Plasmid transfer in Streptococcus faecalis: Production of multiple sex pheromones by recipients

Dunny, Gary M.; Craig, Ronald A.; Carron, Richard L.; Clewell, Don B.

doi:10.1016/0147-619x(79)90029-5

Cited by 260 publications

(255 citation statements)

References 16 publications

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“…The concentration of cAD1 in CF was determined by the microdilution assay method described previously (3). The cAD1 titer is defined as the highest dilution of culture filtrate which induced aggregate formation in responder cells.…”

Section: Methodsmentioning

confidence: 99%

Regulation of the pAD1 sex pheromone response in Enterococcus faecalis: effects of host strain and traA, traB, and C region mutants on expression of an E region pheromone-inducible lacZ fusion

Weaver

Clewell

1990

J Bacteriol

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Pheromone-induced conjugal transfer of the hemolysin-bacteriocin plasmid pADl of Enterococcusfaecalis is regulated by a cluster of determinants designated traA, traB, and regions C and E. The E region is believed to include a positive regulator that controls many structural genes related to conjugation. The pheromoneinducible Tn917-lac fusion NR5, located in the E region, is regulated by the products of traA, traB, and the C region. To more closely examine the effects of these genes on the induction of E region products, inserts in each of these genes were combined with the NR5 fusion in a novel approach involving triparental matings with a pADi miniplasmid and recombinational mutagenesis. Results indicate that (i) the traA gene product is a key repressor of the pheromone response; (ii) the traB gene product, in cooperation with a gene within or regulated by the E region, controls pheromone shutdown; (iii) a primary function of the C region gene product is in pheromone sensing, with secondary functions in pheromone shutdown and negative regulation; and (iv) the host in which the plasmid resides has a dramatic effect on the regulation of the NR5 fusion in traB and C region mutants. Numerous parallels were observed between the regulation of the NR5 fusion and the regulation of the aggregation and transfer response. These parallels aided in further defining the functions of particular regulatory determinants as well as further establishing the link between the regulation of the E region and the regulation of the aggregation and transfer response.The Enterococcus faecalis plasmid pAD1 (59.6 kilobases) encodes a conjugative transfer system that is induced specifically by a small peptide pheromone, cAD1, produced by potential recipients. When exposed to pheromone, plasmidcontaining cells respond by producing: (i) an aggregation substance which facilitates the formation of mating or selfaggregates, (ii) surface proteins that have been correlated with aggregation substance, and (iii) other functions believed to be essential for the actual physical transfer of the plasmid DNA. Once a recipient acquires pAD1, cAD1 activity is shut down, while the activity of pheromones specific for other plasmids is unaffected. In addition, plasmid-containing cells produce a specific peptide, iAD1, which specifically and competitively inhibits cAD1 activity (for a recent review, see reference 2).The pAD1 pheromone response was previously shown to be regulated by a cluster of genes located in the 15-kilobase EcoRI B restriction fragment (4, 7, 13). Miniplasmids consisting solely of this fragment and containing pheromoneinducible lacZ fusions were found to regulate the fusions in a manner identical to that observed when the same fusions were present on the intact plasmid (14). These results indicated that the regulatory genes on the pAD1 EcoRI B fragment were not only essential but also sufficient for directing a normal pheromone response. Mutational analyses with Tn917 and transcriptional analyses with Tn917-lac have identified genetic det...

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Section: Methodsmentioning

confidence: 99%

Regulation of the pAD1 sex pheromone response in Enterococcus faecalis: effects of host strain and traA, traB, and C region mutants on expression of an E region pheromone-inducible lacZ fusion

Weaver

Clewell

1990

J Bacteriol

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“…Similar results were obtained using donors carrying pAD1, pAMγ1 or pOB1. 108,109 We also found that when donors were exposed to a recipientfiltrate for a couple of hours prior to mixing with recipient cells, conjugative transfer of plasmid DNA occurred efficiently in short (e.g., 10 min) matings, in contrast to the 90 min period generally needed in conventional mating experiments before transfer became optimal. Without prior exposure of donors to recipient filtrate, little if any transfer was observed in 10 min.…”

Section: Sex Pheromonesmentioning

confidence: 93%

Tales of conjugation and sex pheromones

Clewell¹

2011

Mobile Genetic Elements

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REVIEW REVIEW use of a technique that had recently been reported by Godson and Sinsheimer 6 that utilized a detergent mixture of Brij 58 (a neutral detergent) and deoxycholate for lysis of phageinfected E. coli. During a relatively low-speed centrifugation step, the vast majority of chromosomal DNA pelleted leaving the supernatant greatly enriched in plasmid DNA. Sucrose gradient analyses of these "cleared lysates" revealed a ColE1-protein complex sedimenting slightly faster (24S) than the 23S protein-free supercoiled DNA. Surprisingly, when the 24S complex was exposed to conditions expected to affect protein structure such as proteases, strong ionic detergents or heat, the 24S complex converted to a 17S relaxed (open circular) configuration. The relaxation event involved the introduction of a strand-specific nick in the supercoiled plasmid; we therefore called the 24S entity a "relaxation complex". 7,8 The percentage of plasmid DNA that was present in the complexed state varied from less than 30%, when glucose was in the growth medium, to more than 80% when glucose was absent.

