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ELife May 2023Iron-sulfur clusters (FeS) are ancient and ubiquitous protein cofactors that play fundamental roles in many aspects of cell biology. These cofactors cannot be scavenged...
Iron-sulfur clusters (FeS) are ancient and ubiquitous protein cofactors that play fundamental roles in many aspects of cell biology. These cofactors cannot be scavenged or trafficked within a cell and thus must be synthesized in any subcellular compartment where they are required. We examined the FeS synthesis proteins found in the relict plastid organelle, called the apicoplast, of the human malaria parasite . Using a chemical bypass method, we deleted four of the FeS pathway proteins involved in sulfur acquisition and cluster assembly and demonstrated that they are all essential for parasite survival. However, the effect that these deletions had on the apicoplast organelle differed. Deletion of the cysteine desulfurase SufS led to disruption of the apicoplast organelle and loss of the organellar genome, whereas the other deletions did not affect organelle maintenance. Ultimately, we discovered that the requirement of SufS for organelle maintenance is not driven by its role in FeS biosynthesis, but rather, by its function in generating sulfur for use by MnmA, a tRNA modifying enzyme that we localized to the apicoplast. Complementation of MnmA and SufS activity with a bacterial MnmA and its cognate cysteine desulfurase strongly suggests that the parasite SufS provides sulfur for both FeS biosynthesis and tRNA modification in the apicoplast. The dual role of parasite SufS is likely to be found in other plastid-containing organisms and highlights the central role of this enzyme in plastid biology.
Topics: Humans; Plasmodium falciparum; Apicoplasts; Sulfur; Iron; RNA, Transfer; Iron-Sulfur Proteins
PubMed: 37166116
DOI: 10.7554/eLife.84491 -
ELife Mar 2024The apicoplast is a four-membrane plastid found in the apicomplexans, which harbors biosynthesis and organelle housekeeping activities in the matrix. However, the...
The apicoplast is a four-membrane plastid found in the apicomplexans, which harbors biosynthesis and organelle housekeeping activities in the matrix. However, the mechanism driving the flux of metabolites, in and out, remains unknown. Here, we used TurboID and genome engineering to identify apicoplast transporters in . Among the many novel transporters, we show that one pair of apicomplexan monocarboxylate transporters (AMTs) appears to have evolved from a putative host cell that engulfed a red alga. Protein depletion showed that AMT1 and AMT2 are critical for parasite growth. Metabolite analyses supported the notion that AMT1 and AMT2 are associated with biosynthesis of isoprenoids and fatty acids. However, stronger phenotypic defects were observed for AMT2, including in the inability to establish parasite virulence in mice. This study clarifies, significantly, the mystery of apicoplast transporter composition and reveals the importance of the pair of AMTs in maintaining the apicoplast activity in apicomplexans.
Topics: Animals; Mice; Toxoplasma; Parasites; Apicoplasts; Fatty Acids; Organic Chemicals; Protozoan Proteins
PubMed: 38502570
DOI: 10.7554/eLife.88866 -
PLoS Pathogens Oct 2023Isoprenoid precursor synthesis is an ancient and fundamental function of plastid organelles and a critical metabolic activity of the apicoplast in Plasmodium malaria...
Isoprenoid precursor synthesis is an ancient and fundamental function of plastid organelles and a critical metabolic activity of the apicoplast in Plasmodium malaria parasites [1-3]. Over the past decade, our understanding of apicoplast properties and functions has increased enormously [4], due in large part to our ability to rescue blood-stage parasites from apicoplast-specific dysfunctions by supplementing cultures with isopentenyl pyrophosphate (IPP), a key output of this organelle [5,6]. In this Pearl, we explore the interdependence between isoprenoid metabolism and apicoplast biogenesis in P. falciparum and highlight critical future questions to answer.
Topics: Animals; Apicoplasts; Parasites; Plasmodium falciparum; Malaria, Falciparum; Protozoan Proteins
PubMed: 37883328
DOI: 10.1371/journal.ppat.1011713 -
Genes Aug 2018The organelles of endosymbiotic origin, plastids, and mitochondria, evolved through the serial acquisition of endosymbionts by a host cell. These events were... (Review)
Review
The organelles of endosymbiotic origin, plastids, and mitochondria, evolved through the serial acquisition of endosymbionts by a host cell. These events were accompanied by gene transfer from the symbionts to the host, resulting in most of the organellar proteins being encoded in the cell nuclear genome and trafficked into the organelle via a series of translocation complexes. Much of what is known about organelle protein translocation mechanisms is based on studies performed in common model organisms; e.g., yeast and humans or . However, studies performed in divergent organisms are gradually accumulating. These studies provide insights into universally conserved traits, while discovering traits that are specific to organisms or clades. Apicomplexan parasites feature two organelles of endosymbiotic origin: a secondary plastid named the apicoplast and a mitochondrion. In the context of the diseases caused by apicomplexan parasites, the essential roles and divergent features of both organelles make them prime targets for drug discovery. This potential and the amenability of the apicomplexan Toxoplasma gondii to genetic manipulation motivated research about the mechanisms controlling both organelles' biogenesis. Here we provide an overview of what is known about apicomplexan organelle protein import. We focus on work done mainly in T. gondii and provide a comparison to model organisms.
