Euglenids represent a group of protists with diverse modes of feeding. To date, only a partial genomic sequence of Euglena gracilis and transcriptomes of several phototrophic and secondarily osmotrophic species are available, while primarily heterotrophic euglenids are seriously undersampled. In this work, we begin to fill this gap by presenting genomic and transcriptomic drafts of a primary osmotroph, Rhabdomonas costata. The current genomic assembly length of 100 Mbp is 14× smaller than that of E. gracilis. Despite being too fragmented for comprehensive gene prediction it provided fragments of the mitochondrial genome and comparison of the transcriptomic and genomic data revealed features of its introns, including several candidates for nonconventional types. A set of 39,456 putative R. costata proteins was predicted from the transcriptome. Annotation of the mitochondrial core metabolism provides the first data on the facultatively anaerobic mitochondrion of R. costata, which in most respects resembles the mitochondrion of E. gracilis with a certain level of streamlining. R. costata can synthetise thiamine by enzymes of heterogenous provenances and haem by a mitochondrial-cytoplasmic C4 pathway with enzymes orthologous to those found in E. gracilis. The low percentage of green algae-affiliated genes supports the ancestrally osmotrophic status of this species.

• MeSH
• biologická evoluce MeSH
• Chromatium genetika   metabolismus   MeSH
• Euglenida genetika   metabolismus   MeSH
• exony genetika   MeSH
• fylogeneze MeSH
• genom MeSH
• heterotrofní procesy MeSH
• introny genetika   MeSH
• mitochondrie genetika   MeSH
• sekvenční analýza DNA metody   MeSH
• transkriptom genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: The supergroup Euglenozoa unites heterotrophic flagellates from three major clades, kinetoplastids, diplonemids, and euglenids, each of which exhibits extremely divergent mitochondrial characteristics. Mitochondrial genomes (mtDNAs) of euglenids comprise multiple linear chromosomes carrying single genes, whereas mitochondrial chromosomes are circular non-catenated in diplonemids, but circular and catenated in kinetoplastids. In diplonemids and kinetoplastids, mitochondrial mRNAs require extensive and diverse editing and/or trans-splicing to produce mature transcripts. All known euglenozoan mtDNAs exhibit extremely short mitochondrial small (rns) and large (rnl) subunit rRNA genes, and absence of tRNA genes. How these features evolved from an ancestral bacteria-like circular mitochondrial genome remains unanswered. RESULTS: We sequenced and assembled 20 euglenozoan single-cell amplified genomes (SAGs). In our phylogenetic and phylogenomic analyses, three SAGs were placed within kinetoplastids, 14 within diplonemids, one (EU2) within euglenids, and two SAGs with nearly identical small subunit rRNA gene (18S) sequences (EU17/18) branched as either a basal lineage of euglenids, or as a sister to all euglenozoans. Near-complete mitochondrial genomes were identified in EU2 and EU17/18. Surprisingly, both EU2 and EU17/18 mitochondrial contigs contained multiple genes and one tRNA gene. Furthermore, EU17/18 mtDNA possessed several features unique among euglenozoans including full-length rns and rnl genes, six mitoribosomal genes, and nad11, all likely on a single chromosome. CONCLUSIONS: Our data strongly suggest that EU17/18 is an early-branching euglenozoan with numerous ancestral mitochondrial features. Collectively these data contribute to untangling the early evolution of euglenozoan mitochondria.

• Klíčová slova
• Evolution, Mitochondrial ribosome, Phylogeny, Single-cell amplified genome,
• MeSH
• Euglenida * genetika   MeSH
• Euglenozoa genetika   MeSH
• europium MeSH
• fylogeneze MeSH
• genom mitochondriální * genetika   MeSH
• genomika MeSH
• mitochondriální DNA MeSH
• RNA transferová MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• europium MeSH
• mitochondriální DNA MeSH
• RNA transferová MeSH

Euglenozoa is a species-rich group of protists, which have extremely diverse lifestyles and a range of features that distinguish them from other eukaryotes. They are composed of free-living and parasitic kinetoplastids, mostly free-living diplonemids, heterotrophic and photosynthetic euglenids, as well as deep-sea symbiontids. Although they form a well-supported monophyletic group, these morphologically rather distinct groups are almost never treated together in a comparative manner, as attempted here. We present an updated taxonomy, complemented by photos of representative species, with notes on diversity, distribution and biology of euglenozoans. For kinetoplastids, we propose a significantly modified taxonomy that reflects the latest findings. Finally, we summarize what is known about viruses infecting euglenozoans, as well as their relationships with ecto- and endosymbiotic bacteria.

