Stabilization of pSW100 from<i>Pantoea stewartii</i>by the F Conjugation System

Lin, Mei-Hui; Liu, Shih‐Tung

doi:10.1128/jb.00846-07

Cited by 5 publications

(9 citation statements)

References 46 publications

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“…Our earlier work established that pSW100 utilizes a 38-bp TraC-binding sequence for segregation (Fig. 5A) and that this sequence is important to the stability of the plasmid (25). This work verified that a pSW100 derivative that contains a Km resistance gene, pSW100K (Fig.…”

Section: Resultssupporting

confidence: 53%

“…Determining plasmid stability. Plasmid stability was determined using a method that has been described elsewhere (25). Cells were cultured in LB-Km broth overnight at 37°C.…”

Section: Vol 192 2010 Activation Of Rnaii Promoter 3655mentioning

confidence: 99%

“…The mob genes and bom enable pSW100 and pSW200 to be mobilized by the F plasmid (19). Plasmid pSW100 has about 10 copies per cell (18) and uses TraC in the sex pilus assembly as an apparatus for segregation to maintain the stability of the plasmid (25). This study finds that a 9-bp sequence region, which is located at Ϫ43 in RNAIIp in pSW200 and ColE1, is an UP element and important to the synthesis of RNAII and the stability of the plasmid.…”

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A Sequence That Affects the Copy Number and Stability of pSW200 and ColE1

Liu

2010

J Bacteriol

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Pantoea stewartii SW2 contains 13 plasmids. One of these plasmids, pSW200, has a replicon that resembles that of ColE1. This study demonstrates that pSW200 contains a 9-bp UP element, 5-AAGATCTTC, which is located immediately upstream of the ؊35 box in the RNAII promoter. A transcriptional fusion study reveals that substituting this 9-bp sequence reduces the activity of the RNAII promoter by 78%. The same mutation also reduced the number of plasmid copies from 13 to 5, as well as the plasmid stability. When a similar sequence in a ColE1 derivative, pYCW301, is mutated, the copy number of the plasmid also declines from 34 to 16 per cell. Additionally, inserting this 9-bp sequence stabilizes an unstable pSW100 derivative, pSW142K, which also contains a replicon resembling that of ColE1, indicating the importance of this sequence in maintaining the stability of the plasmid. In conclusion, the 9-bp sequence upstream of the ؊35 box in the RNAII promoter is required for the efficient synthesis of RNAII and maintenance of the stability of the plasmids in the ColE1 family.

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Section: Resultssupporting

confidence: 53%

“…Determining plasmid stability. Plasmid stability was determined using a method that has been described elsewhere (25). Cells were cultured in LB-Km broth overnight at 37°C.…”

Section: Vol 192 2010 Activation Of Rnaii Promoter 3655mentioning

confidence: 99%

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A Sequence That Affects the Copy Number and Stability of pSW200 and ColE1

Liu

2010

J Bacteriol

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“…The contribution of conjugal transfer to maintenance of plasmid pKJK5 in E. coli has also been demonstrated in gastrointestinal tracts of rats (1). More recently, a study by Lin and Liu (22) showed that a Pantoea stewartii plasmid, pSW100, is stabilized by the F conjugation system in E. coli. Those authors showed that mutations in genes encoding the sex pilus reduced plasmid stability and hypothesized that pSW100 might use sex pilus assembly as a partitioning apparatus to maintain stability.…”

Section: Discussionmentioning

confidence: 98%

Linear Plasmid SLP2 Is Maintained by Partitioning, Intrahyphal Spread, and Conjugal Transfer inStreptomyces