PubMed: 30110980
DOI: 10.3390/genes9080412 -
Computational and Structural... 2013Autophagy is a catabolic process that normally utilizes the lysosome. The far-reaching implications of this system in disease are being increasingly understood. Studying... (Review)
Review
Autophagy is a catabolic process that normally utilizes the lysosome. The far-reaching implications of this system in disease are being increasingly understood. Studying autophagy is complicated by its role in cell survival and programmed cell death and the involvement of the canonical marker of autophagy, Atg8/LC3, in numerous non-autophagic roles. The malaria parasite, Plasmodium, has conserved certain aspects of the autophagic machinery but for what purpose has long remained a mystery. Major advances have recently been gained and suggest a role for Atg8 in apicoplast maintenance, degradation of heme inside the food vacuole, and possibly trafficking of proteins or organelles outside the parasite membrane. Autophagy may also participate in programmed cell death under drug treatment or as a selective tool to limit parasite load. We review the current findings and discuss discrepancies in the field of autophagy in the Plasmodium parasite.
PubMed: 24688742
DOI: 10.5936/csbj.201308002 -
Traffic (Copenhagen, Denmark) May 2008The accurate targeting of proteins to their final destination is an essential process in all living cells. Apicomplexans are obligate intracellular protozoan parasites... (Review)
Review
The accurate targeting of proteins to their final destination is an essential process in all living cells. Apicomplexans are obligate intracellular protozoan parasites that possess a compartmental organization similar to that of free-living eukaryotes but can be viewed as professional secretory cells. Establishment of parasitism involves the sequential secretion from highly specialized secretory organelles, including micronemes, rhoptries and dense granules. Additionally, apicomplexans harbor a tubular mitochondrion, a nonphotosynthetic plastid organelle termed the apicoplast, acidocalcisomes and an elaborated inner membrane complex composed of flattened membrane cisternae that are derived from the secretory pathway. Given the multitude of destinations both inside and outside the parasite, the endoplasmic reticulum/Golgi of the apicomplexans constitutes one of the most busy roads intersections in eukaryotic traffic.
Topics: Animals; Endoplasmic Reticulum; Golgi Apparatus; Humans; Plasmodium falciparum; Plastids; Protein Transport; Protozoan Proteins; Toxoplasma
PubMed: 18331382
DOI: 10.1111/j.1600-0854.2008.00713.x -
International Journal For Parasitology Mar 2004Toxoplasma gondii is an obligate intracellular protozoan parasite which is a significant human and veterinary pathogen. Other members of the phylum Apicomplexa are also... (Review)
Review
Toxoplasma gondii is an obligate intracellular protozoan parasite which is a significant human and veterinary pathogen. Other members of the phylum Apicomplexa are also important pathogens including Plasmodium species (i.e. malaria), Eimeria species, Neospora, Babesia, Theileria and Cryptosporidium. Unlike most of these organisms, T. gondii is readily amenable to genetic manipulation in the laboratory. Cell biology studies are more readily performed in T. gondii due to the high efficiency of transient and stable transfection, the availability of many cell markers, and the relative ease with which the parasite can be studied using advanced microscopic techniques. Thus, for many experimental questions, T. gondii remains the best model system to study the biology of the Apicomplexa. Our understanding of the mechanisms of drug resistance, the biology of the apicoplast, and the process of host cell invasion has been advanced by studies in T. gondii. Heterologous expression of apicomplexan proteins in T. gondii has frequently facilitated further characterisation of proteins that could not be easily studied. Recent studies of Apicomplexa have been complemented by genome sequencing projects that have facilitated discovery of surprising differences in cell biology and metabolism between Apicomplexa. While results in T. gondii will not always be applicable to other Apicomplexa, T. gondii remains an important model system for understanding the biology of apicomplexan parasites.
Topics: Animals; Antiprotozoal Agents; Genes, Protozoan; Models, Biological; Purines; Pyrimidines; Toxoplasma
PubMed: 15003501
DOI: 10.1016/j.ijpara.2003.12.009 -
International Journal For Parasitology May 2021Apicomplexans are the causative agents of numerous important infectious diseases including malaria and toxoplasmosis. Most of them harbour a chloroplast-like organelle...
Apicomplexans are the causative agents of numerous important infectious diseases including malaria and toxoplasmosis. Most of them harbour a chloroplast-like organelle called the apicoplast that is essential for the parasites' metabolism and survival. While most apicoplast proteins are nuclear encoded, the organelle also maintains its own genome, a 35 kb circle. In this study we used Toxoplasma gondii to identify and characterise essential proteins involved in apicoplast genome replication and to understand how apicoplast genome segregation unfolds over time. We demonstrated that the DNA replication enzymes Prex, DNA gyrase and DNA single stranded binding protein localise to the apicoplast. We show in knockdown experiments that apicoplast DNA Gyrase A and B, and Prex are required for apicoplast genome replication and growth of the parasite. Analysis of apicoplast genome replication by structured illumination microscopy in T. gondii tachyzoites showed that apicoplast nucleoid division and segregation initiate at the beginning of S phase and conclude during mitosis. Thus, the replication and division of the apicoplast nucleoid is highly coordinated with nuclear genome replication and mitosis. Our observations highlight essential components of apicoplast genome maintenance and shed light on the timing of this process in the context of the overall parasite cell cycle.
