Transcriptome, proteome and draft genome of Euglena gracilis

Abstract

Background: Photosynthetic euglenids are major contributors to fresh water ecosystems. Euglena gracilis in particular has noted metabolic flexibility, reflected by an ability to thrive in a range of harsh environments. E. gracilis has been a popular model organism and of considerable biotechnological interest, but the absence of a gene catalogue has hampered both basic research and translational efforts.

Results: We report a detailed transcriptome and partial genome for E. gracilis Z1. The nuclear genome is estimated to be around 500 Mb in size, and the transcriptome encodes over 36,000 proteins and the genome possesses less than 1% coding sequence. Annotation of coding sequences indicates a highly sophisticated endomembrane system, RNA processing mechanisms and nuclear genome contributions from several photosynthetic lineages. Multiple gene families, including likely signal transduction components, have been massively expanded. Alterations in protein abundance are controlled post-transcriptionally between light and dark conditions, surprisingly similar to trypanosomatids.

Conclusions: Our data provide evidence that a range of photosynthetic eukaryotes contributed to the Euglena nuclear genome, evidence in support of the 'shopping bag' hypothesis for plastid acquisition. We also suggest that euglenids possess unique regulatory mechanisms for achieving extreme adaptability, through mechanisms of paralog expansion and gene acquisition.

Keywords: Cellular evolution; Euglena gracilis; Excavata; Gene architecture; Horizontal gene transfer; Plastid; Secondary endosymbiosis; Splicing; Transcriptome.

Fig. 1

a–d Euglena gracilis exon structure. The predicted gene structure of several selected contigs is shown, including the mapped transcripts (red), predicted splice sites and intergenic regions. Note that transcripts 524 and 326, (panel b) which encompass essentially the same portions of the genome, demonstrate possible differential exon inclusion, indicating differential open reading frame organisation and possible alternate splicing. Black boxes indicate exons, with predicted splice site dinucleotides indicated above. Transcripts are shown as arrows with the arrowhead indicating the predicted direction of transcription. Protein product annotations are indicated in parentheses. Contig sizes are shown in kilobase; note that each contig is not drawn to the same scale. Further examples of predicted contig gene organisation are given in Additional file 1: Figure S1

Fig. 2

Expression level changes induced by light are mainly post-transcriptional. Alterations to the transcriptome and proteome in response to ambient light or complete darkness were analysed using RNA-seq and SILAC/LCMS² proteomics respectively. Data are plotted for individual transcripts/polypeptides as the log₁₀ ratio between the two conditions, light (L) and dark (D), with protein on the y-axis and RNA on the x-axis. The presence of a number of proteins that were detected exclusively under one or other condition (hence infinite ratio) are indicated in green (for light) and blue (for dark). With the exception of a few transcripts, which are plastid encoded (green dots), there is little alteration to RNA abundance, but considerable changes to protein levels. Raw data for transcriptome/proteome analysis are provided in Additional file 3

Fig. 3

Euglena gracilis shares orthologs with a diverse array of lineages. Panel a (top): Histogram of E. gracilis orthologous groups clustering with selected eukaryotic lineages as determined with OrthoFinder. The x-axis shows the number of orthogroups and y-axis shows the taxon groupings representative of selected eukaryotic groups. Histogram bars highlighted in green indicate orthogroups shared with photosynthetic organisms. Panel a (lower): taxa sharing orthogroups with E. gracilis, where black circles correspond to the presence of orthogroup members while light gray circles correspond to the absence of orthogroup members in the genome. Black tie bars linking black circles are for clarity only. Eukaryotic taxon groupings are colored accordingly: gray, Euglena and kinetoplastida; white, other members of the Excavata excluding Euglenozoa; brown, SAR, pink, red algae; light green, green algae; dark green, land (vascular) plants and dark gray, Unikona. An expanded version of this figure, broken down by species is given as Additional file 1: Figure S4. Panel b: The number of E. gracilis proteins that clustered (BS > 75%) in their single-protein phylogenetic tree with taxonomic group are indicated on the x-axis

Fig. 4

Large paralog gene families are present in the Euglena gracilis genome. Several orthogroups contain many E. gracilis paralogs. The phylogenetic distribution of one large orthogroup, the nucleotidylcyclase III domain-containing proteins, is shown. Lineage groupings are colour coded: gray, all eukaryotes (and collapsed for clarity); red, N. gruberi; amber, B. saltans; and green, E. gracilis. Clades containing only Euglena sequences are boxed in green. Each sequence has been assigned a domain composition (colour gradient black to teal to blue), number of predicted trans-membrane domains (colour coded red to orange to black gradient). To obtain this phylogenetic tree, sequences with likely low coverage (less than 30% of the length of the overall alignment) were removed during alignment to avoid conflicting homology or artefact generation. Domain compositions identified are nucleotidylcyclase III, BLUF, NIT, P-loopNTPase, HAMP and Cache1

Fig. 5

Euglena gracilis has flexible and fault-tolerant mitochondrial metabolism. Proteins involved in mitochondrial pathways and complexes are shown, including: tricarboxylic acid (TCA) cycle, pyruvate dehydrogenase, fatty acid metabolism, complexes I-V of respiratory chain, ubiquinone biosynthesis, sulfate assimilation pathway, Fe-S cluster assembly and export, TIM/TOM complex and mitochondrial import machinery. Colour codes: dark blue, nucleus encoded, present in predicted mitochondrial proteome; light blue, present in transcriptome without evidence for mitochondrial localization; light blue/white, mitochondrion-encoded proteins identified previously [39]; grey, expected in nuclear transcriptome and not found; grey/white, expected in mitochondrial genome and not found. The E. gracilis mitochondrion can produce energy under both aerobic and anaerobic conditions and has workarounds for the main mitochondrial pathways, such as TCA cycle and respiratory chain, which may in part explain the outstanding adaptability of this organism

Fig. 6

The Euglena gracilis plastid possesses broad metabolic potential. Proteins involved in core plastid metabolic pathways were identified and include glycolysis/gluconeogenesis, carbon fixation, fatty acid biosynthesis, carotenoid biosynthesis, isoprenoid biosynthesis, and chlorophyll biosynthesis. Colour codes: green, nucleus encoded, present in predicted chloroplast proteome; amber, plastid encoded, present in predicted chloroplast proteome; light green/white, combination of green and amber in case of multiple subunits/isoforms; and gray, expected but not found