A family of diatom-like silicon transporters in the siliceous loricate choanoflagellates

Abstract

Biosilicification is widespread across the eukaryotes and requires concentration of silicon in intracellular vesicles. Knowledge of the molecular mechanisms underlying this process remains limited, with unrelated silicon-transporting proteins found in the eukaryotic clades previously studied. Here, we report the identification of silicon transporter (SIT)-type genes from the siliceous loricate choanoflagellates Stephanoeca diplocostata and Diaphanoeca grandis. Until now, the SIT gene family has been identified only in diatoms and other siliceous stramenopiles, which are distantly related to choanoflagellates among the eukaryotes. This is the first evidence of similarity between SITs from different eukaryotic supergroups. Phylogenetic analysis indicates that choanoflagellate and stramenopile SITs form distinct monophyletic groups. The absence of putative SIT genes in any other eukaryotic groups, including non-siliceous choanoflagellates, leads us to propose that SIT genes underwent a lateral gene transfer event between stramenopiles and loricate choanoflagellates. We suggest that the incorporation of a foreign SIT gene into the stramenopile or choanoflagellate genome resulted in a major metabolic change: the acquisition of biomineralized silica structures. This hypothesis implies that biosilicification has evolved multiple times independently in the eukaryotes, and paves the way for a better understanding of the biochemical basis of silicon transport through identification of conserved sequence motifs.

Figure 1.

The loricate choanoflagellates (a,b) S. diplocostata and (c,d) D. grandis. Photographs taken using phase contrast at 100× magnification. Schematic figures are taken from the Micro*scope v. 6.0 website (http://starcentral.mbl.edu/microscope, drawings by Won-Je Lee) and used under a Creative Commons licence. Lor, Lorica; CB, cell body; Fl, flagellum; Col, collar. Bacteria in the photographs are annotated with asterisks (*). Scale bar, 5 μm. (Online version in colour.)

Figure 2.

Alignment and identification of conserved residues and TM domains in stramenopile and choanoflagellate SITs. The black line is the TMPred SdSIT topology prediction. The orange line is the diatom SIT TMD prediction [30]. Red boxes indicate conserved motifs of three or more residues. The double blue circles mark the ‘CMLD’ motif in stramenopiles. Conserved charged or hydroxylated residues are noted as per [13,30]; purple arrows, negatively charged; red arrows, positively charged; green arrows, hydroxyl-containing. Oo, Ochromonas ovalis (chrysophyte, non-diatom stramenopile); Cf, Cylindrotheca fusiformis (pennate diatom); Pt, Phaeodactylum tricornutum (pennate diatom); Tp, Thalassiosira pseudonana (centric diatom); Sa, Synedra acus (pennate diatom).

Figure 3.

Phylogenetic tree demonstrates that choanoflagellate SITs are monophyletic within the stramenopile SITs. The tree was produced using PhyML ML analysis with the LG + Γ model from an alignment of 769 amino acid residues. The tree produced using RaxML and the WAG model had identical topology. Numbers at nodes are a percentage of 100 bootstrap replicates (PhyML bootstrap value/RaxML bootstrap value). Blue, pennate diatom; green, centric diatom; brown, non-diatom stramenopile; red, choanoflagellate. Dash (−/−) indicates a bootstrap value less than or equal to 70%. Scale bar indicates the average number of amino acid substitutions per site.

Figure 4.

Schematic of the SdSIT protein showing transmembrane structure and orientations of conserved motifs and residues. Conservation was determined from the choanoflagellate and stramenopile SIT alignment (see figure 2 and electronic supplementary material). TMDs (grey cylinders) are taken from the SdSITa TMPred topology and numbered accordingly. Conserved motifs are in boxes, circles are conserved residues as per figure 2: red, positively charged; purple, negatively charged; green, hydroxyl group containing.