Identification and characterization of genes encoding sex pheromone cAM373 activity in <i>Enterococcus faecalis</i> and <i>Staphylococcus aureus</i>

Flannagan, Susan E.; Clewell, Don B.

doi:10.1046/j.1365-2958.2002.02922.x

Cited by 60 publications

(47 citation statements)

References 62 publications

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“…pSU18bac* contains a point mutation in the promoter Ϫ10 box, which results in the sequence CATAAT. From here the SalI/KpnI fragment that contains the bac promoter was subcloned into pAM434 (21), yielding pAM434b*.…”

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confidence: 99%

Replication of Enterococcus faecalis Pheromone-Responding Plasmid pAD1: Location of the Minimal Replicon and oriV Site and RepA Involvement in Initiation of Replication

et al. 2004

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The hemolysin-determining plasmid pAD1 is a member of a widely disseminated family of highly conjugative elements commonly present in clinical isolates of Enterococcus faecalis. The determinants repA, repB, and repC, as well as adjacent iteron sequences, are believed to play important roles in pAD1 replication and maintenance. The repA gene encodes an initiator protein, whereas repB and repC encode proteins related to stability and copy number. The present study focuses specifically on repA and identifies a replication origin (oriV) within a central region of the repA determinant. A small segment of repA carrying oriV was able to support replication in cis of a plasmid vector otherwise unable to replicate, if an intact RepA was supplied in trans. We demonstrate that under conditions in which RepA is expressed from an artificial promoter, a segment of DNA carrying only repA is sufficient for stable replication in E. faecalis. We also show that RepA binds specifically to oriV DNA at several sites containing inverted repeat sequences (i.e., IR-1) and nonspecifically to single-stranded DNA, and related genetic analyses confirm that these sequences play an important role in replication. Finally, we reveal a relationship between the internal structure of RepA and its ability to recognize oriV. An in-frame deletion within repA resulting in loss of 105 nucleotides, including at least part of oriV, did not eliminate the ability of the altered RepA protein to initiate replication using an intact origin provided in trans. The relationship of RepA to other known initiator proteins is also discussed.pAD1 is a 60-kb, conjugative plasmid originally identified in Enterococcus faecalis DS16 (12,23,49). It encodes a cytolysin (hemolysin/bacteriocin) that contributes to virulence in animal models (8,37,40) and is one of numerous plasmids in E. faecalis that facilitate a response to peptide sex pheromones secreted by plasmid-free (recipient) bacteria. pAD1 responds to the pheromone cAD1 and represents a widely disseminated family of cytolysin plasmids commonly associated with clinical infections in humans (38) and which, for the most part, are members of the same incompatibility group (13,34). (For recent reviews, see references 9 and 10.)Nucleotide sequence data relating to plasmids from different incompatibility groups (e.g., pAD1, pAM373, pCF10, and pPD1) and responding to four different pheromones have shown that the regions associated with replication and maintenance are organized similarly; in all cases, this region is located adjacent to that involved in regulation of the pheromone response (9, 10). In the case of pAD1 the key determinants associated with plasmid maintenance are repA, repB, and repC (Fig. 1A). On the basis of sequence homology, repA is believed to encode the initiator of vegetative replication, whereas repB and repC most likely represent a partition system (50, 54). When a segment carrying these three determinants was cloned on an E. coli plasmid vector, it enabled the chimera to replicate in E. faecalis (52...

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Replication of Enterococcus faecalis Pheromone-Responding Plasmid pAD1: Location of the Minimal Replicon and oriV Site and RepA Involvement in Initiation of Replication

et al. 2004

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“…This degradation is important, because signal peptides may be harmful to the cell, as they can interfere with membrane integrity and block protein translocation via the Sec machinery (25,46,165). In some cases, the cleaved fragments of signal peptides are released from the membrane and function in signal transduction pathways, both in eukaryotes (87,101) and in bacteria (6,41). Notably, multiple enzymes, such as the bacterial RseP and signal peptide peptidase A (SPPA) enzymes, appear to be involved in signal peptide degradation in species ranging from bacteria to humans.…”

Section: Signal Peptide Hydrolases Degrade Signal Peptides Within or mentioning

confidence: 99%

“…RseP was furthermore shown to cleave a ␤-lactamase signal peptide fused to the maltose binding protein in E. coli (2) and also the signal peptides of several prelipoproteins in Enterococcus faecalis and S. aureus, to generate sex pheromones (6,41). An SPP function of the S2P RasP would be consistent with the secretion defects described for a rasP mutant of B. subtilis (54), as signal peptides have a known inhibitory effect on preprotein translocation across the membrane (25,46,165).…”

