Diplomonads are anaerobic, flagellated protists, being part of the Metamonada group of Eukaryotes. Diplomonads either live as endobionts (parasites and commensals) of animals or free-living in low-oxygen environments. Genomic information is available for parasitic diplomonads like Giardia intestinalis and Spironucleus salmonicida, while little is known about the genomic arrangements of free-living diplomonads. We have generated the first reference genome of a free-living diplomonad, Hexamita inflata. The final version of the genome assembly is fragmented (1241 contigs) but substantially larger (142 Mbp) than the parasitic diplomonad genomes (9.8-14.7 Mbp). It encodes 79,341 proteins; 29,874 have functional annotations and 49,467 are hypothetical proteins. Interspersed repeats comprise 34% of the genome (9617 Retroelements, 2676 DNA transposons). The large expansion of protein-encoding capacity and the interspersed repeats are the major reasons for the large genome size. This genome from a free-living diplomonad will be the basis for further studies of the Diplomonadida lineage and the evolution of parasitism-free living style transitions.

• MeSH
• Diplomonadida * genetika   MeSH
• genom protozoální * MeSH
• retroelementy MeSH
• rozptýlené repetitivní sekvence MeSH
• Publikační typ
• časopisecké články MeSH
• dataset MeSH
• Názvy látek
• retroelementy MeSH

BACKGROUND: The endoplasmic reticulum (ER)-mitochondria membrane contact sites (MCS) are extensively studied in aerobic eukaryotes; however, little is known about MCS in anaerobes with reduced forms of mitochondria named hydrogenosomes. In several eukaryotic lineages, the direct physical tether between ER and the outer mitochondrial membrane is formed by ER-mitochondria encounter structure (ERMES). The complex consists of four core proteins (Mmm1, Mmm2, Mdm12, and Mdm10) which are involved in phospholipid trafficking. Here we investigated ERMES distribution in organisms bearing hydrogenosomes and employed Trichomonas vaginalis as a model to estimate ERMES cellular localization, structure, and function. RESULTS: Homology searches revealed that Parabasalia-Anaeramoebae, anaerobic jakobids, and anaerobic fungi are lineages with hydrogenosomes that retain ERMES, while ERMES components were gradually lost in Fornicata, and are absent in Preaxostyla and Archamoebae. In T. vaginalis and other parabasalids, three ERMES components were found with the expansion of Mmm1. Immunofluorescence microscopy confirmed that Mmm1 localized in ER, while Mdm12 and Mmm2 were partially localized in hydrogenosomes. Pull-down assays and mass spectrometry of the ERMES components identified a parabasalid-specific Porin2 as a substitute for the Mdm10. ERMES modeling predicted a formation of a continuous hydrophobic tunnel of TvMmm1-TvMdm12-TvMmm2 that is anchored via Porin2 to the hydrogenosomal outer membrane. Phospholipid-ERMES docking and Mdm12-phospholipid dot-blot indicated that ERMES is involved in the transport of phosphatidylinositol phosphates. The absence of enzymes involved in hydrogenosomal phospholipid metabolism implies that ERMES is not involved in the exchange of substrates between ER and hydrogenosomes but in the unidirectional import of phospholipids into hydrogenosomal membranes. CONCLUSIONS: Our investigation demonstrated that ERMES mediates ER-hydrogenosome interactions in parabasalid T. vaginalis, while the complex was lost in several other lineages with hydrogenosomes.

• Klíčová slova
• Anaerobiosis, Cardiolipin, ERMES, Endoplasmic reticulum, Hydrogenosomes, Structure, Trichomonas vaginalis,
• MeSH
• anaerobióza MeSH
• endoplazmatické retikulum * metabolismus   MeSH
• fosfolipidy metabolismus   MeSH
• membránové proteiny * metabolismus   MeSH
• mitochondrie metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fosfolipidy MeSH
• membránové proteiny * MeSH

Trichomonas vaginalis is a parasitic protist that infects the human urogenital tract. During the infection, trichomonads adhere to the host mucosa, acquire nutrients from the vaginal/prostate environment, and release small extracellular vesicles (sEVs) that contribute to the trichomonad adherence and modulate the host-parasite communication. Approximately 40-70% of T. vaginalis strains harbor a double-stranded RNA virus called Trichomonasvirus (TVV). Naked TVV particles have the potential to stimulate a proinflammatory response in human cells, however, the mode of TVV release from trichomonads to the environment is not clear. In this report, we showed for the first time that TVV particles are released from T. vaginalis cells within sEVs. The sEVs loaded with TVV stimulated a higher proinflammatory response of human HaCaT cells in comparison to sEVs from TVV negative parasites. Moreover, a comparison of T. vaginalis isogenic TVV plus and TVV minus clones revealed a significant impact of TVV infection on the sEV proteome and RNA cargo. Small EVs from TVV positive trichomonads contained 12 enriched and 8 unique proteins including membrane-associated BspA adhesine, and about a 2.5-fold increase in the content of small regulatory tsRNA. As T. vaginalis isolates are frequently infected with TVV, the release of TVV via sEVs to the environment represents an important factor with the potential to enhance inflammation-related pathogenesis during trichomoniasis.

• Klíčová slova
• TVV, Trichomonasvirus, exosome, extracellular vesicle, proteomics, tsRNA,
• Publikační typ
• časopisecké články MeSH