• Klíčová slova
• Diplonemida, Euglenida, Kinetoplastida, microbial eukaryotes, phylogeny, systematics,
• MeSH
• ekosystém MeSH
• Euglenozoa klasifikace   genetika   fyziologie   virologie   MeSH
• fylogeneze MeSH
• Mimiviridae patogenita   MeSH
• symbióza MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

Euglena gracilis is a photosynthetic flagellate possessing chlorophyte-derived secondary plastids that are enclosed by only three enveloping membranes, unlike most secondary plastids, which are surrounded by four membranes. It has generally been assumed that the two innermost E. gracilis plastid envelopes originated from the primary plastid, while the outermost is of eukaryotic origin. It was suggested that nucleus-encoded plastid proteins pass through the middle and innermost plastid envelopes of E. gracilis by machinery homologous to the translocons of outer and inner chloroplast membranes, respectively. Although recent genomic, transcriptomic, and proteomic data proved the presence of a reduced form of the translocon of inner membrane, they failed to identify any outer-membrane translocon homologs, which raised the question of the origin of E. gracilis's middle plastid envelope. Here, we compared the lipid composition of whole cells of the pigmented E. gracilis strain Z and two bleached mutants that lack detectable plastid structures, W10BSmL and WgmZOflL We determined the lipid composition of E. gracilis strain Z mitochondria and plastids, and of plastid subfractions (thylakoids and envelopes), using HPLC high-resolution tandem mass spectrometry, thin-layer chromatography, and gas chromatography-flame ionization detection analytical techniques. Phosphoglycerolipids are the main structural lipids in mitochondria, while glycosyldiacylglycerols are the major structural lipids of plastids and also predominate in extracts of whole mixotrophic cells. Glycosyldiacylglycerols were detected in both bleached mutants, indicating that mutant cells retain some plastid remnants. Additionally, we discuss the origin of the E. gracilis middle plastid envelope based on the lipid composition of envelope fraction.

• MeSH
• buněčná membrána chemie   MeSH
• chloroplasty chemie   MeSH
• Euglena gracilis chemie   MeSH
• genetická variace MeSH
• genotyp MeSH
• lipidy chemie   MeSH
• mutace MeSH
• plastidy chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• lipidy MeSH

Euglena gracilis is a metabolically flexible, photosynthetic, and adaptable free-living protist of considerable environmental importance and biotechnological value. By label-free liquid chromatography tandem mass spectrometry, a total of 1,786 proteins were identified from the E. gracilis purified mitochondria, representing one of the largest mitochondrial proteomes so far described. Despite this apparent complexity, protein machinery responsible for the extensive RNA editing, splicing, and processing in the sister clades diplonemids and kinetoplastids is absent. This strongly suggests that the complex mechanisms of mitochondrial gene expression in diplonemids and kinetoplastids occurred late in euglenozoan evolution, arising independently. By contrast, the alternative oxidase pathway and numerous ribosomal subunits presumed to be specific for parasitic trypanosomes are present in E. gracilis. We investigated the evolution of unexplored protein families, including import complexes, cristae formation proteins, and translation termination factors, as well as canonical and unique metabolic pathways. We additionally compare this mitoproteome with the transcriptome of Eutreptiella gymnastica, illuminating conserved features of Euglenida mitochondria as well as those exclusive to E. gracilis. This is the first mitochondrial proteome of a free-living protist from the Excavata and one of few available for protists as a whole. This study alters our views of the evolution of the mitochondrion and indicates early emergence of complexity within euglenozoan mitochondria, independent of parasitism.

• Klíčová slova
• Euglena gracilis, Euglenozoa, mitochondria, proteome, protist,
• MeSH
• Euglena gracilis metabolismus   MeSH
• mitochondriální proteiny metabolismus   MeSH
• proteom * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• mitochondriální proteiny MeSH
• proteom * MeSH

