Spy: a new group of eukaryotic DNA transposons without target site duplications

Abstract

Class 2 or DNA transposons populate the genomes of most eukaryotes and like other mobile genetic elements have a profound impact on genome evolution. Most DNA transposons belong to the cut-and-paste types, which are relatively simple elements characterized by terminal-inverted repeats (TIRs) flanking a single gene encoding a transposase. All eukaryotic cut-and-paste transposons so far described are also characterized by target site duplications (TSDs) of host DNA generated upon chromosomal insertion. Here, we report a new group of evolutionarily related DNA transposons called Spy, which also include TIRs and DDE motif-containing transposase but surprisingly do not create TSDs upon insertion. Instead, Spy transposons appear to transpose precisely between 5'-AAA and TTT-3' host nucleotides, without duplication or modification of the AAATTT target sites. Spy transposons were identified in the genomes of diverse invertebrate species based on transposase homology searches and structure-based approaches. Phylogenetic analyses indicate that Spy transposases are distantly related to IS5, ISL2EU, and PIF/Harbinger transposases. However, Spy transposons are distinct from these and other DNA transposon superfamilies by their lack of TSD and their target site preference. Our findings expand the known diversity of DNA transposons and reveal a new group of eukaryotic DDE transposases with unusual catalytic properties.

Keywords: DNA transposon; Spy; target site duplication; transposition.

Fig. 1.—

Characters of silkworm Spy transposons. (A) Sequence alignments for Spy-6_BMo family. The TIRs and flanking sequences are shown. (B) Seven examples of alignments of the flanking sequences of Spy insertions with a paralogous sequences found within the same genome but devoid of the transposon. The TIRs of the element is underlined. (C) Structure of Spy-6_BMo. Black triangles represent the TIRs. ORFs are depicted as solid black boxes and the position of the DDE triad is shown.

Fig. 2.—

Target sites of the Spy family were tested using PCR and sequencing. (A) Results of PCR search for indel silkworm strains (DaZao, BiBo, and HeiGao) among four insertion sites of four silkworm Spy families (Spy-2, -4, -6, and -7_BMo). The black arrow points to the corresponding Spy lack at this genomic location. The white arrow represents the corresponding Spy occupied at this genomic location. (B) Results of sequencing for above locations. The target sequences are marked with black bold font, TIRs are marked with underline, and flank sequences are in blank.

Fig. 3.—

Taxonomic distribution of the Spy, PIF/Harbinger, and ISL2EU transposons as well as character of Spy TIRs. (A) Taxonomic distribution of three groups across the eukaryotic tree of life. Black and white boxes indicated presence and absence, respectively. (B) TIRs of all Spy identified in this study. Sequences are major-rule consensus derived from the alignment of multiple copies of each family.

Fig. 4.—

The results of phylogenetic analysis, coding capacity, and conserved transposase motifs of IS5, Spy, ISL2EU, and PIF/Harbinger. (A) The alignment of DDE domain of each superfamily after redundancy elimination. Distances between the conserved blocks are indicated in the number of amino acid residues. Conserved residues within each superfamily are highlight in black or gray. The DDE triad identified here is marked with asterisks below alignments. Common conserved residues among four superfamilies are marked with letter above the alignments. Unique conserved residues of each superfamily are marked with blank triangle below the alignment. (B) Phylogenetic tree based on DDE domain sequences of each superfamily. In front of the colon represents corresponding IS5, Spy, ISL2EU, or PIF/Harbinger elements name; behind the colon represents species. (C) Structure of each superfamily. Black triangles represent the TIRs. ORFs are depicted as solid black boxes, and the position of the DDE triad and additional domains is shown above. Target sequences are shown in flank.

Fig. 5.—

The secondary structure of DDE domain-containing transposase for Spy, ISL2EU, and PIF/Harbinger elements. The DDE triad is marked with red triangles below sequence.