Figure 1. Novel domesticated PiggyBac transposases in Paramecium.

(A) Domain organization of the PiggyBac transposase (PB) from T. ni and of Paramecium PiggyBac-related proteins (Pgm and PgmLs). The Pfam domain DDE_Tnp_1_7 is shown as a bipartite orange domain, with the RNase H fold corresponding to its right part (conserved catalytic D residues are indicated by vertical bars). The DDE_Tnp_1-like zinc ribbon is in grey. Id: % of amino acid identity; sim: % of similarity. (B) Protein sequence alignment of the residues surrounding the three catalytic aspartic acids (DDD). Following secondary structure prediction, sequence alignments were adjusted manually, using the expected position of the three catalytic D residues in the first and fourth β strands and immediately downstream of the fourth α helix of the RNase H fold domain, respectively (Hickman et al., 2010). ‘?' indicates that the expected α4 helix could not be predicted using the PSIPRED secondary structure prediction software.

Figure 1—figure supplement 1. MUSCLE alignment of the cysteine-rich domains of ciliate domesticated PB transposases and other PB transposases.

The analysis involved 62 amino acid sequences of PB transposases and domesticated transposases from ciliates and other species. Amino acid sequences encompassing the cysteine-rich domain of each protein were aligned using MUSCLE (http://www.ebi.ac.uk/Tools/msa/muscle/). All sequences used for the alignment are displayed in Supplementary file 3. Complete accession numbers can be found in Figure 1—figure supplement 2.

Figure 1—figure supplement 2. Maximum Likelihood tree of ciliate domesticated PB transposases and other PB transposases.

The tree includes 69 amino acid sequences of PB transposases and domesticated PB transposases from ciliates and other species. To construct the tree, the alignment of all transposase core domains (Supplementary file 1) was edited to remove specific insertions restricted to one particular PgmL family. All accession numbers are indicated, except for the PiggyBat transposase from Myotis lucifugus (Mitra et al., 2013). PB and domesticated PB proteins from Tetrahymena thermophila are in blue. The evolutionary history was inferred by using the Maximum Likelihood method based on the JTT matrix-based model (Jones et al., 1992). The tree with the highest log likelihood (−35952.17) is shown. The percentage of trees in which the associated taxa clustered together is shown next to the branches (bootstrap = 100). Initial tree(s) for the heuristic search were obtained automatically by applying Neighbor-Join and BioNJ algorithms to a matrix of pairwise distances estimated using a JTT model, and then selecting the topology with superior log likelihood value. A discrete Gamma distribution was used to model evolutionary rate differences among sites (five categories (+G, parameter = 2.8098)). There were a total of 541 positions in the final dataset. The tree is drawn to scale, with branch lengths measured as the number of substitutions per site. Evolutionary analyses were conducted in MEGA7 (Kumar et al., 2016).