Genetic analysis of F sex factor cistrons needed for surface exclusion in Escherichia coli

Achtman, Mark; Manning, Paul A.; Kušećek, Barica; Schwuchow, S; Willetts, Neil

doi:10.1016/0022-2836(80)90065-0

Cited by 69 publications

(37 citation statements)

References 24 publications

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“…Even though ICEs appear to have fairly plastic genomes (7,8), their exclusion genes are conserved. Although entry exclusion in plasmids has been studied for many years (1,10,11,21), there is relatively scant knowledge of the extent of the diversity of exclusion determinants within plasmid families. Further definition of the diversity of exclusion genes in plasmid families will provide useful tools for analyses of the bases of exclusion in plasmids and ultimately further our understanding of the conjugative process.…”

Section: Exclusion Activity Of Eexs and Eexr Exclusion Group Proteinsmentioning

confidence: 99%

“…Similar to F (3) and other conjugative plasmids (11), SXT and R391 carry genes for an entry exclusion system mediated by two inner membrane proteins, one (TraG) in the donor and the other (Eex) in the recipient (16). Entry exclusion systems function to specifically inhibit redundant conjugative transfers between cells that carry the same element (1,10,11,21). We recently showed that even though SXT and R391 have nearly identical conjugative transfer genes, these ICEs do not exclude each other (16).…”

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The SXT/R391 Family of Integrative Conjugative Elements Is Composed of Two Exclusion Groups

Marrero

Waldor

2007

J Bacteriol

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Conjugative elements often encode entry exclusion systems that convert host cells into poor recipients for identical or similar elements. The diversity of exclusion systems within families of conjugative elements has received little attention. We report here the most comprehensive study to date of the diversity of exclusion determinants within a single family of conjugative elements. Unexpectedly, our analyses indicate that there are only two exclusion groups among the diverse members of the SXT/R391 family of integrative conjugative elements.Integrative and conjugative elements (ICEs) are a diverse group of mobile genetic elements with the capacity to incorporate and disseminate genes encoding a variety of properties, including antibiotic resistance, into many bacterial hosts (6). ICEs are self-transmissible, and they transfer from cell to cell via conjugation, similar to conjugative plasmids. However, unlike conjugative plasmids, ICEs do not replicate autonomously; instead, they integrate into the host chromosome. SXT and R391 are closely related ICEs derived from clinical isolates of Vibrio cholerae and Providencia rettgeri, respectively (9, 23). SXT and R391 were the first recognized members of the SXT/ R391 family of ICEs, which now includes ϳ25 members (5). These ICEs were grouped as a family since they encode nearly identical integrases that mediate the integration of the elements into the host chromosome at the prfC locus (8,14). In addition, the members of the SXT/R391 family of ICEs appear to carry a conserved set of genes involved in DNA transfer (8).The conjugal transfer genes in SXT and R391 are distantly related to those found in the F plasmid (4). Similar to F (3) and other conjugative plasmids (11), SXT and R391 carry genes for an entry exclusion system mediated by two inner membrane proteins, one (TraG) in the donor and the other (Eex) in the recipient (16). Entry exclusion systems function to specifically inhibit redundant conjugative transfers between cells that carry the same element (1,10,11,21). We recently showed that even though SXT and R391 have nearly identical conjugative transfer genes, these ICEs do not exclude each other (16). Thus, cells harboring SXT inhibit the acquisition of SXT but not R391 and vice versa. We found that element-specific exclusion activity is mediated by Eex variants EexS and EexR and TraG variants TraG S and TraG R (encoded within SXT and R391, respectively). For example, exclusion was observed when EexS was produced in the recipient and TraG S in the donor; however, EexS did not exclude ICE transfer from a donor producing TraG R . The specificity of the Eex variants is dictated by residues found in their respective carboxyl termini. While the first 86 amino-terminal residues of EexS and EexR are 87% identical, the remaining 56 carboxyl-terminal residues are only 41% identical. Interestingly, a stretch of only 3 amino acids (amino acids 606, 607, and 608) determines TraG's exclusion specificity. At these positions, TraG S has the amino acid sequence P-G-E, wher...

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Section: Exclusion Activity Of Eexs and Eexr Exclusion Group Proteinsmentioning

confidence: 99%

mentioning

confidence: 99%

The SXT/R391 Family of Integrative Conjugative Elements Is Composed of Two Exclusion Groups

Marrero

Waldor

2007

J Bacteriol

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“…The entry exclusion systems of the F and F-like plasmids are the most intensively studied (65,74). Two genes encoded by the F Tra operon, traT and traS, were shown to contribute independently to the Eex phenotype (2,33). The exclusion functions of IncI 1 plasmids R144, R64, and ColIb were shown to belong to one entry exclusion group (26).…”

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TrbK, a small cytoplasmic membrane lipoprotein, functions in entry exclusion of the IncP alpha plasmid RP4

