The African trypanosome, Trypanosoma brucei, has developed into a flexible and robust experimental model for molecular and cellular parasitology, allowing us to better combat these and related parasites that cause worldwide suffering. Diminishing case numbers, due to efficient public health efforts, and recent development of new drug treatments have reduced the need for continued study of T. brucei in a disease context. However, we argue that this pathogen has been instrumental in revolutionary discoveries that have widely informed molecular and cellular biology and justifies continuing research as an experimental model. Ongoing work continues to contribute towards greater understanding of both diversified and conserved biological features. We discuss multiple examples where trypanosomes pushed the boundaries of cell biology and hope to inspire researchers to continue exploring these remarkable protists as tools for magnifying the inner workings of cells.
- Klíčová slova
- GPI-anchor, RNA editing, alternative oxidase, endocytosis, genetic code, glycosome, tRNA import,
- MeSH
- molekulární biologie MeSH
- Trypanosoma brucei brucei * genetika MeSH
- Trypanosoma * genetika MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
The diverse GTPases of the dynamin superfamily play various roles in the cell, as exemplified by the dynamin-related proteins (DRPs) Mgm1 and Opa1, which remodel the mitochondrial inner membrane in fungi and metazoans, respectively. Via an exhaustive search of genomic and metagenomic databases, we found previously unknown DRP types occurring in diverse eukaryotes and giant viruses (phylum Nucleocytoviricota). One novel DRP clade, termed MidX, combined hitherto uncharacterized proteins from giant viruses and six distantly related eukaryote taxa (Stramenopiles, Telonemia, Picozoa, Amoebozoa, Apusomonadida, and Choanoflagellata). MidX stood out because it was not only predicted to be mitochondria-targeted but also to assume a tertiary structure not observed in other DRPs before. To understand how MidX affects mitochondria, we exogenously expressed MidX from Hyperionvirus in the kinetoplastid Trypanosoma brucei, which lacks Mgm1 or Opa1 orthologs. MidX massively affected mitochondrial morphology from inside the matrix, where it closely associates with the inner membrane. This unprecedented mode of action contrasts to those of Mgm1 and Opa1, which mediate inner membrane remodeling in the intermembrane space. We speculate that MidX was acquired in Nucleocytoviricota evolution by horizontal gene transfer from eukaryotes and is used by giant viruses to remodel host mitochondria during infection. MidX's unique structure may be an adaptation for reshaping mitochondria from the inside. Finally, Mgm1 forms a sister group to MidX and not Opa1 in our phylogenetic analysis, throwing into question the long-presumed homology of these DRPs with similar roles in sister lineages.
- Klíčová slova
- Nucleocytoviricota, Mgm1, Opa1, dynamin superfamily, mitochondria, protists,
- MeSH
- dynaminy genetika metabolismus MeSH
- fylogeneze MeSH
- mitochondriální proteiny genetika metabolismus MeSH
- mitochondrie genetika metabolismus MeSH
- obří viry * genetika metabolismus MeSH
- Saccharomyces cerevisiae genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- dynaminy MeSH
- mitochondriální proteiny MeSH
Mitochondrial cristae expand the surface area of respiratory membranes and ultimately allow for the evolutionary scaling of respiration with cell volume across eukaryotes. The discovery of Mic60 homologs among alphaproteobacteria, the closest extant relatives of mitochondria, suggested that cristae might have evolved from bacterial intracytoplasmic membranes (ICMs). Here, we investigated the predicted structure and function of alphaproteobacterial Mic60, and a protein encoded by an adjacent gene Orf52, in two distantly related purple alphaproteobacteria, Rhodobacter sphaeroides and Rhodopseudomonas palustris. In addition, we assessed the potential physical interactors of Mic60 and Orf52 in R. sphaeroides. We show that the three α helices of mitochondrial Mic60's mitofilin domain, as well as its adjacent membrane-binding amphipathic helix, are present in alphaproteobacterial Mic60. The disruption of Mic60 and Orf52 caused photoheterotrophic growth defects, which are most severe under low light conditions, and both their disruption and overexpression led to enlarged ICMs in both studied alphaproteobacteria. We also found that alphaproteobacterial Mic60 physically interacts with BamA, the homolog of Sam50, one of the main physical interactors of eukaryotic Mic60. This interaction, responsible for making contact sites at mitochondrial envelopes, has been conserved in modern alphaproteobacteria despite more than a billion years of evolutionary divergence. Our results suggest a role for Mic60 in photosynthetic ICM development and contact site formation at alphaproteobacterial envelopes. Overall, we provide support for the hypothesis that mitochondrial cristae evolved from alphaproteobacterial ICMs and have therefore improved our understanding of the nature of the mitochondrial ancestor.
