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The rulB gene of plasmid pWW0 is a hotspot for the site-specific insertion of integron-like elements found in the chromosomes of environmental Pseudomonas fluorescens group bacteria.

Rhodes G, Bosma H, Studholme D, Arnold DL, Jackson RW, Pickup RW - Environ. Microbiol. (2014)

Bottom Line: Here, for the first time, we show that rulB on plasmid pWW0 is a hotspot for the active site-specific integration of related integron-like elements (ILEs) found in six environmental pseudomonads (strains FH1-FH6).Integration into rulB on pWW0 occurred at position 6488 generating a 3 bp direct repeat.Collectively, these ILEs share features with the previously described type III protein secretion system effector ILEs and are considered important to host survival and transfer of fitness enhancing and (a)virulence genes between bacteria.

View Article: PubMed Central - PubMed

Affiliation: Centre for Ecology and Hydrology, Lancaster Environment Centre, Library Avenue, Bailrigg, Lancaster, LA1 4AP, UK.

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Alignment of intergenic regions found immediately upstream and downstream of rulAB′ on integron-like elements.A. Alignment of the predicted promoter region and LexA binding site upstream of rulA. The conserved CTG-N10-CAG LexA binding site motif (yellow), the −35 box (bold and underlined) and the −10 box (underlined bold italics) are highlighted.B. The 118–119 bp intergenic region between the known 5′-GAT-3′ insertion point in pWW0 and the predicted ATG start codon of ORF1 (xerC/xerD) aligned with chromosomal locations in strains FH1, FH4 and FH5, and other close relatives. aNo ATG start codon for rulA in FH5 chromosome.
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fig04: Alignment of intergenic regions found immediately upstream and downstream of rulAB′ on integron-like elements.A. Alignment of the predicted promoter region and LexA binding site upstream of rulA. The conserved CTG-N10-CAG LexA binding site motif (yellow), the −35 box (bold and underlined) and the −10 box (underlined bold italics) are highlighted.B. The 118–119 bp intergenic region between the known 5′-GAT-3′ insertion point in pWW0 and the predicted ATG start codon of ORF1 (xerC/xerD) aligned with chromosomal locations in strains FH1, FH4 and FH5, and other close relatives. aNo ATG start codon for rulA in FH5 chromosome.

Mentions: Comparison of the sequences of ILEFH1, ILEFH4 and ILEFH5 with proposed ILEs in pGRT1, PAGI-5, RGP63, pUM505 and another candidate region on the chromosome of P. syringae pv. tomato DC3000 showed that all share structural features with the recently proposed type III protein secretion system effector (T3SE) ILEs (Jackson et al., 2011). T3SE ILEs have T3SE gene(s) orientated so that the transcription is towards the 3′ end of the integrase gene and therefore not under the influence of the integrase Pc promoter. Although we have not identified T3SE genes on the ILEs here, this feature is shared with the integrated genes downstream of tetR in the ILEFH1 and ILEFH5, and on pGRT1, but not with all sequences downstream of tetR in FH4, pUMU505, PAGI-5 and RGP63 (Fig. 3). In addition, we have been unable to demonstrate the presence of a Pc promoter in the upstream integrase gene. However, even if present, its influence would not be exerted on rulA or disrupted rulB' that flank the element since they are transcribed in the opposite direction. In T3SE ILEs, insertion into the rulAB operon is considered likely to be under the influence of the LexA repressor because of a LexA binding region in the rulAB promoter (Jackson et al., 2011). Consistent with this, we found LexA1 binding sites with the characteristic CTG-N10-CAG motif upstream of rulA in each of the chromosomally located ILEs of FH1, FH4 and FH5 as well in plasmids pWW0, pGRT1 and genomic islands PAGI-5, RGP63 and pUM505 (Fig. 4A).


The rulB gene of plasmid pWW0 is a hotspot for the site-specific insertion of integron-like elements found in the chromosomes of environmental Pseudomonas fluorescens group bacteria.

Rhodes G, Bosma H, Studholme D, Arnold DL, Jackson RW, Pickup RW - Environ. Microbiol. (2014)

Alignment of intergenic regions found immediately upstream and downstream of rulAB′ on integron-like elements.A. Alignment of the predicted promoter region and LexA binding site upstream of rulA. The conserved CTG-N10-CAG LexA binding site motif (yellow), the −35 box (bold and underlined) and the −10 box (underlined bold italics) are highlighted.B. The 118–119 bp intergenic region between the known 5′-GAT-3′ insertion point in pWW0 and the predicted ATG start codon of ORF1 (xerC/xerD) aligned with chromosomal locations in strains FH1, FH4 and FH5, and other close relatives. aNo ATG start codon for rulA in FH5 chromosome.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4542609&req=5

fig04: Alignment of intergenic regions found immediately upstream and downstream of rulAB′ on integron-like elements.A. Alignment of the predicted promoter region and LexA binding site upstream of rulA. The conserved CTG-N10-CAG LexA binding site motif (yellow), the −35 box (bold and underlined) and the −10 box (underlined bold italics) are highlighted.B. The 118–119 bp intergenic region between the known 5′-GAT-3′ insertion point in pWW0 and the predicted ATG start codon of ORF1 (xerC/xerD) aligned with chromosomal locations in strains FH1, FH4 and FH5, and other close relatives. aNo ATG start codon for rulA in FH5 chromosome.
Mentions: Comparison of the sequences of ILEFH1, ILEFH4 and ILEFH5 with proposed ILEs in pGRT1, PAGI-5, RGP63, pUM505 and another candidate region on the chromosome of P. syringae pv. tomato DC3000 showed that all share structural features with the recently proposed type III protein secretion system effector (T3SE) ILEs (Jackson et al., 2011). T3SE ILEs have T3SE gene(s) orientated so that the transcription is towards the 3′ end of the integrase gene and therefore not under the influence of the integrase Pc promoter. Although we have not identified T3SE genes on the ILEs here, this feature is shared with the integrated genes downstream of tetR in the ILEFH1 and ILEFH5, and on pGRT1, but not with all sequences downstream of tetR in FH4, pUMU505, PAGI-5 and RGP63 (Fig. 3). In addition, we have been unable to demonstrate the presence of a Pc promoter in the upstream integrase gene. However, even if present, its influence would not be exerted on rulA or disrupted rulB' that flank the element since they are transcribed in the opposite direction. In T3SE ILEs, insertion into the rulAB operon is considered likely to be under the influence of the LexA repressor because of a LexA binding region in the rulAB promoter (Jackson et al., 2011). Consistent with this, we found LexA1 binding sites with the characteristic CTG-N10-CAG motif upstream of rulA in each of the chromosomally located ILEs of FH1, FH4 and FH5 as well in plasmids pWW0, pGRT1 and genomic islands PAGI-5, RGP63 and pUM505 (Fig. 4A).

Bottom Line: Here, for the first time, we show that rulB on plasmid pWW0 is a hotspot for the active site-specific integration of related integron-like elements (ILEs) found in six environmental pseudomonads (strains FH1-FH6).Integration into rulB on pWW0 occurred at position 6488 generating a 3 bp direct repeat.Collectively, these ILEs share features with the previously described type III protein secretion system effector ILEs and are considered important to host survival and transfer of fitness enhancing and (a)virulence genes between bacteria.

View Article: PubMed Central - PubMed

Affiliation: Centre for Ecology and Hydrology, Lancaster Environment Centre, Library Avenue, Bailrigg, Lancaster, LA1 4AP, UK.

Show MeSH
Related in: MedlinePlus