bims-toxgon 2025-02-23 papers

bims-toxgon

Biomed News

on Toxoplasma gondii metabolism

Issue of 2025–02–23
nine papers selected by
Lakesh Kumar, BITS Pilani

Toxoplasma gondii PROP1 is critical for autophagy and parasite viability during chronic infection.
Effect of the flavonoid compound glabridin on tachyzoites and bradyzoites of Toxoplasma gondii.
Mechanism-Based Inhibition of Histone Deacetylase 6 by a Selenocyanate Is Subject to Redox Modulation.
Genetic and functional dissection of the glutamate-proline pathway reveals a shortcut for glutamate catabolism in Leishmania.
Phylogenomics of coral-infecting corallicolids reveal multiple independent losses of chlorophyll biosynthesis in apicomplexan parasites.
Plasmodium blood stage development requires the chromatin remodeller Snf2L.
Conformational flexibility associated with remote residues regulates the kinetic properties of glutamate dehydrogenase.
Mitochondrial glutamic-oxaloacetic transaminase (GOT2) in the growth of C2C12 myoblasts.
SFP6 fluorescent probes for imaging SAM dynamics in living cells.

mSphere. 2025 Feb 21. e0082924

Toxoplasma gondii PROP1 is critical for autophagy and parasite viability during chronic infection.

Pariyamon Thaprawat, Fengrong Wang, Shreya Chalasani, Tracey L Schultz, Manlio Di Cristina, Vern B Carruthers.

  Macroautophagy is an important cellular process involving lysosomal degradation of cytoplasmic components, facilitated by autophagy-related proteins. In the protozoan parasite Toxoplasma gondii, autophagy has been demonstrated to play a key role in adapting to stress and the persistence of chronic infection. Despite limited knowledge about the core autophagy machinery in T. gondii, two PROPPIN family proteins (TgPROP1 and TgPROP2) have been identified with homology to Atg18/WIPI. Prior research in acute-stage tachyzoites suggests that TgPROP2 is predominantly involved in a non-autophagic function, specifically apicoplast biogenesis, while TgPROP1 may be involved in canonical autophagy. Here, we investigated the distinct roles of TgPROP1 and TgPROP2 in chronic stage T. gondii bradyzoites, revealing a critical role for TgPROP1, but not TgPROP2, in bradyzoite autophagy. Conditional knockdown of TgPROP2 did not impair bradyzoite autophagy. In contrast, TgPROP1 KO parasites had impaired autolysosome formation, reduced cyst burdens in chronically infected mice, and decreased viability. Together, our findings clarify the indispensable role of TgPROP1 to T. gondii autophagy and chronic infection.
IMPORTANCE: It is estimated that up to a third of the human population is chronically infected with Toxoplasma gondii; however, little is known about how this parasite persists long term within its hosts. Autophagy is a self-eating pathway that has recently been shown to play a key role in parasite persistence, yet few proteins that carry out this process during T. gondii chronic infection are known. Here, we provide evidence for a non-redundant role of TgPROP1, a protein important in the early steps of the autophagy pathway. Genetic disruption of TgPROP1 resulted in impaired autophagy and chronic infection of mice. Our results reveal a critical role for TgPROP1 in autophagy and underscore the importance of this pathway in parasite persistence.

Keywords:  Toxoplasma gondii; autophagy; bradyzoite; persistance

DOI:  https://doi.org/10.1128/msphere.00829-24
Parasit Vectors. 2025 Feb 17. 18(1): 56

Effect of the flavonoid compound glabridin on tachyzoites and bradyzoites of Toxoplasma gondii.

Yanhua Qiu, Weiwei Wang, Qing Wang, Hongling Lin, Yubin Bai, Jiyu Zhang.