- Klíčová slova
- Cereibacter, MICOS, Rhodobacter, Rhodopseudomonas, chromatophores, endosymbosis, eukaryogenesis, eukaryote, purple bacteria,
- MeSH
- Alphaproteobacteria * genetika metabolismus MeSH
- biologická evoluce MeSH
- mitochondriální membrány metabolismus MeSH
- mitochondriální proteiny * metabolismus MeSH
- mitochondrie metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- mitochondriální proteiny * MeSH
Kinesins are motor proteins found in all eukaryotic lineages that move along microtubules to mediate cellular processes such as mitosis and intracellular transport. In trypanosomatids, the kinesin superfamily has undergone a prominent expansion, resulting in one of the most diverse kinesin repertoires that includes the two kinetoplastid-restricted families X1 and X2. Here, we characterize in Trypanosoma brucei TbKifX2A, an orphaned X2 kinesin. TbKifX2A tightly interacts with TbPH1, a kinesin-like protein with a likely inactive motor domain, a rarely reported occurrence. Both TbKifX2A and TbPH1 localize to the microtubule quartet (MtQ), a characteristic but poorly understood cytoskeletal structure that wraps around the flagellar pocket as it extends to the cell body anterior. The proximal proteome of TbPH1 revealed two other interacting proteins, the flagellar pocket protein FP45 and intriguingly another X2 kinesin, TbKifX2C. Simultaneous ablation of TbKifX2A/TbPH1 results in the depletion of FP45 and TbKifX2C and also an expansion of the flagellar pocket, among other morphological defects. TbKifX2A is the first motor protein to be localized to the MtQ. The observation that TbKifX2C also associates with the MtQ suggests that the X2 kinesin family may have co-evolved with the MtQ, both kinetoplastid-specific traits.
- Klíčová slova
- Trypanosoma, cytoskeleton, kinesin, microtubule quartet, microtubules, morphogenesis,
- MeSH
- cytoskelet metabolismus MeSH
- kineziny * genetika metabolismus MeSH
- mikrotubuly metabolismus MeSH
- PH-doména MeSH
- protozoální proteiny * genetika metabolismus MeSH
- Trypanosoma brucei brucei * metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- kineziny * MeSH
- protozoální proteiny * MeSH
The mitochondrial protein import machinery of trypanosomatids is highly divergent from that of the well-studied models such as baker's yeast. A notable example is that the central catalyst of the mitochondrial intermembrane space import and assembly pathway (MIA), named Mia40, is missing in trypanosomatids. Mia40 works in a two-step process. First it recognizes by direct binding reduced MIA substrate proteins and then catalyzes their oxidative folding to produce intramolecular disulfide bridges. It was recently proposed that a thioredoxin-like subunit of the trypanosomal mitochondrial contact site and cristae organizing system (MICOS) called TbMic20 may be the Mia40 replacement. Our study performed on procyclic stage of the parasite revealed that each of the two cysteines in TbMic20's active site is essential for the stability of MIA substrate proteins although they do not form a disulfide bridge in vivo. The two cysteines of Mia40's active site form an intramolecular disulfide bridge at steady state, which is a prerequisite for its oxidative folding of MIA substrates. Thus, we conclude that TbMic20 is unlikely to represent a bona fide Mia40 replacement and plays a still unresolved role in the stability and/or import of MIA substrates in trypanosomatids. Despite this, the effect of TbMic20 depletion and mutation indicates that the trypanosomal MICOS complex still plays a vital role in the maturation and/or stability of proteins imported by the MIA pathway.
