Mobility and Generation of Mosaic Non-Autonomous Transposons by Tn3-Derived Inverted-Repeat Miniature Elements (TIMEs)

Abstract

Functional transposable elements (TEs) of several Pseudomonas spp. strains isolated from black shale ore of Lubin mine and from post-flotation tailings of Zelazny Most in Poland, were identified using a positive selection trap plasmid strategy. This approach led to the capture and characterization of (i) 13 insertion sequences from 5 IS families (IS3, IS5, ISL3, IS30 and IS1380), (ii) isoforms of two Tn3-family transposons – Tn5563a and Tn4662a (the latter contains a toxin-antitoxin system), as well as (iii) non-autonomous TEs of diverse structure, ranging in size from 262 to 3892 bp. The non-autonomous elements transposed into AT-rich DNA regions and generated 5- or 6-bp sequence duplications at the target site of transposition. Although these TEs lack a transposase gene, they contain homologous 38-bp-long terminal inverted repeat sequences (IRs), highly conserved in Tn5563a and many other Tn3-family transposons. The simplest elements of this type, designated TIMEs (Tn3 family-derived Inverted-repeat Miniature Elements) (262 bp), were identified within two natural plasmids (pZM1P1 and pLM8P2) of Pseudomonas spp. It was demonstrated that TIMEs are able to mobilize segments of plasmid DNA for transposition, which results in the generation of more complex non-autonomous elements, resembling IS-driven composite transposons in structure. Such transposon-like elements may contain different functional genetic modules in their core regions, including plasmid replication systems. Another non-autonomous element “captured” with a trap plasmid was a TIME derivative containing a predicted resolvase gene and a res site typical for many Tn3-family transposons. The identification of a portable site-specific recombination system is another intriguing example confirming the important role of non-autonomous TEs of the TIME family in shuffling genetic information in bacterial genomes. Transposition of such mosaic elements may have a significant impact on diversity and evolution, not only of transposons and plasmids, but also of other types of mobile genetic elements.

Mechanisms Involved in Acquisition of bla NDM Genes by IncA/C 2 and IncFII Y Plasmids

“…In all of these plasmids, bla NDM lies within a 5,945-bp region matching Tn125 that includes 101 bp of ISAba125 and a fragment of ISCR27. This region is flanked by two copies of a 256-bp Tn3-derived invertedrepeat transposable element (TIME), each bounded by 38-bp IRs (42). These TIMEs, previously described as miniature invertedrepeat transposable elements (MITEs), may have been responsible for capturing the bla NDM region from a pNDM-BJ01-like plasmid (18,19,43).…”

“…TIMEs create a 5-to 6-bp DR on transposition like the Tn3 transposons from which they appear to be derived (42). In these plasmids, the 5-bp sequences adjacent to the "inside" of each TIME flanking the rmtC region are identical (TAT AA).…”

Autonomous and non-autonomous Tn3-family transposons and their role in the evolution of mobile genetic elements

“…11 This approach permitted the characterization of several typical non-composite transposons: (i-ii) Tn3434 and Tn6122 -cryptic elements 9,12 , (iii) Tn5393 -conferring streptomycin resistance 9,12,13 (iv) Tn4662a -containing a predicted toxin-antitoxin module, and (v) Tn5563a -carrying part of the mer mercury resistance operon. 10 Interestingly, we also identified several TEs of unusual structure, which significantly broaden our knowledge on the diversity and evolution of Tn3 family transposons. These atypical elements are (i) a composite transposon TnPpa1 of Paracoccus pantotrophus DSM 11072 (Alphaproteobacteria) 14 and (ii) a group of non-composite transposons of diverse structure isolated from Pseudomonas spp.…”

“…5 We have analyzed functional TEs of the Tn3-family in a pool of environmental strains of the genera Paracoccus (Alphaproteobacteria) 9 and Pseudomonas (Gammaproteobacteria). 10 These elements were identified using trap plasmids which enable positive selection of transposition events and therefore the direct "capture" of even phenotypically silent elements. 11 This approach permitted the characterization of several typical non-composite transposons: (i-ii) Tn3434 and Tn6122 -cryptic elements 9,12 , (iii) Tn5393 -conferring streptomycin resistance 9,12,13 (iv) Tn4662a -containing a predicted toxin-antitoxin module, and (v) Tn5563a -carrying part of the mer mercury resistance operon.…”

The genetic analysis of Enterobacteriaceae plasmids to provide insights into the acquisition and spread of the blaNDM gene

“…TIME create 5-6 bp DR on transposition like the Tn3 transposons from which they appear to be derived (259). In these plasmids the 5 bp sequences adjacent to the "inside" of each TIME flanking the rmtC region are identical (TATAA).…”