Table 5_Detection and functional analysis of horizontal gene transfer events in the ciliate Euplotes.docx

BackgroundHorizontal gene transfer (HGT), the movement of heritable materials between distantly related organisms, is a key evolutionary force shaping eukaryotic genomes. Euplotes are free-living unicellular eukaryotes belonging to the phylum Ciliophora, and are tended to establish endosymbiotic relationships with different bacteria. However, the scale of HGT in Euplotes, and its possible roles in driving their diversification and adaptation remains unexplored.
MethodsA large-scale phylogeny-based bacterial HGT detection was performed across five genome sequenced Euplotes. Gene structure and expression of the HGT-acquired genes were analyzed based on the transcriptome data. Putative functions of these genes were annotated based on BLAST search in the protein family (Pfam), the Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Quantitative polymerase chain reaction (qPCR) and RNA interference (RNAi) were performed to validate the function of the prevalent HGT-acquired genes encoding mannan endo-1,4-β-mannosidase (Man) from E. amieti.
ResultsWe systematically examined HGT in five Euplotes genomes and found that they acquired a total of 342 genes exhibiting diverse functions, including enzymes involved in carbohydrate metabolism, sulfur metabolism, and the cell signaling. HGT-acquired genes displayed similar genomic features with the native genes, including GC content, the proportion of intron-contained gene, and coding sequences (CDS) length, implying ancient acquisition events. Five putative endosymbiont-derived genes encoding glycoside hydrolases from E. vannus were identified. Furthermore, among the 342 HGT candidates, only seven HGT families were putatively transferred into the last common ancestor of all five Euplotes. Further qPCR analysis showed that the mRNA levels of mannan endo-1,4-β-mannosidase A (Ea-ManA) and mannan endo-1,4-β-mannosidase B (Ea-ManB) increased after feeding with Chlorogonium elongatum in E. amieti. Knockdown of Ea-Man genes by RNAi increased mortality which suggested that Ea-Man genes are essential for E. amieti.
ConclusionBased on these findings, we suggest that the endosymbionts of Euplotes are potential donor organisms for HGT-acquired genes, and HGT is a prevalent mechanism that is actively used in Euplotes to expand their adaptive capabilities.