- Klíčová slova
- Intermembrane space, MICOS, Mitochondrion, Oxidative folding, Protein import, Trypanosoma,
- MeSH
- chlorprofam metabolismus MeSH
- cystein metabolismus MeSH
- disulfidy MeSH
- mitochondriální importní komplex MeSH
- mitochondriální proteiny metabolismus MeSH
- oxidace-redukce MeSH
- Saccharomyces cerevisiae - proteiny * genetika MeSH
- Saccharomyces cerevisiae metabolismus MeSH
- sbalování proteinů MeSH
- thioredoxiny genetika metabolismus MeSH
- transport proteinů MeSH
- transportní proteiny mitochondriální membrány genetika MeSH
- transportní proteiny metabolismus MeSH
- Trypanosoma brucei brucei * genetika metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- chlorprofam MeSH
- cystein MeSH
- disulfidy MeSH
- MIA40 protein, S cerevisiae MeSH Prohlížeč
- mitochondriální importní komplex MeSH
- mitochondriální proteiny MeSH
- Saccharomyces cerevisiae - proteiny * MeSH
- thioredoxiny MeSH
- transportní proteiny mitochondriální membrány MeSH
- transportní proteiny MeSH
Mitochondrial cristae are polymorphic invaginations of the inner membrane that are the fabric of cellular respiration. Both the mitochondrial contact site and cristae organization system (MICOS) and the F1FO-ATP synthase are vital for sculpting cristae by opposing membrane-bending forces. While MICOS promotes negative curvature at crista junctions, dimeric F1FO-ATP synthase is crucial for positive curvature at crista rims. Crosstalk between these two complexes has been observed in baker's yeast, the model organism of the Opisthokonta supergroup. Here, we report that this property is conserved in Trypanosoma brucei, a member of the Discoba clade that separated from the Opisthokonta ∼2 billion years ago. Specifically, one of the paralogs of the core MICOS subunit Mic10 interacts with dimeric F1FO-ATP synthase, whereas the other core Mic60 subunit has a counteractive effect on F1FO-ATP synthase oligomerization. This is evocative of the nature of MICOS-F1FO-ATP synthase crosstalk in yeast, which is remarkable given the diversification that these two complexes have undergone during almost 2 eons of independent evolution. Furthermore, we identified a highly diverged, putative homolog of subunit e, which is essential for the stability of F1FO-ATP synthase dimers in yeast. Just like subunit e, it is preferentially associated with dimers and interacts with Mic10, and its silencing results in severe defects to cristae and the disintegration of F1FO-ATP synthase dimers. Our findings indicate that crosstalk between MICOS and dimeric F1FO-ATP synthase is a fundamental property impacting crista shape throughout eukaryotes. IMPORTANCE Mitochondria have undergone profound diversification in separate lineages that have radiated since the last common ancestor of eukaryotes some eons ago. Most eukaryotes are unicellular protists, including etiological agents of infectious diseases, like Trypanosoma brucei. Thus, the study of a broad range of protists can reveal fundamental features shared by all eukaryotes and lineage-specific innovations. Here, we report that two different protein complexes, MICOS and F1FO-ATP synthase, known to affect mitochondrial architecture, undergo crosstalk in T. brucei, just as in baker's yeast. This is remarkable considering that these complexes have otherwise undergone many changes during their almost 2 billion years of independent evolution. Thus, this crosstalk is a fundamental property needed to maintain proper mitochondrial structure even if the constituent players considerably diverged.
- Klíčová slova
- ATP synthase, MICOS, Trypanosoma, evolution, mitochondria,
- Publikační typ
- časopisecké články MeSH
The mitochondrion is crucial for ATP generation by oxidative phosphorylation, among other processes. Cristae are invaginations of the mitochondrial inner membrane that house nearly all the macromolecular complexes that perform oxidative phosphorylation. The unicellular parasite Trypanosoma brucei undergoes during its life cycle extensive remodeling of its single mitochondrion, which reflects major changes in its energy metabolism. While the bloodstream form (BSF) generates ATP exclusively by substrate-level phosphorylation and has a morphologically highly reduced mitochondrion, the insect-dwelling procyclic form (PCF) performs oxidative phosphorylation and has an expanded and reticulated organelle. Here, we have performed high-resolution 3D reconstruction of BSF and PCF mitochondria, with a particular focus on their cristae. By measuring the volumes and surface areas of these structures in complete or nearly complete cells, we have found that mitochondrial cristae are more prominent in BSF than previously thought and their biogenesis seems to be maintained during the cell cycle. Furthermore, PCF cristae exhibit a surprising range of volumes in situ, implying that each crista is acting as an independent bioenergetic unit. Cristae appear to be particularly enriched in the region of the organelle between the nucleus and kinetoplast, the mitochondrial genome, suggesting this part has distinctive properties.
