scholarly journals Untargeted Metabolomics Uncovers the Essential Lysine Transporter in Toxoplasma gondii

Metabolites ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 476
Author(s):  
Joachim Kloehn ◽  
Matteo Lunghi ◽  
Emmanuel Varesio ◽  
David Dubois ◽  
Dominique Soldati-Favre

Apicomplexan parasites are responsible for devastating diseases, including malaria, toxoplasmosis, and cryptosporidiosis. Current treatments are limited by emerging resistance to, as well as the high cost and toxicity of existing drugs. As obligate intracellular parasites, apicomplexans rely on the uptake of many essential metabolites from their host. Toxoplasma gondii, the causative agent of toxoplasmosis, is auxotrophic for several metabolites, including sugars (e.g., myo-inositol), amino acids (e.g., tyrosine), lipidic compounds and lipid precursors (cholesterol, choline), vitamins, cofactors (thiamine) and others. To date, only few apicomplexan metabolite transporters have been characterized and assigned a substrate. Here, we set out to investigate whether untargeted metabolomics can be used to identify the substrate of an uncharacterized transporter. Based on existing genome- and proteome-wide datasets, we have identified an essential plasma membrane transporter of the major facilitator superfamily in T. gondii—previously termed TgApiAT6-1. Using an inducible system based on RNA degradation, TgApiAT6-1 was depleted, and the mutant parasite’s metabolome was compared to that of non-depleted parasites. The most significantly reduced metabolite in parasites depleted in TgApiAT6-1 was identified as the amino acid lysine, for which T. gondii is predicted to be auxotrophic. Using stable isotope-labeled amino acids, we confirmed that TgApiAT6-1 is required for efficient lysine uptake. Our findings highlight untargeted metabolomics as a powerful tool to identify the substrate of orphan transporters.

2019 ◽  
Vol 295 (3) ◽  
pp. 701-714 ◽  
Author(s):  
Aarti Krishnan ◽  
Joachim Kloehn ◽  
Matteo Lunghi ◽  
Dominique Soldati-Favre

The Apicomplexa phylum comprises diverse parasitic organisms that have evolved from a free-living ancestor. These obligate intracellular parasites exhibit versatile metabolic capabilities reflecting their capacity to survive and grow in different hosts and varying niches. Determined by nutrient availability, they either use their biosynthesis machineries or largely depend on their host for metabolite acquisition. Because vitamins cannot be synthesized by the mammalian host, the enzymes required for their synthesis in apicomplexan parasites represent a large repertoire of potential therapeutic targets. Here, we review recent advances in metabolic reconstruction and functional studies coupled to metabolomics that unravel the interplay between biosynthesis and salvage of vitamins and cofactors in apicomplexans. A particular emphasis is placed on Toxoplasma gondii, during both its acute and latent stages of infection.


2002 ◽  
Vol 184 (5) ◽  
pp. 1444-1448 ◽  
Author(s):  
Jayna L. Ditty ◽  
Caroline S. Harwood

ABSTRACT Charged amino acids in the predicted transmembrane portion of PcaK, a permease from Pseudomonas putida that transports 4-hydroxybenzoate (4-HBA), were required for 4-HBA transport, and they were also required for P. putida to have a chemotactic response to 4-HBA. An essential amino acid motif (DGXD) containing aspartate residues is located in the first transmembrane segment of PcaK and is conserved in the aromatic acid/H+ symporter family of the major facilitator superfamily of transporters.


