bims-toxgon Biomed News
on Toxoplasma gondii metabolism
Issue of 2025–03–09
thirteen papers selected by
Lakesh Kumar, BITS Pilani



  1. Nat Commun. 2025 Mar 01. 16(1): 2095
      The apicoplast, a relic plastid organelle derived from secondary endosymbiosis, is crucial for many medically relevant Apicomplexa. While it no longer performs photosynthesis, the organelle retains several essential metabolic pathways. In this study, we examine the four primary metabolic pathways in the Toxoplasma gondii apicoplast, along with an accessory pathway, and identify conditions that can bypass these. Contrary to the prevailing view that the apicoplast is indispensable for T. gondii, we demonstrate that bypassing all pathways renders the apicoplast non-essential. We further show that T. gondii lacking an apicoplast (T. gondii-Apico) can be maintained indefinitely in culture, establishing a unique model to study the functions of this organelle. Through comprehensive metabolomic, transcriptomic, and proteomic analyses of T. gondii-Apico we uncover significant adaptation mechanisms following loss of the organelle and identify numerous putative apicoplast proteins revealed by their decreased abundance in T. gondii-Apico. Moreover, T. gondii-Apico parasites exhibit reduced sensitivity to apicoplast targeting compounds, providing a valuable tool for discovering new drugs acting on the organelle. The capability to culture T. gondii without its plastid offers new avenues for exploring apicoplast biology and developing novel therapeutic strategies against apicomplexan parasites.
    DOI:  https://doi.org/10.1038/s41467-025-57302-x
  2. Commun Biol. 2025 Mar 07. 8(1): 384
      Toxoplasma gondii, an intracellular pathogenic protist with a remarkable ability to infect a wide range of host cells, displays an equally exceptional design of its carbon metabolism. There are, however, critical gaps in our understanding of the metabolic network in T. gondii. We characterized the mito-nuclear metabolism and organelle coupling during its acute infection (lytic cycle). The major enzymes of the TCA cycle, i.e., citrate synthase (CS1), succinyl-CoA synthase alpha subunit (SCSα), succinate dehydrogenase (SDHA) and FAD malate dehydrogenase (MDH-FAD) located in the parasite mitochondrion support its asexual reproduction but are not needed for its survival. The SCSα and SDHA mutants are nearly avirulent in a mouse model, and they can protect the host against a lethal challenge infection. Genetic deletion of MDH-FAD dysregulated glucose-derived carbon flux, leading to a collapse of the mitochondrial membrane potential. The parasite also harbors a cytosolic isoform of MDH and a nuclear malic enzyme (ME) contributing to malate oxidation; however, only the latter is essential for the lytic cycle. Expression of ME in the nucleus is crucial for the parasite development. Besides, conditional knockdown of ME impairs the histone acetylation and disrupts the expression of several genes in tachyzoites. Our work discloses novel network design features of T. gondii and highlights the therapeutic and vaccination potential of the parasite metabolism.
    DOI:  https://doi.org/10.1038/s42003-025-07823-4
  3. J Biol Chem. 2025 Mar 03. pii: S0021-9258(25)00221-2. [Epub ahead of print] 108372
      The cytosolic Ca2+ concentration of all cells is highly regulated demanding the coordinated operation of Ca2+ pumps, channels, exchangers and binding proteins. In the protozoan parasite Toxoplasma gondii calcium homeostasis, essential for signaling, governs critical virulence traits. However, the identity of most molecular players involved in signaling and homeostasis in T. gondii are unknown or poorly characterized. In this work we studied a putative calcium proton exchanger, TgGT1_319550 (TgCAXL1), which belongs to a family of Ca2+/proton exchangers that localize to the Golgi apparatus. We localized TgCAXL1 to the Golgi and the endoplasmic reticulum (ER) of T. gondii and validated its role as a Ca2+/proton exchanger by yeast complementation. Characterization of a knock-out mutant for TgCAXL1 (Δcaxl) underscored the role of TgCAXL1 in Ca2+ storage by the ER and acidic stores, most likely the Golgi. Most interestingly, TgCAXL1 function is linked to the Ca2+ pumping activity of the Sarcoplasmic Reticulum Ca2+-ATPase (TgSERCA). TgCAXL1 functions in cytosolic pH regulation and recovery from acidic stress. Our data showed for the first time the role of the Golgi in storing and modulating Ca2+ signaling in T. gondii and the potential link between pH regulation and TgSERCA activity, which is essential for filling intracellular stores with Ca2+.