- Klíčová slova
- Cristae, kinetoplastid, mitochondria, tomography,
- MeSH
- buněčné jádro MeSH
- buněčný cyklus MeSH
- mitochondrie MeSH
- stadia vývoje MeSH
- Trypanosoma brucei brucei * MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Receptor adenylate cyclases (RACs) on the surface of trypanosomatids are important players in the host-parasite interface. They detect still unidentified environmental signals that affect the parasites' responses to host immune challenge, coordination of social motility, and regulation of cell division. A lesser known class of oxygen-sensing adenylate cyclases (OACs) related to RACs has been lost in trypanosomes and expanded mostly in Leishmania species and related insect-dwelling trypanosomatids. In this work, we have undertaken a large-scale phylogenetic analysis of both classes of adenylate cyclases (ACs) in trypanosomatids and the free-living Bodo saltans. We observe that the expanded RAC repertoire in trypanosomatids with a two-host life cycle is not only associated with an extracellular lifestyle within the vertebrate host, but also with a complex path through the insect vector involving several life cycle stages. In Trypanosoma brucei, RACs are split into two major clades, which significantly differ in their expression profiles in the mammalian host and the insect vector. RACs of the closely related Trypanosoma congolense are intermingled within these two clades, supporting early RAC diversification. Subspecies of T. brucei that have lost the capacity to infect insects exhibit high numbers of pseudogenized RACs, suggesting many of these proteins have become redundant upon the acquisition of a single-host life cycle. OACs appear to be an innovation occurring after the expansion of RACs in trypanosomatids. Endosymbiont-harboring trypanosomatids exhibit a diversification of OACs, whereas these proteins are pseudogenized in Leishmania subgenus Viannia. This analysis sheds light on how ACs have evolved to allow diverse trypanosomatids to occupy multifarious niches and assume various lifestyles.
- Klíčová slova
- Kinetoplastida, adenylate cyclase, oxygen, phylogenetics, receptor, trypanosomatid,
- MeSH
- adenylátcyklasy genetika MeSH
- duplikace genu MeSH
- fylogeneze * MeSH
- genom protozoální MeSH
- interakce hostitele a patogenu genetika MeSH
- molekulární evoluce * MeSH
- Trypanosomatina enzymologie genetika MeSH
- upregulace MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- adenylátcyklasy MeSH
Cristae are infoldings of the mitochondrial inner membrane jutting into the organelle's innermost compartment from narrow stems at their base called crista junctions. They are emblematic of aerobic mitochondria, being the fabric for the molecular machinery driving cellular respiration. Electron microscopy revealed that diverse eukaryotes possess cristae of different shapes. Yet, crista diversity has not been systematically examined in light of our current knowledge about eukaryotic evolution. Since crista form and function are intricately linked, we take a holistic view of factors that may underlie both crista diversity and the adherence of cristae to a recognizable form. Based on electron micrographs of 226 species from all major lineages, we propose a rational crista classification system that postulates cristae as variations of two general morphotypes: flat and tubulo-vesicular. The latter is most prevalent and likely ancestral, but both morphotypes are found interspersed throughout the eukaryotic tree. In contrast, crista junctions are remarkably conserved, supporting their proposed role as diffusion barriers that sequester cristae contents. Since cardiolipin, ATP synthase dimers, the MICOS complex, and dynamin-like Opa1/Mgm1 are known to be involved in shaping cristae, we examined their variation in the context of crista diversity. Moreover, we have identified both commonalities and differences that may collectively be manifested as diverse variations of crista form and function.