Metabolites ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 61
Author(s):  
Aarti Krishnan ◽  
Dominique Soldati-Favre

Obligate intracellular pathogens have coevolved with their host, leading to clever strategies to access nutrients, to combat the host’s immune response, and to establish a safe niche for intracellular replication. The host, on the other hand, has also developed ways to restrict the replication of invaders by limiting access to nutrients required for pathogen survival. In this review, we describe the recent advancements in both computational methods and high-throughput –omics techniques that have been used to study and interrogate metabolic functions in the context of intracellular parasitism. Specifically, we cover the current knowledge on the presence of amino acid biosynthesis and uptake within the Apicomplexa phylum, focusing on human-infecting pathogens: Toxoplasma gondii and Plasmodium falciparum. Given the complex multi-host lifecycle of these pathogens, we hypothesize that amino acids are made, rather than acquired, depending on the host niche. We summarize the stage specificities of enzymes revealed through transcriptomics data, the relevance of amino acids for parasite pathogenesis in vivo, and the role of their transporters. Targeting one or more of these pathways may lead to a deeper understanding of the specific contributions of biosynthesis versus acquisition of amino acids and to design better intervention strategies against the apicomplexan parasites.


Author(s):  
Ana Paula Sato ◽  
Frederico Fontanelli Vaz ◽  
Aline Luiza Konell ◽  
Marilia de Oliveira Koch ◽  
Rafaela Furioso Ferreira ◽  
...  

Abstract Toxoplasma gondii, Neospora caninum and Sarcocystis neurona are obligate intracellular parasites within the phylum Apicomplexa. The red-tailed Amazon parrot (Amazona brasiliensis) is a near-threatened species of psittacine that is endemic to the Atlantic Forest of Brazil and has been designated as a bioindicator because of its sensitivity to environmental qualitative status and changes. The aim of this study was to evaluate the presence of antibodies against T. gondii, N. caninum and S. neurona in wild red-tailed Amazon parrot nestlings on Rasa Island, Brazil. Blood samples were collected from 51 parrots and plasma samples were stored at – 20 °C until immunofluorescence antibody tests (IFAT) were performed. Antigen slides were prepared using tachyzoites of T. gondii (RH strain) and, N. caninum (NC-1 strain) and using merozoites of S. neurona (SNR37 strain). Plasma samples were tested at initial dilutions of 1:16 for T. gondii, 1:50 for N. caninum and 1:5 for S. neurona. An anti-chicken antibody conjugated with FITC was used as a secondary antibody at 1:50 dilution. No antibodies for any of these three protozoa were found, thus suggesting that these wild red-tailed Amazon parrot nestlings had not been exposed to these parasites.


2002 ◽  
Vol 66 (1) ◽  
pp. 21-38 ◽  
Author(s):  
Naomi S. Morrissette ◽  
L. David Sibley

SUMMARY The Apicomplexa are a phylum of diverse obligate intracellular parasites including Plasmodium spp., the cause of malaria; Toxoplasma gondii and Cryptosporidium parvum, opportunistic pathogens of immunocompromised individuals; and Eimeria spp. and Theileria spp., parasites of considerable agricultural importance. These protozoan parasites share distinctive morphological features, cytoskeletal organization, and modes of replication, motility, and invasion. This review summarizes our current understanding of the cytoskeletal elements, the properties of cytoskeletal proteins, and the role of the cytoskeleton in polarity, motility, invasion, and replication. We discuss the unusual properties of actin and myosin in the Apicomplexa, the highly stereotyped microtubule populations in apicomplexans, and a network of recently discovered novel intermediate filament-like elements in these parasites.


2009 ◽  
Vol 78 (2) ◽  
pp. 651-660 ◽  
Author(s):  
Diana Marcela Penarete-Vargas ◽  
Marie Noelle Mévélec ◽  
Sarah Dion ◽  
Edouard Sèche ◽  
Isabelle Dimier-Poisson ◽  
...  