    Keywords:  Calcium signaling; SERCA; Toxoplasma gondii; calcium proton exchanger; pH homeostasis
    DOI:  https://doi.org/10.1016/j.jbc.2025.108372
  4. Chembiochem. 2025 Mar 06. e202500096
      Protein-based probes constructed via genetically encoding acetyl lysine (AcK) or its close analogs represent an important way to detect protein lysine deacetylases. Existing reported probes exhibit excellent sensitivity to NAD+-dependent sirtuins but lack responsiveness to Zn2+-dependent histone deacetylases (HDACs). Herein, we reformed the probe design by replacing the genetically encoded AcK with trifluoroacetyl lysine (TfAcK) and generated fluorescent and bioluminescent probes that could respond specifically to HDAC8 recombinantly expressed in E. coli and to endogenous HDACs in mammalian cells. We believe these probes would benefit the biological investigation of HDAC8 and promisingly some other HDACs, as well as the discovery of innovative HDAC inhibitors.
    Keywords:  HADC; Protein deacetylase; genetic code expansion; sirtuin; trifluoroacetyl lysine
    DOI:  https://doi.org/10.1002/cbic.202500096
  5. Nat Commun. 2025 Mar 06. 16(1): 2259
      cGMP signal-activated ookinete gliding is essential for mosquito midgut infection of Plasmodium in malaria transmission. During ookinete development, cGMP synthesizer GCβ polarizes to a unique localization "ookinete extrados site" (OES) until ookinete maturation and activates cGMP signaling for initiating parasite motility. However, the mechanism underlying GCβ translocation from cytosol to OES remains elusive. Here, we use protein proximity labeling to search the GCβ-interacting proteins in ookinetes of the rodent malaria parasite P. yoelii, and find the top hit Sir2A, a NAD+-dependent sirtuin family deacetylase. Sir2A interacts with GCβ throughout ookinete development. In mature ookinetes, Sir2A co-localizes with GCβ at OES in a mutually dependent manner. Parasites lacking Sir2A lose GCβ localization at OES, ookinete gliding, and mosquito infection, phenocopying GCβ deficiency. GCβ is acetylated at gametocytes but is deacetylated by Sir2A for OES localization at mature ookinetes. We further demonstrate that the level of NAD+, an essential co-substrate for sirtuin, increases during the ookinete development. NAD+ at its maximal level in mature ookinetes promotes Sir2A-catalyzed GCβ deacetylation, ensuring GCβ localization at OES. This study highlights the spatiotemporal coordination of cytosolic NAD+ level and NAD+-dependent Sir2A in regulating GCβ deacetylation and dynamic localization for Plasmodium ookinete gliding.
    DOI:  https://doi.org/10.1038/s41467-025-57517-y
  6. mSphere. 2025 Mar 04. e0004325
      Ben Liffner studies the cell biology of apicomplexan parasites. In this mSphere of Influence article, he reflects on two key papers: "Three-dimensional ultrastructure of Plasmodium falciparum throughout cytokinesis" by R. M. Rudlaff, S. Kraemer, J. Marshman, J. D. Dvorin, et al. (PLoS Pathog 16:e1008587, 2020, https://doi.org/10.1371/journal.ppat.1008587) and "Expansion microscopy provides new insights into the cytoskeleton of malaria parasites including the conservation of a conoid" by E. Bertiaux, A. C. Balestra, L. Bournonville, V. Louvel, et al. (PLoS Biol 19:e3001020, 2021, https://doi.org/10.1371/journal.pbio.3001020). These two studies provided Ben with the conceptual framework to understand how parasites are organized in three dimensions, and the technique of ultrastructure expansion microscopy that he has since used to investigate this intriguing area of biology.