1996

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TrbK is the only plasmid-encoded gene product involved in entry exclusion of the broad-host-range plasmid RP4. The corresponding gene, trbK, coding for a protein of 69 amino acid residues maps in the Tra2 region within the mating pair formation genes. TrbK carries a lipid moiety at the N-terminal cysteine of the mature 47-residue polypeptide. The mutant protein TrbKC23G cannot be modified or proteolytically processed but still acts in entry exclusion with reduced efficiency. An 8-amino-acid truncation at the C terminus of TrbK results in a complete loss of the entry exclusion activity but still allows the protein to be processed. TrbK localizes predominately to the cytoplasmic membrane. Its function depends on presence in the recipient cell but not in the donor cell. TrbK excludes plasmids of homologous systems of the P complex; it is inert towards the IncI system. The likely target for TrbK action is the mating pair formation system, because DNA or any of the components of the relaxosome were excluded as possible targets.Soon after the discovery of the transfer of genetic material by bacterial conjugation, Lederberg and coworkers found that the presence of a plasmid in the recipient cell inhibits transfer of an identical or a closely related plasmid (38). Two distinct processes were found to be responsible for this phenomenon, incompatibility and entry or surface exclusion (48). Entry exclusion functions of several self-transmissible plasmids and of a mobilizable plasmid have been described. These include the plasmids of the incompatibility groups F (1, 2), IncI (20,24,25,27,56), IncP (22,42,44), and IncN (55,75,76) and the mobilizable plasmid ColE1 (77), and the sex pheromone plasmids pAD1 and pCF10 of gram-positive bacteria (18,35,49,73).In general, entry exclusion functions interact only within homologous transfer systems. Entry exclusion (Eex) systems acting on related transfer systems belong to one entry exclusion group. The entry exclusion systems of the F and F-like plasmids are the most intensively studied (65, 74). Two genes encoded by the F Tra operon, traT and traS, were shown to contribute independently to the Eex phenotype (2, 33). The exclusion functions of IncI 1 plasmids R144, R64, and ColIb were shown to belong to one entry exclusion group (26). Two polypeptides responsible for entry exclusion were identified for R144 and R64 (20, 27). Other transfer systems like that of the IncN plasmid pKM101 encode only one gene product which functions in entry exclusion (55).Although the entry exclusion determinants of several plasmids are known, little information is available about the mechanisms by which they inhibit the conjugative DNA transfer into the recipient cell. TraT of the F plasmid is a surface-exposed outer membrane lipoprotein and was thought to inhibit stable mating pair formation (1,54,65). TraS of the F plasmid and ExcA, the 19-kDa polypeptide of the IncI 1 plasmid R144, were shown to localize to the cytoplasmic membrane and were thought to inhibit DNA transfer after mating pairs have been...

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“…F ϩ cells in exponential growth inhibit the redundant entry of highly related F plasmids in a process called surface exclusion. Mutational analysis has identified two genetically distinct surface exclusion systems encoded by the F genes traS and traT (2). While the traT gene product appears to reduce the ability of cells to maintain stable mating aggregates, the mechanism through which traS acts remains unproven (1,34).…”

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Redundant transfer of F' plasmids occurs between Escherichia coli cells during nonlethal selections

Peters

Benson

1995

J Bacteriol

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Surface exclusion is the mechanism by which F plasmids prevent the redundant entry of additional F plasmids into the host cell during exponential growth. This mechanism is relaxed in cells that are in stationary phase. Using genetically marked F plasmids and host strains, we extend this finding to Escherichia coli populations during extended nonlethal selection in bacterial lawns. We show that a high level of redundant transfer occurs between these nongrowing cells during the selection. This result has implications for the mechanism of adaptive mutagenesis.Our lab is investigating nonlethal bacterial selection systems in which cells appear to undergo adaptive mutagenesis. Adaptive mutagenesis implicates environmental stress factors as a cause of specific advantageous mutations and not just as a selective force that enriches for random beneficial mutations within the population. The nonlethal genetic selection systems used to study adaptive mutation share basic tenets (11). In these systems beneficial mutations occur in the absence of significant cell growth and without the accumulation of nonadvantageous mutations (6,7,14). One of the best-studied systems utilizes an Escherichia coli K-12 strain in which the chromosomal lac genes are deleted and which carries an FЈ lac plasmid containing an out-of-frame ⌽(lacI q -lacZ)33 fusion. The strain is unable to grow on minimal lactose plates (Lac Ϫ ). When plated on lactose media, Lac ϩ colonies appear after 2 days and new colonies continue to form for many days (7). Most of the Lac ϩ revertants contain deletions within small mononucleotide repeats that restore a reading frame to the lacZ gene (13,23). An important question is that of how these alterations occur without cell division and chromosome replication (7). Long-standing research concerning F plasmid biology suggests a solution to this paradox.Conjugative F plasmids were first described nearly 50 years ago (20). It was soon noted that although F plasmids efficiently block the entry of additional F plasmids, this blockage is relaxed in stationary-phase cells (19), yielding F ϩ cells that are phenotypically F Ϫ (termed F Ϫ phenocopies) (19). The extent to which F ϩ cells relaxed this block was estimated by incubating exponentially growing liquid cultures of F ϩ cells with F ϩ cells from stationary-phase cultures (33). Procedures that optimized the F Ϫ phenocopy condition were later developed (4). These procedures involved growing cells into stationary phase and maintaining them at a specific pH in storage media (4). Given that stationary-phase F ϩ cells do not block the redundant entry of additional FЈ plasmids, a high level of transfer might occur between F ϩ cells during nonlethal selections. Because FЈ plasmid transfer involves replication of the plasmid genome in both the donor and recipient cells, this may provide a mechanism of preferential mutagenesis for plasmid-encoded genes. Moreover, it results in multiple plasmids within the same cell, thus allowing various recombination events to occur.To determine whe...

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Genetic analysis of F sex factor cistrons needed for surface exclusion in Escherichia coli

Cited by 69 publications

References 24 publications

The SXT/R391 Family of Integrative Conjugative Elements Is Composed of Two Exclusion Groups

The SXT/R391 Family of Integrative Conjugative Elements Is Composed of Two Exclusion Groups

TrbK, a small cytoplasmic membrane lipoprotein, functions in entry exclusion of the IncP alpha plasmid RP4

Redundant transfer of F' plasmids occurs between Escherichia coli cells during nonlethal selections

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