes autonomous rolling-circle replication when in the plasmid form. We found that the origin of transfer functions as a double-stranded origin of replication and identified a single-stranded origin of replication. It was long thought that ICEs do not undergo autonomous replication. Our work adds to the evidence that ICEs replicate autonomously as part of their normal life cycle and indicates that diverse ICEs use the same replicative mechanism. Integrative and conjugative elements (ICEs), also called conjugative transposons, are mobile genetic elements that encode proteins that mediate transfer of the element from the host cell (donor) to a recipient by conjugation. ICEs often contain additional (cargo) genes that can provide a selective advantage to the host cells (reviewed in references 1 and 2). Most ICEs have been identified based on their cargo genes and the phenotypes conferred. For example, many ICEs carry genes encoding antibiotic resistances. The horizontal dissemination of ICEs and their associated cargo genes is a major driver of bacterial genome plasticity and evolution and the s...

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Section: Discussionmentioning

confidence: 91%

Autonomous Replication of the Conjugative Transposon Tn916

2016

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“…Previous studies with the conjugative transposon Tn916 and IncJ elements (18,22,32) have revealed that these elements do not prevent the transfer of a similar or identical element into a host already harboring such an element. Similar to this, our work showed that the presence of either R391 or SXT in a recipient cell had minimal to no detectable effect on the capability of the cell to serve as a recipient for the other element.…”

Section: Discussionmentioning

confidence: 99%

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confidence: 99%

“…Unlike the case for phages and plasmids, relatively little is known about whether similar conjugative transposons can coexist within the same host cell. The well-studied conjugative transposon Tn916, which does not integrate site specifically, can be present in more than one copy at different sites in the host cell chromosome, and the presence of Tn916 in a recipient cell does not inhibit acquisition of a second copy (18,26).We previously identified a conjugative transposon-like element, the SXT element, encoding resistance to sulfamethoxazole (Su r ), trimethoprim (Tm r ), chloramphenicol (Cm r ), and streptomycin (Sm r ) in the recently emerged O139 serogroup of Vibrio cholerae (30). The SXT element (henceforth referred to as SXT), or very closely related elements, have subsequently been detected in all O1 and O139 V. cholerae clinical isolates from the Indian subcontinent (11).…”

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confidence: 99%

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Formation of Chromosomal Tandem Arrays of the SXT Element and R391, Two Conjugative Chromosomally Integrating Elements That Share an Attachment Site

et al. 2001

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The SXT element, a conjugative, self-transmissible, integrating element (a constin) originally derived from a Vibrio cholerae O139 isolate from India, and IncJ element R391, originally derived from a South African Providencia rettgeri isolate, were found to be genetically and functionally related. Both of these constins integrate site specifically into the Escherichia coli chromosome at an identical attachment site within the 5 end of prfC. They encode nearly identical integrases, which are required for chromosomal integration, excision, and extrachromosomal circularization of these elements, and they have similar tra genes. Therefore, these closely related constins have virtually identical mechanisms for chromosomal integration and dissemination. The presence of either element in a recipient cell did not significantly reduce its ability to acquire the other element, indicating that R391 and SXT do not encode surface exclusion determinants. In cells harboring both elements, SXT and R391 were integrated in tandem fashion on the chromosome, and homologous recombination appeared to play little or no role in the formation of these arrays. Interference between R391 and SXT was detected by measuring the frequency of loss of an unselected resident element upon introduction of a second selected element. In these assays, R391 was found to have a stronger effect on SXT stability than vice versa. The level of expression and/or activity of the donor and recipient integrases may play a role in the interference between these two related constins.Similar genetic elements tend not to coexist within the same host cell. Instead, related elements of the same class usually "repel" one another in some fashion. For different types of genetic elements, the molecular bases of incompatibility differ. Plasmid incompatibility, for example, is generally mediated by competition for replication and/or partitioning systems (19). Conjugative plasmids also frequently inhibit host cell entry of related plasmids by altering the host cell's surface (1). Similarly, some bacteriophages alter the surface of host cells to exclude other phages. Phages can also prevent the replication of similar phages through immunity mechanisms (25). Thus, both plasmid and phage incompatibility can depend either on preventing entry of new DNA into a potential host or on inhibiting the replication of new DNA after it has breached the host cell barrier.Chromosomally integrating mobile genetic elements that are transferred between cells via conjugation-often referred to as conjugative transposons-have been found with increasing frequency in both gram-negative and gram-positive bacteria. Unlike the case for phages and plasmids, relatively little is known about whether similar conjugative transposons can coexist within the same host cell. The well-studied conjugative transposon Tn916, which does not integrate site specifically, can be present in more than one copy at different sites in the host cell chromosome, and the presence of Tn916 in a recipient cell does not inhibit...

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“…Furthermore, the presence of a conjugative transposon in the recipient strain in mating does not affect the frequency of integration of a second such element (8,16). These properties make conjugative transposons valuable for mutagenesis, especially in gram-positive bacteria.…”

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Conjugative transposition of Tn916: the transposon int gene is required only in the donor

1992

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Conjugative transposition of transposon Tn916 has been shown to proceed by excision of the transposon in the donor strain and insertion of this element in the recipient. This process requires the product of the transposon int gene. We report here the surprising finding that the int gene is required only in the donor during conjugative transposition. We find that Tn916 int-l, whose int gene has been inactivated by an insertion mutation, transposes when a complementing wild-type int gene is present only in the donor during mating. When the int+ gene is present in a plasmid and is expressed from the spac promoter, conjugative transposition is very inefficient. However, when the Int+ function is supplied from a coresident distantly linked Tn916 tra-641 mutant, which is defective in a function required for conjugation, efficient conjugative transposition of Tn916 int-i occurs. This suggests either that Int is not required for integration of Tn916 in gram-positive bacteria or that the protein is transferred from the donor to the transconjugant during the mating event. When the nonconjugative plasmid pAT145 was present in the donor, it was rarely cotransferred with Tn916. This suggests that complete fusion of mating cells is not common during conjugative transposition.Conjugative transposons promote bacterial conjugation, during which they excise from a donor and then insert into a new target in the recipient genome (recent reviews include references 6, 21, 22, and 29). All such transposons share regions of DNA homology and carry the tetM gene, a determinant of tetracyline resistance which is expressed in bacteria of many different genera (6). The promiscuity of conjugative transposons is an important clinical problem, since they are transferred among unrelated gram-positive bacteria and serve as vectors for the spread of antibiotic resistance. Although these elements do not promote conjugation between Escherichia coli strains (7, 11), they do transpose in this organism. These elements integrate into many different sites in the genome of their hosts, and multiple transposons are often found in transconjugants (10). Furthermore, the presence of a conjugative transposon in the recipient strain in mating does not affect the frequency of integration of a second such element (8, 16). These properties make conjugative transposons valuable for mutagenesis, especially in gram-positive bacteria.The best-studied conjugative transposons are the 16.4-kb Tn916 (9) and TnI545, a 25.3-kb transposon. In addition to tetM, the latter also carries antibiotic resistance determinants for erythromycin and kanamycin (3). The terminal 250 bp of these two elements are essentially identical, and they have similar restriction maps.

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The presence of conjugative transposon Tn916 in the recipient strain does not impede transfer of a second copy of the element

Cited by 48 publications

References 33 publications

Autonomous Replication of the Conjugative Transposon Tn916

Autonomous Replication of the Conjugative Transposon Tn916

Formation of Chromosomal Tandem Arrays of the SXT Element and R391, Two Conjugative Chromosomally Integrating Elements That Share an Attachment Site

Conjugative transposition of Tn916: the transposon int gene is required only in the donor

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