    Keywords:  Plasmodium; apicomplexan parasites; electron microscopy; expansion microscopy
    DOI:  https://doi.org/10.1128/msphere.00043-25
  7. bioRxiv. 2025 Feb 20. pii: 2025.02.20.639246. [Epub ahead of print]
      Pyrophosphate is a byproduct of numerous cellular reactions that use ATP or other nucleoside triphosphates to synthesize DNA, RNA, protein, and other molecules. Its degradation into monophosphate is thus crucial for the survival and proliferation of all life forms. The human malaria parasite Plasmodium falciparum encodes two classes of pyrophosphatases to hydrolyze pyrophosphate. The first consists of P. falciparum proton pumping vacuolar pyrophosphatases (PfVP1 and PfVP2), which localize to the parasite's subcellular membranes and work as proton pumps. The second includes P. falciparum soluble pyrophosphatases (PfsPPases), which have not been well characterized. Interestingly, the gene locus of PfsPPase encodes two isoforms, PfsPPase1 (PF3D7_0316300.1) and PfsPPase2 (PF3D7_0316300.2). PfsPPase2 contains a 51- amino acid organellar localization peptide that is absent in PfsPPase1. Here, we combine reverse genetics and biochemical approaches to identify the localization of PfsPPase1 and PfsPPase2 and elucidate their individual functions. We show that PfsPPases are essential for the asexual blood stage. While PfsPPase1 solely localizes to the cytoplasm, PfsPPase2 exhibits multiple localizations including the mitochondrion, the apicoplast, and, to a lesser extent, the cytoplasm. Our data suggest that P. falciparum has taken a unique evolutionary trajectory in pyrophosphate metabolism by utilizing a leader sequence to direct sPPases to the mitochondrion and apicoplast. This differs from model eukaryotes as they generally encode multiple sPPases at distinct genetic loci to facilitate pyrophosphate degradation in cytosolic and organellar compartments. Our study highlights PfsPPases as promising targets for the development of novel antimalarial drugs.
    DOI:  https://doi.org/10.1101/2025.02.20.639246
  8. Parasites Hosts Dis. 2025 Feb;63(1): 37-49
      Cancer immunotherapy is widely used to treat various cancers to augment the weakened host immune response against tumors. Dendritic cells (DCs) are specialized antigen-presenting cells that play dual roles in inducing innate and adaptive immunity. Toxoplasma gondii is a protozoan parasite that exhibits anti-tumor activity against certain types of cancers. However, little is known about the anti-tumor effects of T. gondii or tumor/parasite antigen-pulsed DCs (DC vaccines, DCV) in breast cancer. In this study, C57BL/6 mice were administered E0771 mouse breast cancer cells (Cancer-injected) subcutaneously, T. gondii Me49 cysts orally (TG-injected), or DCs pulsed with breast cancer cell lysate antigen and T. gondii lysate antigens (DCV-injected) intraperitoneally. Tumor size and immunological characteristics were subsequently evaluated. We also evaluated matrix metalloproteinase (MMP)-2 and MMP-9 levels in E0771 mouse breast cancer cells co-cultured with T. gondii or DCs by RT-PCR. The tumor volumes of mice injected with breast cancer cells and antigen-pulsed DCs (Cancer/DCV-injected mice) were similar to those of Cancer-injected mice; however, they were significantly reduced in T. gondii-infected tumor-bearing (TG/Cancer-injected) mice. Moreover, tumor volumes were significantly reduced by adding antigen-pulsed DCs (TG/Cancer/DCV-injected mice) compared to TG/Cancer-injected mice. The levels of IFN-γ, serum IgG2a levels, and CD8+ T cell populations were significantly higher in DCV- and TG-injected mice than in control mice, while no significant differences between Cancer- and Cancer/DCV-injected mice were observed. The levels of IFN-γ, the IgG2a levels, and the percentage of CD8+ T cells were significantly increased in TG/Cancer- and TG/Cancer/DCV-injected mice than in Cancer-injected mice. IFN-γ levels and serum IgG2a levels were further increased in TG/Cancer/DCV-injected mice than in TG/Cancer-injected mice. The MMP-2 and MMP-9 mRNA expressions were significantly decreased in mouse breast cancer cells co-cultured with live T. gondii, T. gondii lysate antigen, or antigen-pulsed DCs (DCV) but not in inactivated DCs. These results indicate that T. gondii induces anti-tumor effects in breast cancer-bearing mice through the induction of strong Th1 immune responses, but not in antigen-pulsed DCs alone. The addition of antigen-pulsed DCs further augments the anti-tumor effects of T. gondii.