ABSTRACT Neospora caninum and Toxoplasma gondii are closely related, obligate intracellular parasites infecting a wide range of vertebrate hosts and causing abortion and neonatal morbidity and mortality. Several lines of evidence suggest that cross immunity between these two pathogens could be exploited in the design of strategies for heterologous vaccination. We assessed the ability of an attenuated strain of T. gondii (“mic1-3KO strain”) conferring strong protection against chronic and congenital toxoplasmosis to protect mice against lethal N. caninum infection. Mice immunized with mic1-3KO tachyzoites by the oral and intraperitoneal routes developed a strong cellular Th1 response and displayed significant protection against lethal heterologous N. caninum infection, with survival rates of 70% and 80%, respectively, whereas only 30% of the nonimmunized mice survived. We report here the acquisition of heterologous protective immunity against N. caninum following immunization with a live attenuated mic1-3KO strain of T. gondii.


2021 ◽  
Author(s):  
Stephen J Fairweather ◽  
Esther Rajendran ◽  
Martin Blume ◽  
Kiran Javed ◽  
Birte Steinhoefel ◽  
...  

Intracellular parasites of the phylum Apicomplexa are dependent on the scavenging of essential amino acids from their hosts. We previously identified a large family of apicomplexan-specific plasma membrane-localized amino acid transporters, the ApiATs, and showed that the Toxoplasma gondii transporter TgApiAT1 functions in the selective uptake of arginine. TgApiAT1 is essential for parasite virulence, but dispensable for parasite growth in medium containing high concentrations of arginine, indicating the presence of at least one other arginine transporter. Here we identify TgApiAT6-1 as the second arginine transporter. Using a combination of parasite assays and heterologous characterisation of TgApiAT6-1 in Xenopus laevis oocytes, we demonstrate that TgApiAT6-1 is a general cationic amino acid transporter that mediates both the high-affinity uptake of lysine and the low-affinity uptake of arginine. TgApiAT6-1 is the primary lysine transporter in the disease-causing tachyzoite stage of T. gondii and is essential for parasite proliferation. We demonstrate that the uptake of cationic amino acids by TgApiAT6-1 is "trans-stimulated" by cationic and neutral amino acids and is likely promoted by an inwardly negative membrane potential. These findings demonstrate that T. gondii has evolved overlapping transport mechanisms for the uptake of essential cationic amino acids, and we draw together our findings into a comprehensive model that highlights the finely-tuned, regulated processes that mediate cationic amino acid scavenging by these intracellular parasites.


2013 ◽  
Author(s):  
Ting-Kai Liu

Toxoplasma gondii is a kind of obligate intracellular parasites that are capable of infecting virtually all warm-blooded animals. It is one of the most common parasites in human. Serological studies estimated that up to a third of the global population has been chronically infected with the parsites. Toxoplasma gondii is also used as a model orgnisim of Apicomplexans that includes Plasmodium – the parasites that cause malaria. Gene knockout is a very important way to study gene function in all organisms. When it comes to Toxoplasma, it’s very difficult as this haploid parasite has strong adaptability to circumvent the condition of gene defects by gene duplication. High throughput screening at very early stage of transfection is very important for generating a true knockout of this parasite. For some genes that are required forToxoplasma gondii, the high throughput screening is necessary. Different labs use different protocols. After reading and practicing protocols from different papers or different labs, here I established an efficient pipeline for this purpose.


2013 ◽  
Author(s):  
Ting-Kai Liu

Toxoplasma gondii is a kind of obligate intracellular parasites that are capable of infecting virtually all warm-blooded animals. It is one of the most common parasites in human. Serological studies estimated that up to a third of the global population has been chronically infected with the parsites. Toxoplasma gondii is also used as a model orgnisim of Apicomplexans that includes Plasmodium – the parasites that cause malaria. Gene knockout is a very important way to study gene function in all organisms. When it comes to Toxoplasma, it’s very difficult as this haploid parasite has strong adaptability to circumvent the condition of gene defects by gene duplication. High throughput screening at very early stage of transfection is very important for generating a true knockout of this parasite. For some genes that are required forToxoplasma gondii, the high throughput screening is necessary. Different labs use different protocols. After reading and practicing protocols from different papers or different labs, here I established an efficient pipeline for this purpose.


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