   BACKGROUND: Toxoplasma gondii (T. gondii) is one of the most prevalent parasites worldwide. At present, the majority of drugs used for the treatment of toxoplasmosis target the tachyzoite stage of T. gondii and are largely ineffective against bradyzoites. Furthermore, these treatments are typically accompanied by adverse events. Consequently, there is an urgent need for the development of novel drugs that are both safe and effective against T. gondii.
METHODS: A total of 20 flavonoids were preliminarily screened for their anti-T. gondii activity using microscopy. Next, the cell counting kit (CCK)-8 method was employed to assess the toxicity of glabridin (GLA) to host cells, while the RH strain of T.0gondii, which expresses β-galactosidase, was utilized to evaluate the inhibitory, anti-invasive, and antiproliferative effects of GLA on T. gondii. In addition, the Prugniaud (PRU) strain was employed to investigate the impact of GLA on the bradyzoites of T. gondii. Subsequently, the effect of GLA on the ultrastructure of T. gondii was examined via transmission electron microscopy (TEM), followed by an assessment of the influence of GLA on the autophagy and mitochondria of T. gondii through monodansylcadaverine (MDC), MitoTracker™ red CMXRos, and CM-HDCFDA and MitoSOX Red staining.
RESULTS: Among the 20 flavonoids assessed, GLA exhibited the most potent anti-T. gondii activity. Indeed, it significantly inhibited both the invasive and proliferative abilities of T. gondii, thereby disrupting its lytic cycle. Moreover, GLA markedly reduced the number of bradyzoites and concurrently inhibited cyst growth. Meanwhile, ultrastructural analysis revealed that GLA induced mitochondrial swelling, membrane rupture, and autophagy in T. gondii. Finally, fluorescent probe staining provided further evidence that GLA triggers mitochondrial dysfunction and autophagy in this parasite.
CONCLUSIONS: Our findings collectively indicate that the flavonoid compound GLA exhibits significant activity against both T. gondii tachyzoites and bradyzoites. The underlying mechanism of action potentially involves the induction of autophagy and mitochondrial dysfunction and the disruption of the membrane of T. gondii, thereby offering new avenues for treating toxoplasmosis and establishing a theoretical reference for future research.

Keywords:   Toxoplasma gondii ; Autophagy; Flavonoid compound; Glabridin; Membrane disruption; Mitochondrial dysfunction

DOI:  https://doi.org/10.1186/s13071-025-06695-1
J Am Chem Soc. 2025 Feb 17.

Mechanism-Based Inhibition of Histone Deacetylase 6 by a Selenocyanate Is Subject to Redox Modulation.

Juana Goulart Stollmaier, Briana Abigail R Czarnecki, David W Christianson.

Organoselenocyanates have attracted considerable attention in recent years due to their therapeutic potential and versatility in medicinal chemistry. Here, we report on the mechanism of inhibition by 5-phenylcarbamoylpentyl selenocyanide (SelSA-2), an analogue of the well-characterized histone deacetylase inhibitor suberoylanilide hydroxamic acid (SAHA, a.k.a. Vorinostat). We show that histone deacetylases 6 and 10 promote selenocyanate hydrolysis to generate a selenolate anion, and we explore the redox chemistry of selenium as it modulates inhibitory activity through reversible formation of the diselenide. The 2.15 Å-resolution crystal structure of histone deacetylase 6 cocrystallized with SelSA-2 conclusively demonstrates that it is not the selenocyanate, but instead a zinc-bound selenolate anion, that is the active pharmacophore responsible for enzyme inhibition.

DOI: https://doi.org/10.1021/jacs.5c00157
FEBS J. 2025 Feb 17.

Genetic and functional dissection of the glutamate-proline pathway reveals a shortcut for glutamate catabolism in Leishmania.

Gustavo Daniel Campagnaro, Angela Kaysel Cruz.

  Trypanosomatids are early-divergent eukaryotes that have adapted to parasitism. During their life cycles, these parasites switch between a mammalian and an invertebrate host, and the ability to adapt their metabolism to different nutritional sources is instrumental for their success. In the invertebrate host, these protists have access to high amounts of amino acids and efficiently utilise it for energy production. Proline is a particularly efficient energy source for trypanosomes. Glutamate is also efficiently used by Trypanosoma cruzi and can be converted into proline as part of the glutamate-proline pathway prior to its intramitochondrial catabolism. By employing a series of genetic modifications and functional analysis, we show here that Leishmania parasites, the causative agents of leishmaniases, can utilise proline, glutamate and glutamine as energy sources, and although these parasites possess all the genes necessary for the biosynthesis of proline from glutamate, this pathway has, at best, limited function, with at least one of its components (pyrroline-5-carboxylate reductase) assuming divergent functions in different life cycle stages of the parasite. In fact, we show that the catabolism of glutamate is independent of proline biosynthesis and the former is most likely directly imported into the mitochondrion and catabolised to recover the cellular redox metabolism and increase mitochondrial membrane potential. Moreover, our data suggest a relevant role for glutamate dehydrogenase in nutritional stress response in Leishmania. These findings highlight relevant differences in amino acid metabolism between Trypanosoma and Leishmania and suggest a diversification in amino acid metabolic pathways within Trypanosomatidae.

Keywords:  Leishmania; glutamate–proline pathway; mitochondrial metabolism; moonlight activity; proline–glutamate

DOI:  https://doi.org/10.1111/febs.70030
Curr Biol. 2025 Feb 11. pii: S0960-9822(25)00058-2. [Epub ahead of print]

Phylogenomics of coral-infecting corallicolids reveal multiple independent losses of chlorophyll biosynthesis in apicomplexan parasites.

Victoria K L Jacko-Reynolds, Waldan K Kwong, Samuel J Livingston, Morelia Trznadel, Anthony M Bonacolta, Gordon Lax, Jade Shivak, Nicholas A T Irwin, Mark J A Vermeij, Javier Del Campo, Patrick J Keeling.