The order Trypanosomatida has been well studied due to its pathogenicity and the unique biology of the mitochondrion. In Trypanosoma brucei, four DNA polymerases, namely PolIA, PolIB, PolIC, and PolID, related to bacterial DNA polymerase I (PolI), were shown to be localized in mitochondria experimentally. These mitochondrion-localized DNA polymerases are phylogenetically distinct from other family A DNA polymerases, such as bacterial PolI, DNA polymerase gamma (Polγ) in human and yeasts, "plant and protist organellar DNA polymerase (POP)" in diverse eukaryotes. However, the diversity of mitochondrion-localized DNA polymerases in Euglenozoa other than Trypanosomatida is poorly understood. In this study, we discovered putative mitochondrion-localized DNA polymerases in broad members of three major classes of Euglenozoa-Kinetoplastea, Diplonemea, and Euglenida-to explore the origin and evolution of trypanosomatid PolIA-D. We unveiled distinct inventories of mitochondrion-localized DNA polymerases in the three classes: (1) PolIA is ubiquitous across the three euglenozoan classes, (2) PolIB, C, and D are restricted in kinetoplastids, (3) new types of mitochondrion-localized DNA polymerases were identified in a prokinetoplastid and diplonemids, and (4) evolutionarily distinct types of POP were found in euglenids. We finally propose scenarios to explain the inventories of mitochondrion-localized DNA polymerases in Kinetoplastea, Diplonemea, and Euglenida.

• Klíčová slova
• DNA replication, Diplonemea, Euglenida, Kinetoplastea, Prokinetoplastina, Trypanosomatida, family A DNA polymerase, plant and protist organellar DNA polymerase,
• Publikační typ
• časopisecké články MeSH

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.

• Klíčová slova
• Cellular evolution, Euglena gracilis, Excavata, Gene architecture, Horizontal gene transfer, Plastid, Secondary endosymbiosis, Splicing, Transcriptome,
• MeSH
• buněčné jádro MeSH
• Euglena gracilis genetika   metabolismus   MeSH
• genom * MeSH
• plastidy MeSH
• proteom * MeSH
• transkriptom * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• proteom * MeSH

The mitochondrial DNA of diplonemid and kinetoplastid protists is known for its suite of bizarre features, including the presence of concatenated circular molecules, extensive trans-splicing and various forms of RNA editing. Here we report on the existence of another remarkable characteristic: hyper-inflated DNA content. We estimated the total amount of mitochondrial DNA in four kinetoplastid species (Trypanosoma brucei, Trypanoplasma borreli, Cryptobia helicis, and Perkinsela sp.) and the diplonemid Diplonema papillatum. Staining with 4',6-diamidino-2-phenylindole and RedDot1 followed by color deconvolution and quantification revealed massive inflation in the total amount of DNA in their organelles. This was further confirmed by electron microscopy. The most extreme case is the ∼260 Mbp of DNA in the mitochondrion of Diplonema, which greatly exceeds that in its nucleus; this is, to our knowledge, the largest amount of DNA described in any organelle. Perkinsela sp. has a total mitochondrial DNA content ~6.6× greater than its nuclear genome. This mass of DNA occupies most of the volume of the Perkinsela cell, despite the fact that it contains only six protein-coding genes. Why so much DNA? We propose that these bloated mitochondrial DNAs accumulated by a ratchet-like process. Despite their excessive nature, the synthesis and maintenance of these mtDNAs must incur a relatively low cost, considering that diplonemids are one of the most ubiquitous and speciose protist groups in the ocean. © 2018 IUBMB Life, 70(12):1267-1274, 2018.

• Klíčová slova
• DNA content, kinetoplast DNA, mitochondrial DNA, protist,
• MeSH
• Euglenozoa genetika   MeSH
• fylogeneze MeSH
• Kinetoplastida genetika   MeSH
• mitochondriální DNA genetika   izolace a purifikace   ultrastruktura   MeSH
• mitochondrie genetika   MeSH
• trans-splicing genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• mitochondriální DNA MeSH

Mitochondria are double membrane organelles of endosymbiotic origin, best known for constituting the centre of energetics of a eukaryotic cell. They contain their own mitochondrial genome, which as a consequence of gradual reduction during evolution typically contains less than two dozens of genes. In this review, we highlight the extremely diverse architecture of mitochondrial genomes and mechanisms of gene expression between the three sister groups constituting the phylum Euglenozoa - Euglenida, Diplonemea and Kinetoplastea. The earliest diverging euglenids possess a simplified mitochondrial genome and a conventional gene expression, whereas both are highly complex in the two other groups. The expression of their mitochondrial-encoded proteins requires extensive post-transcriptional modifications guided by complex protein machineries and multiple small RNA molecules. Moreover, the least studied diplonemids, which have been recently discovered as a highly abundant component of the world ocean plankton, possess one of the most complicated mitochondrial genome organisations known to date.

• Klíčová slova
• euglenozoa, mitochondria, mitochondrial genome,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH