Transposons families/Tn554 family

From TnPedia
Revision as of 16:03, 8 April 2021 by TnCentral (talk | contribs)
Jump to navigation Jump to search


Tn554 was originally identified in Staphylococcus aureus as a determinant specifying spectinomycin resistance and inducible erythromycin (macrolide-lincosamide-streptogramin B (MLS)2) which was able to establish by transduction in a rec-deficient host [1] and was later recognised as a transposon [2]. Early genetic studies with a chromosomally located copy suggested that it might undergo site-specific integration on transduction since it appeared to be linked to specific chromosomal markers [2]. Moreover, when inserted into a plasmid, Tn554 it could be transferred alone during transduction, without the accompanying plasmid markers implying transposition from the plasmid vector into the transducing phage [3]. The observation that its transduction frequency was extremely high compared to other markers but was dramatically reduced if the recipient cell already carried a Tn554 copy in its chromosomal site led to the idea that its activity was regulated by a repressor [3,4] although it is possible that this was due simply to the occupation of the primary Tn554 integration site. It was also observed that in cases where the entire Tn554-carrying plasmid was introduced either by transduction or transformation, a copy of the transposon could often be found in the recipient chromosome, suggesting some form of zygotic induction [5] occurred from the plasmid when introduced into naïve cells [6]. Southern hybridization of restriction digested genomic DNA from 15 different Staphylococcus strains from a number of sources using radioactive labelled Tn554 DNA clearly revealed the presence of a Tn554 copy at the same location, and in one case, an additional copy was present [6]. Site-specific insertion was confirmed using a plasmid target with a clone site [4] and using this system, it was also shown that insertions could occur into secondary sites at a lower frequency. Electron microscopy heteroduplex (see General Information/IS History) analysis between plasmids with and without a Tn554 insertion revealed a single strand Tn554 insertion loop. The fact that when introduced into a second strain, the Tn554-carrying plasmids did not change (i.e. retained the transposon) while Tn544 transposition into the chromosome had occurred, suggested that transposition in this case may be replicative [4]. However, this is certainly not a formal demonstration of replicative transposition, especially in view of the possibility that transposition appears to occur by site-specific excision of a circular transposon copy and subsequent site- specific integration. This must be revisited using more powerful biochemical approaches. Transposition of Tn554 from secondary insertion sites in a number of plasmids occurred exclusively into the chromosomal site but at quite variable frequencies depending on the plasmid and site.

The location of the Tn554 antibiotic resistance genes to the right end of the transposon was identified by restriction fragment cloning while a possible transposition repressor tnpI was identified by its ability to inhibit transposition of an incoming Tn554 copy and localised to the left end [7]. Although the nature of tnpI remains obscure, it was shown that it resides in the left end within the first 89 bp and can act in trans leading to the proposal that this region may contain a binding site which titrates one of the transposition proteins [7]. Moreover, the nucleotide sequences of the Tn554 junctions (three 'primary' and two 'secondary' insertions) revealed that Tn554 does not include terminal inverted repeats (IR) at its ends neither does it generate direct target repeats (DR) typical of many other transposons and insertion sequences [8]. Insertion occurred specifically between two A residues at the primary insertion site with the transposon in the same orientation [8].

The entire Tn554 nucleotide sequence, 6,691 bp [9], revealed six open reading frames named from left to right (Fig. Tn554.1A), tnpA, tnpB, tnpC, three transposition-related genes, the antibiotic resistance genes: the spc determinant, an adenyltransferase, AAD(9), that modifies spectinomycin but not streptomycin and ermA, followed by ermB [9] (Fig. Tn554.1A).