    Keywords:  Th1 immune response; Toxoplasma gondii; anti-tumor activity; breast neoplasms; cancer immunotherapy; dendritic cell vaccine
    DOI:  https://doi.org/10.3347/PHD.24082
  9. Biochem Biophys Res Commun. 2025 Feb 24. pii: S0006-291X(25)00249-9. [Epub ahead of print]754 151535
      Malate dehydrogenase 1 (MDH1) and isocitrate dehydrogenase 1 (IDH1) are two crucial enzymes in the process of energy metabolism. MDH1 plays a crucial role in the malate-aspartate shuttle in the cytoplasm by utilizing the coenzyme NAD/NADH to catalyze the interconversion of malate and oxaloacetate. IDH1 utilizes the coenzyme NADP/NADPH to facilitate the reciprocal transformation between isocitrate and α-ketoglutarate and plays a significant role in the metabolic processes of carbohydrates, lipids, and proteins in the liver. MDH1 and IDH1, along with their posttranslational modifications such as methylation and acetylation can influence the development of many diseases. This article reviews the function of MDH1, IDH1, and their posttranslational changes in various illnesses, aiming to offer new perspectives on disease diagnosis and therapy.
    Keywords:  Energy metabolism; Isocitrate dehydrogenase 1; Malate dehydrogenase 1; Posttranslational modification
    DOI:  https://doi.org/10.1016/j.bbrc.2025.151535
  10. mSphere. 2025 Mar 05. e0098324
      l-Serine and l-threonine have versatile roles in metabolism. In addition to their use in protein synthesis, these amino acids participate in the biosynthesis pathways of other amino acids and even phospholipids. Furthermore, l-serine and l-threonine can be substrates for a serine/threonine dehydratase (Ser/ThrDH), resulting in pyruvate and 2-oxobutyrate, respectively, thus being amino acids with anaplerotic potential. Trypanosoma cruzi, the etiological agent of Chagas disease, uses amino acids in several biological processes: metacyclogenesis, infection, resistance to nutritional and oxidative stress, osmotic control, etc. This study investigated the import and metabolism of l-serine, l-threonine, and glycine in T. cruzi. Our results demonstrate that these amino acids are transported from the extracellular environment into T. cruzi cells through a saturable transport system that fits the Michaelis-Menten model. Our results show that l-serine and l-threonine can sustain epimastigote cell viability under nutritional stress conditions and stimulate oxygen consumption, maintaining intracellular ATP levels. Additionally, our findings indicate that serine plays a role in establishing the mitochondrial membrane potential in T. cruzi. Serine is also involved in energy metabolism via the serine-pyruvate pathway, which stimulates the production and subsequent excretion of acetate and alanine. Our results demonstrate the importance of l-serine and l-threonine in the energy metabolism of T. cruzi and provide new insights into the metabolic adaptations of this parasite during its life cycle.IMPORTANCETrypanosoma cruzi, the parasite responsible for Chagas disease, impacts 5-6 million individuals in the Americas and is rapidly spreading globally due to significant human migration. This parasitic organism undergoes a complex life cycle involving triatomine insects and mammalian hosts, thriving in diverse environments, such as various regions within the insect's digestive tract and mammalian cell cytoplasm. Crucially, its transmission hinges on its adaptive capabilities to varying environments. One of the most challenging environments is the insect's digestive tract, marked by nutrient scarcity between blood meals, redox imbalance, and osmotic stresses induced by the triatomine's metabolism. To endure these conditions, T. cruzi has developed a remarkably versatile metabolic network enabling it to metabolize sugars, lipids, and amino acids efficiently. However, the full extent of metabolites this parasite can thrive on remains incompletely understood. This study reveals that, beyond conventional carbon and energy sources (glucose, palmitic acids, proline, histidine, glutamine, and alanine), three additional metabolites (serine, threonine, and glycine) play vital roles in the parasite's survival during starvation. Remarkably, serine and threonine directly contribute to ATP production through a serine/threonine dehydratase enzyme not previously described in T. cruzi. The significance of this metabolic pathway for the parasite's survival sheds light on how metabolic networks aid in its endurance under extreme conditions and its ability to thrive in diverse metabolic settings.