Topics: Apicoplasts; Cell Division; DNA Gyrase; DNA-Directed DNA Polymerase; Humans; Toxoplasma; Toxoplasmosis
PubMed: 33581138
DOI: 10.1016/j.ijpara.2020.11.004 -
Microbiology Spectrum Feb 2022The apicoplast, which harbors key pathways involved in biosynthesis of vital metabolites, is a unique and essential nonphotosynthetic plastid organelle in apicomplexan...
The apicoplast, which harbors key pathways involved in biosynthesis of vital metabolites, is a unique and essential nonphotosynthetic plastid organelle in apicomplexan parasites. Intriguingly, autophagy-related protein 8 (Atg8), a highly conserved eukaryotic protein, can localize to the outermost membrane of the apicoplast and modulate its inheritance in both and parasites. The Atg8-Atg3 interaction plays a key role in Atg8 lipidation and localization, and our previously work in has suggested that the core Atg8-family interacting motif (AIM) in TgAtg3, FADI, and the R27 residue of TgAtg8 contribute to TgAtg8-TgAtg3 interaction . However, little is known about the function of this interaction or its importance in tachyzoite growth in . Here, we generated two complemented cell lines, TgAtg3 and TgAtg8, based on the TgAtg3 and TgAtg8 conditional knockdown cell lines, respectively. We found that both mutant complemented cell lines were severely affected in terms of tachyzoite growth and displayed delayed death upon conditional knockdown of endogenous TgAtg3 or TgAtg8. Intriguingly, both complemented lines appeared to be defective in TgAtg8 lipidation and apicoplast inheritance. Moreover, we showed that the interaction of TgAtg8 and TgAtg3 is critical for TgAtg8 apicoplast localization. In addition, we found that the TgAtg3 complemented line exhibits an integral mitochondrial network upon ablation of endogenous TgAtg3, which is distinct from TgAtg3-depleted parasites with a fragmented mitochondrial network. Taken together, this work solidifies the contribution of the TgAtg8-TgAtg3 interaction to apicoplast inheritance and the growth of tachyzoites. is a widespread intracellular parasite infecting a variety of warm-blooded animals, including humans. Current frontline treatment of toxoplasmosis suffers many drawbacks, including toxicity, drug resistance, and failure to eradicate tissue cysts, underscoring the need to identify novel drug targets for suppression or treatment of toxoplasmosis. TgAtg8 is thought to serve multiple functions in lipidation and is considered essential to the growth and development of both tachyzoites and bradyzoites. Here, we show that has adapted a conserved Atg8-Atg3 interaction, required for canonical autophagy in other eukaryotes, to function specifically in apicoplast inheritance. Our finding not only highlights the importance of TgAtg8-TgAtg3 interaction in tachyzoite growth but also suggests that this interaction is a promising drug target for the therapy of toxoplasmosis.
Topics: Amino Acid Motifs; Apicoplasts; Humans; Mutation; Protein Binding; Protein Transport; Protozoan Proteins; Toxoplasma; Toxoplasmosis
PubMed: 35196797
DOI: 10.1128/spectrum.01495-21 -
The EMBO Journal Aug 2021Malaria parasites contain an essential organelle called the apicoplast that houses metabolic pathways for fatty acid, heme, isoprenoid, and iron-sulfur cluster...
Malaria parasites contain an essential organelle called the apicoplast that houses metabolic pathways for fatty acid, heme, isoprenoid, and iron-sulfur cluster synthesis. Surprisingly, malaria parasites can survive without the apicoplast as long as the isoprenoid precursor isopentenyl pyrophosphate (IPP) is supplemented in the growth medium, making it appear that isoprenoid synthesis is the only essential function of the organelle in blood-stage parasites. In the work described here, we localized an enzyme responsible for coenzyme A synthesis, DPCK, to the apicoplast, but we were unable to delete DPCK, even in the presence of IPP. However, once the endogenous DPCK was complemented with the E. coli DPCK (EcDPCK), we were successful in deleting it. We were then able to show that DPCK activity is required for parasite survival through knockdown of the complemented EcDPCK. Additionally, we showed that DPCK enzyme activity remains functional and essential within the vesicles present after apicoplast disruption. These results demonstrate that while the apicoplast of blood-stage P. falciparum parasites can be disrupted, the resulting vesicles remain biochemically active and are capable of fulfilling essential functions.
Topics: Apicoplasts; Pantothenic Acid; Phosphotransferases (Alcohol Group Acceptor); Plasmodium falciparum; Protozoan Proteins
PubMed: 34031901
DOI: 10.15252/embj.2020107247