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“…Transfer frequencies were calculated as the number of transconjugants per donor cell (at the end of mating). Pheromone induction and detection of cell aggregation were performed as previously described (11,12).…”

Section: Methodsmentioning

confidence: 99%

Cloning and Genetic Analyses of the Bacteriocin 41 Determinant Encoded on the Enterococcus faecalis Pheromone-Responsive Conjugative Plasmid pYI14: a Novel Bacteriocin Complemented by Two Extracellular Components (Lysin and Activator)

Tomita¹,

Kamei²,

Ike

2008

J Bacteriol

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The conjugative plasmid pYI14 (61 kbp) was isolated from Enterococcus faecalis YI714, a clinical isolate. pYI14 conferred a pheromone response on its host and encoded bacteriocin 41 (bac41). Bacteriocin 41 (Bac41) only showed activity against E. faecalis. Physical mapping of pYI14 showed that it consisted of EcoRI fragments A to P. The clone pHT1100, containing EcoRI fragments A (12.6 kbp) and H (3.5 kbp), conferred the bacteriocin activity on E. faecalis strains. Genetic analysis showed that the determinant was located in a 6.6-kbp region within the EcoRI AH fragments. Six open reading frames (ORFs) were identified in this region and designated ORF7 (bacL 1 ) ORF8 (bacL 2 ), ORF9, ORF10, ORF11 (bacA), and ORF12 (bacI). They were aligned in this order and oriented in the same direction. ORFs bacL 1 , bacL 2 , bacA, and bacI were essential for expression of the bacteriocin in E. faecalis. Extracellular complementation of bacteriocin expression was possible for bacL 1 and -L 2 and bacA mutants. bacL 1 and -L 2 and bacA encoded bacteriocin component L and activator component A, respectively. The products of these genes are secreted into the culture medium and extracellularly complement bacteriocin expression. bacI encoded immunity, providing the host with resistance to its own bacteriocin activity. The bacL 1 -encoded protein had significant homology with lytic enzymes that attack the gram-positive bacterial cell wall. Sequence data for the deduced bacL 1 -encoded protein suggested that it has a domain structure consisting of an N-terminal signal peptide, a second domain with the enzymatic activity, and a third domain with a three-repeat structure directing the proenzyme to its cell surface receptor.Bacteriocins are bacterial proteins or peptides which inhibit the growth of other bacteria that are closely related to the producer strain. They usually exhibit a relatively narrow spectrum of activity and are produced by a wide variety of grampositive and gram-negative bacteria (27). Bacteriocin production is thought to provide the host strain with an ecological or other selective advantage over other strains.Many Enterococcus faecalis clinical isolates produce a bacteriocin (3, 5), and the bacteriocin is frequently encoded on the E. faecalis pheromone-responding conjugative plasmid (6,14,21,46). Several E. faecalis bacteriocins have been genetically and biochemically characterized (15, 35), including the ␤-hemolysin/bacteriocin (cytolysin) (6,7,18,20,22) and the peptide antibiotics AS-48 (33), bacteriocin 21 (47), and bacteriocin 31 (46), which are encoded by the E. faecalis conjugative plasmids pAD1 (58 kbp), pMB2 (58 kbp), pPD1 (59 kbp), and pYI17 (57.5 kbp), respectively. A significant number of E. faecalis clinical isolates produce hemolysin/bacteriocin (10, 26), and more than 50% of the hemolytic clinical isolates carry transferable hemolysin/bacteriocin determinants (21, 26). The hemolysin/bacteriocin of pAD1 is associated with virulence in animal models (4,25,29), and this plasmid is considered to be a typical ...

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Plasmid transfer in Streptococcus faecalis: Production of multiple sex pheromones by recipients

Cited by 260 publications

References 16 publications

Regulation of the pAD1 sex pheromone response in Enterococcus faecalis: effects of host strain and traA, traB, and C region mutants on expression of an E region pheromone-inducible lacZ fusion

Regulation of the pAD1 sex pheromone response in Enterococcus faecalis: effects of host strain and traA, traB, and C region mutants on expression of an E region pheromone-inducible lacZ fusion

Tales of conjugation and sex pheromones

Cloning and Genetic Analyses of the Bacteriocin 41 Determinant Encoded on the Enterococcus faecalis Pheromone-Responsive Conjugative Plasmid pYI14: a Novel Bacteriocin Complemented by Two Extracellular Components (Lysin and Activator)

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