    Keywords:  Trypanosoma cruzi; amino acid metabolism; bioenergetics; nutritional stress; transport
    DOI:  https://doi.org/10.1128/msphere.00983-24
  11. J Mol Biol. 2025 Mar 03. pii: S0022-2836(25)00128-7. [Epub ahead of print] 169062
      Plasmodium falciparum plasmepsin X (PMX) has become a target of choice for the development of new antimalarial drugs due to its essential role across the parasite life cycle. Here we describe the 1.7Å crystallographic structure of PMX noncovalently bound to a potent macrocyclic peptidomimetic inhibitor (7k) possessing a hydroxyethylamine (HEA) scaffold. Upon 7k binding, the enzyme adopts a novel conformation, with significant involvement of the S2'S2 loop (M526-H536) and the S2 flap (F311-G314). This results in partial closure of the active site with widespread interactions in both the prime (S') and the non-prime (S) sites of PMX. The catalytic aspartate residues D266 and D467 directly interact with the HEA pharmacophore. Docking of a 7k derivative, compound 7a, highlights a region in the S3 pocket near the S3 flexible loop (H242-F248) that may be key for ligand stabilisation. The dynamic nature of PMX and its propensity to undergo distinct types of induced fit upon inhibitor binding enables generation of potent inhibitors that target this essential malarial aspartic protease.
    Keywords:  Plasmodium falciparum; aspartic protease; hydroxyethylamine; malaria; plasmepsin
    DOI:  https://doi.org/10.1016/j.jmb.2025.169062
  12. Nat Commun. 2025 Feb 28. 16(1): 2071
      Senescence-associated secretory phenotype (SASP) mediates the biological effects of senescent cells on the tissue microenvironment and contributes to ageing-associated disease progression. ACSS2 produces acetyl-CoA from acetate and epigenetically controls gene expression through histone acetylation under various circumstances. However, whether and how ACSS2 regulates cellular senescence remains unclear. Here, we show that pharmacological inhibition and deletion of Acss2 in mice blunts SASP and abrogates the pro-tumorigenic and immune surveillance functions of senescent cells. Mechanistically, ACSS2 directly interacts with and promotes the acetylation of PAICS, a key enzyme for purine biosynthesis. The acetylation of PAICS promotes autophagy-mediated degradation of PAICS to limit purine metabolism and reduces dNTP pools for DNA repair, exacerbating cytoplasmic chromatin fragment accumulation and SASP. Altogether, our work links ACSS2-mediated local acetyl-CoA generation to purine metabolism through PAICS acetylation that dictates the functionality of SASP, and identifies ACSS2 as a potential senomorphic target to prevent senescence-associated diseases.
    DOI:  https://doi.org/10.1038/s41467-025-57334-3
  13. PLoS One. 2025 ;20(3): e0308055
      Fluorescent proteins are widely used as markers to differentiate genetically modified cells from their wild-type counterparts. In malaria research, the prevalent fluorescent markers include red fluorescent proteins (RFPs) and their derivatives, such as mCherry, along with green fluorescent proteins (GFPs) and their derivatives. Recognizing the need for additional fluorescent markers to facilitate multiplexed imaging, this study introduced parasite lines expressing blue fluorescent protein (BFP). These lines enable simultaneous microscopy studies of proteins tagged with GFP, RFP, or detected by fluorophore-labeled antibodies, enhancing the analysis of complex biological interactions. Expression of BFP throughout the parasite's life cycle was driven by the robust Hsp70 promoter, ensuring stable, detectable protein levels suitable for fluorescent light analysis methods, including flow cytometry and fluorescent microscopy. We generated two Plasmodium berghei (P. berghei) lines expressing cytosolic BFP through double crossover homologous recombination targeting the silent 230p locus: eBFP2 (PbeBFP2) and mTagBFP2 (PbmTagBFP2). We compared these transgenic lines to established mCherry-expressing parasites PbmCherryHsp70 (PbmCherry) across their life cycles. The PbmTagBFP2 parasites exhibited fluorescence approximately 4.5 times brighter than the PbeBFP2 parasites in most life cycle stages. Both BFP-expressing lines developed normally through the entire parasite life cycle, offering a valuable expansion to the toolkit for studying Plasmodium biology at the host-pathogen interface.
    DOI:  https://doi.org/10.1371/journal.pone.0308055