Section: Rsepmentioning

confidence: 99%

Membrane Proteases in the Bacterial Protein Secretion and Quality Control Pathway

Dalbey

Wang

Dijl

2012

Microbiol Mol Biol Rev

129

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SUMMARY Proteolytic cleavage of proteins that are permanently or transiently associated with the cytoplasmic membrane is crucially important for a wide range of essential processes in bacteria. This applies in particular to the secretion of proteins and to membrane protein quality control. Major progress has been made in elucidating the structure-function relationships of many of the responsible membrane proteases, including signal peptidases, signal peptide hydrolases, FtsH, the rhomboid protease GlpG, and the site 1 protease DegS. These enzymes employ very different mechanisms to cleave substrates at the cytoplasmic and extracytoplasmic membrane surfaces or within the plane of the membrane. This review highlights the different ways that bacterial membrane proteases degrade their substrates, with special emphasis on catalytic mechanisms and substrate delivery to the respective active sites.

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“…The integration site occurs in genes of relevance to S. aureus pathogenesis (set19 and essA) and antimicrobial resistance (vraE and fmt1) and in other genes (see Table S4). Interestingly, the integration site also occurs in camS, which encodes a lipoprotein that acts as a pheromone for conjugative plasmids (24). Diverse ICE6013 subfamilies in staphylococci.…”

Section: Resultsmentioning

confidence: 99%

Transposase-Mediated Excision, Conjugative Transfer, and Diversity of ICE 6013 Elements in Staphylococcus aureus

et al. 2017

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ICE6013 represents one of two families of integrative conjugative elements (ICEs) identified in the pan-genome of the human and animal pathogen Staphylococcus aureus. Here we investigated the excision and conjugation functions of ICE6013 and further characterized the diversity of this element. ICE6013 excision was not significantly affected by growth, temperature, pH, or UV exposure and did not depend on recA. The IS30-like DDE transposase (Tpase; encoded by orf1 and orf2) of ICE6013 must be uninterrupted for excision to occur, whereas disrupting three of the other open reading frames (ORFs) on the element significantly affects the level of excision. We demonstrate that ICE6013 conjugatively transfers to different S. aureus backgrounds at frequencies approaching that of the conjugative plasmid pGO1. We found that excision is required for conjugation, that not all S. aureus backgrounds are successful recipients, and that transconjugants acquire the ability to transfer ICE6013. Sequencing of chromosomal integration sites in serially passaged transconjugants revealed a significant integration site preference for a 15-bp AT-rich palindromic consensus sequence, which surrounds the 3-bp target site that is duplicated upon integration. A sequence analysis of ICE6013 from different host strains of S. aureus and from eight other species of staphylococci identified seven divergent subfamilies of ICE6013 that include sequences previously classified as a transposon, a plasmid, and various ICEs. In summary, these results indicate that the IS30-like Tpase functions as the ICE6013 recombinase and that ICE6013 represents a diverse family of mobile genetic elements that mediate conjugation in staphylococci.IMPORTANCE Integrative conjugative elements (ICEs) encode the abilities to integrate into and excise from bacterial chromosomes and plasmids and mediate conjugation between bacteria. As agents of horizontal gene transfer, ICEs may affect bacterial evolution. ICE6013 represents one of two known families of ICEs in the pathogen Staphylococcus aureus, but its core functions of excision and conjugation are not well studied. Here, we show that ICE6013 depends on its IS30-like DDE transposase for excision, which is unique among ICEs, and we demonstrate the conjugative transfer and integration site preference of ICE6013. A sequence analysis revealed that ICE6013 has diverged into seven subfamilies that are dispersed among staphylococci.

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Identification and characterization of genes encoding sex pheromone cAM373 activity in Enterococcus faecalis and Staphylococcus aureus

Cited by 60 publications

References 62 publications

Replication of Enterococcus faecalis Pheromone-Responding Plasmid pAD1: Location of the Minimal Replicon and oriV Site and RepA Involvement in Initiation of Replication

Replication of Enterococcus faecalis Pheromone-Responding Plasmid pAD1: Location of the Minimal Replicon and oriV Site and RepA Involvement in Initiation of Replication

Membrane Proteases in the Bacterial Protein Secretion and Quality Control Pathway

Transposase-Mediated Excision, Conjugative Transfer, and Diversity of ICE 6013 Elements in Staphylococcus aureus

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