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Open accessJournal ArticleDOI: 10.1371/JOURNAL.PPAT.1009301

Complexome profile of Toxoplasma gondii mitochondria identifies divergent subunits of respiratory chain complexes including new subunits of cytochrome bc1 complex.

02 Mar 2021-PLOS Pathogens (Public Library of Science (PLoS))-Vol. 17, Iss: 3
Abstract: The mitochondrial electron transport chain (mETC) and F1Fo-ATP synthase are of central importance for energy and metabolism in eukaryotic cells. The Apicomplexa, important pathogens of humans causing diseases such as toxoplasmosis and malaria, depend on their mETC in every known stage of their complicated life cycles. Here, using a complexome profiling proteomic approach, we have characterised the Toxoplasma mETC complexes and F1Fo-ATP synthase. We identified and assigned 60 proteins to complexes II, IV and F1Fo-ATP synthase of Toxoplasma, of which 16 have not been identified previously. Notably, our complexome profile elucidates the composition of the Toxoplasma complex III, the target of clinically used drugs such as atovaquone. We identified two new homologous subunits and two new parasite-specific subunits, one of which is broadly conserved in myzozoans. We demonstrate all four proteins are essential for complex III stability and parasite growth, and show their depletion leads to decreased mitochondrial potential, supporting their assignment as complex III subunits. Our study highlights the divergent subunit composition of the apicomplexan mETC and F1Fo-ATP synthase complexes and sets the stage for future structural and drug discovery studies.

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Topics: Respiratory chain (56%), Protein subunit (54%), Coenzyme Q – cytochrome c reductase (53%) ... show more
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7 results found


Open accessJournal ArticleDOI: 10.1038/S41467-021-23919-X
Abstract: Our current understanding of mitochondrial functioning is largely restricted to traditional model organisms, which only represent a fraction of eukaryotic diversity. The unusual mitochondrion of malaria parasites is a validated drug target but remains poorly understood. Here, we apply complexome profiling to map the inventory of protein complexes across the pathogenic asexual blood stages and the transmissible gametocyte stages of Plasmodium falciparum. We identify remarkably divergent composition and clade-specific additions of all respiratory chain complexes. Furthermore, we show that respiratory chain complex components and linked metabolic pathways are up to 40-fold more prevalent in gametocytes, while glycolytic enzymes are substantially reduced. Underlining this functional switch, we find that cristae are exclusively present in gametocytes. Leveraging these divergent properties and stage dynamics for drug development presents an attractive opportunity to discover novel classes of antimalarials and increase our repertoire of gametocytocidal drugs.

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Topics: Respiratory chain (59%), Respiratory chain complex (58%), Gametocyte (56%) ... show more

5 Citations


Open accessPosted ContentDOI: 10.1101/2021.01.28.428257
Sarah Pamukcu1, Aude Cerutti1, Yann Bordat1, Sonia Hem1  +2 moreInstitutions (1)
31 Aug 2021-bioRxiv
Abstract: Iron-sulfur (Fe-S) clusters are one of the most ancient and ubiquitous prosthetic groups, and they are required by a variety of proteins involved in important metabolic processes. Apicomplexan parasites have inherited different plastidic and mitochondrial Fe-S clusters biosynthesis pathways through endosymbiosis. We have investigated the relative contributions of these pathways to the fitness of Toxoplasma gondii, an apicomplexan parasite causing disease in humans, by generating specific mutants. Phenotypic analysis and quantitative proteomics allowed us to highlight notable differences in these mutants. Both Fe-S cluster synthesis pathways are necessary for optimal parasite growth in vitro, but their disruption leads to markedly different fates: impairment of the plastidic pathway leads to a loss of the organelle and to parasite death, while disruption of the mitochondrial pathway trigger differentiation into a stress resistance stage. This highlights that otherwise similar biochemical pathways hosted by different sub-cellular compartments can have very different contributions to the biology of the parasites, which is something to consider when exploring novel strategies for therapeutic intervention. Author summary Toxoplasma gondii is a ubiquitous unicellular parasite that harbours two organelles of endosymbiotic origin: the mitochondrion, and a relict plastid named the apicoplast. Each one of these organelles contains its own machinery for synthesizing iron-sulfur clusters, which are important protein co- factors. In this study, we show that interfering with either the mitochondrial or the plastidic iron- sulfur cluster synthesizing machinery has a profound impact on parasite growth. However, while disrupting the plastidic pathway led to an irreversible loss of the organelle and subsequent death of the parasite, disrupting the mitochondrial pathway led to conversion of the parasites into a stress resistance form. We used a comparative quantitative proteomic analysis of the mutants, combined with experimental validation, to provide mechanistic clues into these different phenotypic outcomes. Although the consequences of disrupting each pathway were manifold, our data highlighted distinct changes at the metabolic level. For instance, the plastidic iron-sulfur cluster synthesis pathway is likely important for maintaining the lipid homeostasis of the parasites, while the mitochondrial pathway is clearly crucial for maintaining their respiratory capacity. Interestingly, we have discovered that other mutants severely impacted for mitochondrial function, in particular the respiratory chain, are able to survive and initiate conversion to the stress resistance form. This illustrates a different capacity for T. gondii to adapt for survival in response to distinct metabolic dysregulations.

