Dynamins and dynamin-like proteins (DLPs) belong to a family of large GTPases involved in membrane remodelling events. These include both fusion and fission processes with different dynamin proteins often having a specialised function within the same organism. Trypanosoma brucei is thought to have only one multifunctional DLP (TbDLP). While this was initially reported to function in mitochondrial division only, an additional role in endocytosis and cytokinesis was later also proposed. Since there are two copies of TbDLP present in the trypanosome genome, we investigated potential functional differences between these two paralogs by re-expressing either protein in a TbDLP RNAi background. These paralogs, called TbDLP1 and TbDLP2, are almost identical bar a few amino acid substitutions. Our results, based on cell lines carrying tagged and RNAi-resistant versions of each protein, show that overexpression of TbDLP1 alone is able to rescue the observed endocytosis and growth defects in the mammalian bloodstream form (BSF) of the parasite. While TbDLP2 shows no rescue in our experiments in BSF, this might also be due to lower expression levels of the protein in this life stage. In contrast, both TbDLP proteins apparently play more complementary roles in the insect procyclic form (PCF) since neither TbDLP1 nor TbDLP2 alone can fully restore wildtype growth and morphology in TbDLP-depleted parasites.
- MeSH
- buněčné linie MeSH
- dynaminy chemie genetika metabolismus MeSH
- protozoální proteiny chemie genetika metabolismus MeSH
- RNA interference MeSH
- sekvence aminokyselin MeSH
- sekvenční homologie aminokyselin MeSH
- Trypanosoma brucei brucei metabolismus MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- dynaminy MeSH
- protozoální proteiny MeSH
BACKGROUND: Kinetoplastea is a diverse protist lineage composed of several of the most successful parasites on Earth, organisms whose metabolisms have coevolved with those of the organisms they infect. Parasitic kinetoplastids have emerged from free-living, non-pathogenic ancestors on multiple occasions during the evolutionary history of the group. Interestingly, in both parasitic and free-living kinetoplastids, the heme pathway-a core metabolic pathway in a wide range of organisms-is incomplete or entirely absent. Indeed, Kinetoplastea investigated thus far seem to bypass the need for heme biosynthesis by acquiring heme or intermediate metabolites directly from their environment. RESULTS: Here we report the existence of a near-complete heme biosynthetic pathway in Perkinsela spp., kinetoplastids that live as obligate endosymbionts inside amoebozoans belonging to the genus Paramoeba/Neoparamoeba. We also use phylogenetic analysis to infer the evolution of the heme pathway in Kinetoplastea. CONCLUSION: We show that Perkinsela spp. is a deep-branching kinetoplastid lineage, and that lateral gene transfer has played a role in the evolution of heme biosynthesis in Perkinsela spp. and other Kinetoplastea. We also discuss the significance of the presence of seven of eight heme pathway genes in the Perkinsela genome as it relates to its endosymbiotic relationship with Paramoeba.
- Klíčová slova
- Endosymbiosis, Evolution, Heme, Kinetoplastea, Lateral gene transfer, Paramoeba pemaquidensis, Perkinsela, Prokinetoplastina,
- MeSH
- biologická evoluce MeSH
- Eukaryota klasifikace fyziologie MeSH
- fylogeneze MeSH
- hem metabolismus MeSH
- Kinetoplastida klasifikace genetika fyziologie MeSH
- přenos genů horizontální MeSH
- symbióza MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- hem MeSH
In our previous work we established a T7 polymerase-driven Tetracycline-inducible protein expression system in Leishmania mexicana (Biagi, 1953). We used this system to analyse gene expression profiles during development of L. mexicana in procyclic and metacyclic promastigotes and amastigotes. The transcription of the gene of interest and the T7 polymerase genes was significantly reduced upon cell differentiation. This regulation is not locus-specific. It depends on untranslated regions flanking open reading frames of the genes analysed. In this paper, we report that the previously established conventional inducible protein expression system may not be suitable for studies on differentiation of species of Leishmania Ross, 1903 and protein expression systems might have certain limitations.
- Klíčová slova
- Leishmania mexicana, Tet-inducible system, gene expression, untranslated regions,
- MeSH
- DNA-dependentní DNA-polymerasy metabolismus MeSH
- Leishmania mexicana enzymologie genetika MeSH
- regulace genové exprese * MeSH
- stadia vývoje genetika MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- DNA-dependentní DNA-polymerasy MeSH
- MeSH
- antibakteriální látky toxicita MeSH
- antibiotická rezistence fyziologie MeSH
- mitochondrie účinky léků MeSH
- proteosyntéza účinky léků MeSH
- tetracyklin toxicita MeSH
- Trypanosoma účinky léků fyziologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
- Názvy látek
- antibakteriální látky MeSH
- tetracyklin MeSH
BACKGROUND: The haemoflagellate Trypanosoma lewisi is a kinetoplastid parasite which, as it has been recently reported to cause human disease, deserves increased attention. Characteristic features of all kinetoplastid flagellates are a uniquely structured mitochondrial DNA or kinetoplast, comprised of a network of catenated DNA circles, and RNA editing of mitochondrial transcripts. The aim of this study was to describe the kinetoplast DNA of T. lewisi. METHODS/RESULTS: In this study, purified kinetoplast DNA from T. lewisi was sequenced using high-throughput sequencing in combination with sequencing of PCR amplicons. This allowed the assembly of the T. lewisi kinetoplast maxicircle DNA, which is a homologue of the mitochondrial genome in other eukaryotes. The assembly of 23,745 bp comprises the non-coding and coding regions. Comparative analysis of the maxicircle sequence of T. lewisi with Trypanosoma cruzi, Trypanosoma rangeli, Trypanosoma brucei and Leishmania tarentolae revealed that it shares 78%, 77%, 74% and 66% sequence identity with these parasites, respectively. The high GC content in at least 9 maxicircle genes of T. lewisi (ATPase6; NADH dehydrogenase subunits ND3, ND7, ND8 and ND9; G-rich regions GR3 and GR4; cytochrome oxidase subunit COIII and ribosomal protein RPS12) implies that their products may be extensively edited. A detailed analysis of the non-coding region revealed that it contains numerous repeat motifs and palindromes. CONCLUSIONS: We have sequenced and comprehensively annotated the kinetoplast maxicircle of T. lewisi. Our analysis reveals that T. lewisi is closely related to T. cruzi and T. brucei, and may share similar RNA editing patterns with them rather than with L. tarentolae. These findings provide novel insight into the biological features of this emerging human pathogen.
- MeSH
- anotace sekvence MeSH
- editace RNA MeSH
- fylogeneze MeSH
- kinetoplastová DNA chemie genetika MeSH
- molekulární sekvence - údaje MeSH
- polymerázová řetězová reakce MeSH
- pořadí genů MeSH
- protozoální DNA chemie genetika MeSH
- sekvenční analýza DNA MeSH
- sekvenční homologie nukleových kyselin MeSH
- shluková analýza MeSH
- Trypanosoma lewisi genetika MeSH
- vysoce účinné nukleotidové sekvenování MeSH
- zastoupení bazí MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- kinetoplastová DNA MeSH
- protozoální DNA MeSH