Hsu¹,

Chen²

2010

J Bacteriol

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Low-copy-number plasmids generally encode a partitioning system to ensure proper segregation after replication. Little is known about partitioning of linear plasmids in Streptomyces. SLP2 is a 50-kb low-copynumber linear plasmid in Streptomyces lividans, which contains a typical parAB partitioning operon. In S. lividans and Streptomyces coelicolor, a parAB deletion resulted in moderate plasmid loss and growth retardation of colonies. The latter was caused by conjugal transfer from plasmid-containing hyphae to plasmidless hyphae. Deletion of the transfer (traB) gene eliminated conjugal transfer, lessened the growth retardation of colonies, and increased plasmid loss through sporulation cycles. The additional deletion of an intrahyphal spread gene (spd1) caused almost complete plasmid loss in a sporulation cycle and eliminated all growth retardation. Moreover, deletion of spd1 alone severely reduced conjugal transfer and stability of SLP2 in S. coelicolor M145 but had no effect on S. lividans TK64. These results revealed the following three systems for SLP2 maintenance: partitioning and spread for moving the plasmid DNA along the hyphae and into spores and conjugal transfer for rescuing plasmidless hyphae. In S. lividans, both spread and partitioning appear to overlap functionally, but in S. coelicolor, spread appears to play the main role.

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“…This demonstrates an element of redundancy between the three MacRPAs, and suggests that the heterotrimeric RPA structure observed in P. furiosus is the exception and not the rule. Lin et al (2008) suggest that intramolecular recombination between RPA homologs may have led to the diversity of RPAs found in euryarchaea, which can function in different pathways or cellular processes.…”

Section: Introductionmentioning

confidence: 99%

Genetic and Biochemical Identification of a Novel Single-Stranded DNA-Binding Complex in Haloferax volcanii

Stroud¹,

Liddell²,

Allers³

2012

Front. Microbio.

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Single-stranded DNA (ssDNA)-binding proteins play an essential role in DNA replication and repair. They use oligonucleotide/oligosaccharide-binding (OB)-folds, a five-stranded β-sheet coiled into a closed barrel, to bind to ssDNA thereby protecting and stabilizing the DNA. In eukaryotes the ssDNA-binding protein (SSB) is known as replication protein A (RPA) and consists of three distinct subunits that function as a heterotrimer. The bacterial homolog is termed SSB and functions as a homotetramer. In the archaeon Haloferax volcanii there are three genes encoding homologs of RPA. Two of the rpa genes (rpa1 and rpa3) exist in operons with a novel gene specific to Euryarchaeota; this gene encodes a protein that we have termed RPA-associated protein (rpap). The rpap genes encode proteins belonging to COG3390 group and feature OB-folds, suggesting that they might cooperate with RPA in binding to ssDNA. Our genetic analysis showed that rpa1 and rpa3 deletion mutants have differing phenotypes; only Δrpa3 strains are hypersensitive to DNA damaging agents. Deletion of the rpa3-associated gene rpap3 led to similar levels of DNA damage sensitivity, as did deletion of the rpa3 operon, suggesting that RPA3 and RPAP3 function in the same pathway. Protein pull-downs involving recombinant hexahistidine-tagged RPAs showed that RPA3 co-purifies with RPAP3, and RPA1 co-purifies with RPAP1. This indicates that the RPAs interact only with their respective associated proteins; this was corroborated by the inability to construct rpa1 rpap3 and rpa3 rpap1 double mutants. This is the first report investigating the individual function of the archaeal COG3390 RPA-associated proteins (RPAPs). We have shown genetically and biochemically that the RPAPs interact with their respective RPAs, and have uncovered a novel single-stranded DNA-binding complex that is unique to Euryarchaeota.

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Stabilization of pSW100 fromPantoea stewartiiby the F Conjugation System

Cited by 5 publications

References 46 publications

A Sequence That Affects the Copy Number and Stability of pSW200 and ColE1

A Sequence That Affects the Copy Number and Stability of pSW200 and ColE1

Linear Plasmid SLP2 Is Maintained by Partitioning, Intrahyphal Spread, and Conjugal Transfer inStreptomyces

Genetic and Biochemical Identification of a Novel Single-Stranded DNA-Binding Complex in Haloferax volcanii

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