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Topics: Respiratory chain (54%), Apicoplast (54%), Mitochondrion (50%)

2 Citations


Open accessPosted ContentDOI: 10.1101/2021.01.28.428257
Sarah Pamukcu1, Aude Cerutti1, Sonia Hem1, Valerie Rofidal1  +1 moreInstitutions (2)
28 Jan 2021-bioRxiv
Abstract: Iron-sulfur (Fe-S) clusters are one of the most ancient and ubiquitous prosthetic groups, and they are required by a variety of proteins involved in important metabolic processes. Apicomplexan parasites have inherited different plastidic and mitochondrial Fe-S clusters biosynthesis pathways through endosymbiosis. We have investigated the relative contributions of these pathways to the fitness of Toxoplasma gondii, an apicomplexan parasite causing disease in humans, by generating specific mutants. Phenotypic analysis and quantitative proteomics allowed us to highlight striking differences in these mutants. Both Fe-S cluster synthesis pathways are necessary for optimal parasite growth in vitro, but their disruption leads to markedly different fates: impairment of the plastidic pathway leads to a loss of the organelle and to parasite death, while disruption of the mitochondrial pathway trigger differentiation into a stress resistance stage. This highlights that otherwise similar biochemical pathways hosted by different sub-cellular compartments can have very different contributions to the biology of the parasites, which is something to consider when exploring novel strategies for therapeutic intervention.

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2 Citations


Open accessJournal ArticleDOI: 10.1101/GR.262832.120
Luisa Berná1, Pablo Marquez1, Andrés Cabrera1, Gonzalo Greif1  +3 moreInstitutions (2)
27 Apr 2021-Genome Research
Abstract: Neospora caninum primarily infects cattle, causing abortions, with an estimated impact of a billion dollars on the worldwide economy annually. However, the study of its biology has been unheeded by the established paradigm that it is virtually identical to its close relative, the widely studied human pathogen Toxoplasma gondii By revisiting the genome sequence, assembly, and annotation using third-generation sequencing technologies, here we show that the N. caninum genome was originally incorrectly assembled under the presumption of synteny with T. gondii We show that major chromosomal rearrangements have occurred between these species. Importantly, we show that chromosomes originally named Chr VIIb and VIII are indeed fused, reducing the karyotype of both N. caninum and T. gondii to 13 chromosomes. We reannotate the N. caninum genome, revealing more than 500 new genes. We sequence and annotate the nonphotosynthetic plastid and mitochondrial genomes and show that although apicoplast genomes are virtually identical, high levels of gene fragmentation and reshuffling exist between species and strains. Our results correct assembly artifacts that are currently widely distributed in the genome database of N. caninum and T. gondii and, more importantly, highlight the mitochondria as a previously oversighted source of variability and pave the way for a change in the paradigm of synteny, encouraging rethinking the genome as basis of the comparative unique biology of these pathogens.

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Topics: Neospora caninum (59%), Genome (56%), Synteny (53%) ... show more

2 Citations


Open accessJournal ArticleDOI: 10.1371/JOURNAL.PPAT.1010096
Sarah Pamukcu1, Aude Cerutti1, Yann Bordat1, Sonia Hem1  +2 moreInstitutions (2)
18 Nov 2021-PLOS Pathogens
Abstract: Iron-sulfur (Fe-S) clusters are one of the most ancient and ubiquitous prosthetic groups, and they are required by a variety of proteins involved in important metabolic processes. Apicomplexan parasites have inherited different plastidic and mitochondrial Fe-S clusters biosynthesis pathways through endosymbiosis. We have investigated the relative contributions of these pathways to the fitness of Toxoplasma gondii, an apicomplexan parasite causing disease in humans, by generating specific mutants. Phenotypic analysis and quantitative proteomics allowed us to highlight notable differences in these mutants. Both Fe-S cluster synthesis pathways are necessary for optimal parasite growth in vitro, but their disruption leads to markedly different fates: impairment of the plastidic pathway leads to a loss of the organelle and to parasite death, while disruption of the mitochondrial pathway trigger differentiation into a stress resistance stage. This highlights that otherwise similar biochemical pathways hosted by different sub-cellular compartments can have very different contributions to the biology of the parasites, which is something to consider when exploring novel strategies for therapeutic intervention.

