Mobile genomic islands (GIs) can be excised from the chromosome,then form a circular intermediate and be reintegrated into the chromosome by the GI internal integrase.Some mobile GIs can also be transferred into a new receptor cell by transformation,conjugation,or transduction.The action sites of the integrase are usually flanked direct repeats (DRs) of the GIs.Accurate localization of the flanking sequences is a precondition for determining the mobility of the GI.Mobile GIs are generally associated with transfer RNAs (tRNAs).Based on the correlation between flanking sequences and tRNA sequences,the flanking sequences of 11 putative mobile GIs in Pseudomonas aeruginosa PAO1,P.aeruginosa PA14,P.fluorescens Pf-5 and P.fluorescens Pf0-1 were identified.Among the 11 GIs,Pf0-1GI-1 is responsible for benzoate degradation.PAO1GI-1,Pf5GI-2,Pf5GI-3,and Pf5GI-4 were confirmed experimentally to be excised from a chromosome to form a circular intermediate.The action sites of the integrases are these GIs direct repeats.Due to distinct DRs,cutting sites for the internal integrase of PAO1GI-1,Pf5GI-2,Pf5GI-3 and Pf5GI-4 were determined outside the T-loop of the tRNA Gly gene,outside the anticodon loop of the tRNA Ser gene and tRNALys gene,and at the asymmetric 3'-end of the tRNA Leu gene,respectively.PAO1GI-1 and other mobile GIs may be transferred into many different strains that belong to different phyla because of the clear flanking sequences.This study describes basic information about the action sites of the integrases,assesses the mobility of GIs,and can help design and transfer mobile GIs to candidate strains.
tmRNA,a combination of a tRNA-related fragment and a small mRNA fragment,was confirmed as the integration site of genomic islands(GIs).Using sequence alignment and comparative genomics,68 GIs associated with tmRNA genes were identified among 13 genera of Enterobacteriaceae.Among them,53 GIs were found in Escherichia coli and Salmonella enterica.Among these 53 GIs,tandem GIs were verified in eight S.enterica and two E.coli chromosomes.The downstream regions of the tmRNA genes in most of the E.coli and S.enterica chromosomes include one GI or tandem GIs region and a remnant variable region distal to the tmRNA.The chronology of integration of tandem GIs into the genome indicated that GIs farther from the tmRNA were incorporated into the genome earlier than those nearer from the tmRNA.The integrases of the tmRNA gene-associated GIs can be further categorized into three subtypes:HP1 integrases,PhiCTX integrases,and P4 integrases,which are the most predominant.The GIs were first integrated into the chromosome by the P4 integrase,subsequently by the PhiCTX integrase,and finally by the HP1 integrase.Thus,the tmRNA gene is an important site for investigating the genetics and evolution of tandem GIs.
Based on three distinct traits of genomic islands, a novel approach was developed to search for and determine genomic islands in special strains. Two genomic islands in Pseudomonas aeruginosa PAO1 and 7 genomic islands in Pseudomonas aeruginosa PA14 were defined with this method. Among the 9 genomic islands, 4 islands had been characterized before, while the other 5 islands were initially determined. The insert sites of 6 genomic islands are tRNA sequences, direct repeats of PA14GI-3 are relative to tRNALeu, and direct repeats of PA14GI-2 are at the 3′ end of bifunctional GMP synthase/ glutamine amidotransferase. Only direct repeats of PA14GI-4 are not clear. Among the 5 newly-found genomic islands, it was supposed that PA14GI-2 is a genomic island related to Hg2+ uptake, PA14GI-3 is a secretory activity genomic island, PA14GI-6 is a pathogenicity island, and functions of PA14GI-1 and PA14GI-5 are not clear. Finally, the tyrosine type integrases in PAO1GI-1, PA14GI-5 and PA14GI-7 were analyzed, and their binding and restriction sites were predicted.