... read more


References
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92 results found


Journal ArticleDOI: 10.1006/METH.2001.1262
01 Dec 2001-Methods
Abstract: The two most commonly used methods to analyze data from real-time, quantitative PCR experiments are absolute quantification and relative quantification. Absolute quantification determines the input copy number, usually by relating the PCR signal to a standard curve. Relative quantification relates the PCR signal of the target transcript in a treatment group to that of another sample such as an untreated control. The 2(-Delta Delta C(T)) method is a convenient way to analyze the relative changes in gene expression from real-time quantitative PCR experiments. The purpose of this report is to present the derivation, assumptions, and applications of the 2(-Delta Delta C(T)) method. In addition, we present the derivation and applications of two variations of the 2(-Delta Delta C(T)) method that may be useful in the analysis of real-time, quantitative PCR data.

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Topics: MicroRNA 34a (52%), Cell wall organization (50%)

116,500 Citations


Open accessJournal ArticleDOI: 10.1038/S41598-016-0028-X
Michiru Nishita1, Seung-Yeol Park2, Tadashi Nishio1, Koki Kamizaki1  +8 moreInstitutions (5)
26 Jan 2017-Scientific Reports
Abstract: Signaling through the Ror2 receptor tyrosine kinase promotes invadopodia formation for tumor invasion. Here, we identify intraflagellar transport 20 (IFT20) as a new target of this signaling in tumors that lack primary cilia, and find that IFT20 mediates the ability of Ror2 signaling to induce the invasiveness of these tumors. We also find that IFT20 regulates the nucleation of Golgi-derived microtubules by affecting the GM130-AKAP450 complex, which promotes Golgi ribbon formation in achieving polarized secretion for cell migration and invasion. Furthermore, IFT20 promotes the efficiency of transport through the Golgi complex. These findings shed new insights into how Ror2 signaling promotes tumor invasiveness, and also advance the understanding of how Golgi structure and transport can be regulated.

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Topics: Golgi apparatus (58%), Invadopodia (56%), Intraflagellar transport (51%) ... show more

8,752 Citations


Open access
01 Jan 2005-
Topics: Malaria (68%), Indoor residual spraying (55%)

4,964 Citations


Journal ArticleDOI: 10.1126/SCIENCE.272.5265.1136
24 May 1996-Science
Abstract: The crystal structure of bovine heart cytochrome c oxidase at 2.8 A resolution with an R value of 19.9 percent reveals 13 subunits, each different from the other, five phosphatidyl ethanolamines, three phosphatidyl glycerols and two cholates, two hemes A, and three copper, one magnesium, and one zinc. Of 3606 amino acid residues in the dimer, 3560 have been converged to a reasonable structure by refinement. A hydrogen-bonded system, including a propionate of a heme A (heme a), part of peptide backbone, and an imidazole ligand of CuA, could provide an electron transfer pathway between CuA and heme a. Two possible proton pathways for pumping, each spanning from the matrix to the cytosolic surfaces, were identified, including hydrogen bonds, internal cavities likely to contain water molecules, and structures that could form hydrogen bonds with small possible conformational change of amino acid side chains. Possible channels for chemical protons to produce H2O, for removing the produced water, and for O2, respectively, were identified.

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Topics: Heme A (62%), Heme O (59%), Heme (56%) ... show more

1,981 Citations


Open accessJournal ArticleDOI: 10.1038/NCOMMS5357
Abstract: Single-molecule surface-enhanced Raman scattering (SM-SERS) is one of the vital applications of plasmonic nanoparticles. The SM-SERS sensitivity critically depends on plasmonic hot-spots created at the vicinity of such nanoparticles. In conventional fluid-phase SM-SERS experiments, plasmonic hot-spots are facilitated by chemical aggregation of nanoparticles. Such aggregation is usually irreversible, and hence, nanoparticles cannot be re-dispersed in the fluid for further use. Here, we show how to combine SM-SERS with plasmon polariton-assisted, reversible assembly of plasmonic nanoparticles at an unstructured metal–fluid interface. One of the unique features of our method is that we use a single evanescent-wave optical excitation for nanoparticle assembly, manipulation and SM-SERS measurements. Furthermore, by utilizing dual excitation of plasmons at metal–fluid interface, we create interacting assemblies of metal nanoparticles, which may be further harnessed in dynamic lithography of dispersed nanostructures. Our work will have implications in realizing optically addressable, plasmofluidic, single-molecule detection platforms. Plasmonic hot-spot generation in solution is not reversible for single-molecule surface-enhanced Raman scattering, which limits its applications. Patra et al.tackle this problem by integrating this technique with thermo-plasmon-assisted reconfiguration of nanoparticles at a metal–fluid interface.

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Topics: Plasmonic nanoparticles (68%), Plasmon (52%), Raman scattering (50%)

1,682 Citations


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