  The transition from free-living to parasitic lifestyles induces major shifts in evolution, and nowhere is this more acute than in apicomplexans-obligate intracellular parasites of animals that evolved from photosynthetic algae.1 In other cases where photosynthesis has been lost, including most apicomplexans, chlorophyll is also absent, but in coral-infecting apicomplexans (corallicolids), chlorophyll biosynthesis genes are retained in the plastid genome despite their lack of photosystems.2 This suggests that the loss of photosynthesis and chlorophyll were decoupled in this lineage, but because these observations are only based on plastid genomes, two fundamental questions remain unclear. First, how this impacted apicomplexan evolution as a whole is unclear because there are conflicting phylogenetic positions for corallicolids: plastid gene phylogenies place them at the base of the apicomplexans, whereas nuclear rRNA places them with late-branching coccidians (suborder Eimeriorina).2,3 Second, it is unclear if chlorophyll or a metabolic intermediate is synthesized, as most chlorophyll biosynthesis enzymes are encoded in the nucleus. To address these questions, we have sequenced transcriptomes from two corallicolids, infecting Parazoanthus swiftii and Madracis mirabilis hosts. Phylogenomic data strongly support a late-branching relationship closer with coccidians, specifically with the protococcidians and the newly discovered ichthyocolids. We also find evidence for the expression of nucleus-encoded enzymes involved in chlorophyll biosynthesis in corallicolids and protococcidians. Overall, we conclude that chlorophyll synthesis was likely retained through the early evolution of the group and then lost approximately 10 times independently, emphasizing the impact of parallel evolutionary changes in parasitic transitions.

Keywords:  Apicomplexa; chlorophyll; coral; corallicolids; evolution; parasite; photosynthesis; phylogenetics; plastid

DOI:  https://doi.org/10.1016/j.cub.2025.01.028
Nature. 2025 Feb 19.

Plasmodium blood stage development requires the chromatin remodeller Snf2L.

Maria Theresia Watzlowik, Elisabeth Silberhorn, Sujaan Das, Ritwik Singhal, Kannan Venugopal, Simon Holzinger, Barbara Stokes, Ella Schadt, Lauriane Sollelis, Victoria A Bonnell, Matthew Gow, Andreas Klingl, Matthias Marti, Manuel Llinás, Markus Meissner, Gernot Längst.

The complex life cycle of the malaria parasite Plasmodium falciparum involves several major differentiation stages, each requiring strict control of gene expression. Fundamental changes in chromatin structure and epigenetic modifications during life cycle progression suggest a central role for these mechanisms in regulating the transcriptional program of malaria parasite development1-6. P. falciparum chromatin is distinct from other eukaryotes, with an extraordinarily high AT content (>80%)7 and highly divergent histones resulting in atypical DNA packaging properties8. Moreover, the chromatin remodellers that are critical for shaping chromatin structure are not conserved and are unexplored in P. falciparum. Here we identify P. falciparum Snf2L (PfSnf2L, encoded by PF3D7_1104200) as an ISWI-related ATPase that actively repositions P. falciparum nucleosomes in vitro. Our results demonstrate that PfSnf2L is essential, regulating both asexual development and sexual differentiation. PfSnf2L globally controls just-in-time transcription by spatiotemporally determining nucleosome positioning at the promoters of stage-specific genes. The unique sequence and functional properties of PfSnf2L led to the identification of an inhibitor that specifically kills P. falciparum and phenocopies the loss of correct gene expression timing. The inhibitor represents a new class of antimalarial transmission-blocking drugs, inhibiting gametocyte formation.

DOI: https://doi.org/10.1038/s41586-025-08595-x
Protein Sci. 2025 Mar;34(3): e70038

Conformational flexibility associated with remote residues regulates the kinetic properties of glutamate dehydrogenase.

Barsa Kanchan Jyotshna Godsora, Parijat Das, Prasoon Kumar Mishra, Anjali Sairaman, Sandip Kaledhonkar, Narayan S Punekar, Prasenjit Bhaumik.

  Glutamate dehydrogenase (GDH) is a pivotal metabolic enzyme in all living organisms, and some of the GDHs exhibit substrate-dependent homotropic cooperativity. However, the mode of allosteric communication during the homotropic effect in GDHs remains poorly understood. In this study, we examined two homologous GDHs, Aspergillus niger GDH (AnGDH) and Aspergillus terreus GDH (AtGDH), with differing substrate utilization kinetics to uncover the factors driving their distinct behavior. We report the crystal structures and first-ever cryo-EM structures of apo- AtGDH and AnGDH that captured arrays of conformational ensembles. A wider mouth opening (~ 21 Å) is observed for the cooperative AnGDH as compared to the non-cooperative AtGDH (~17 Å) in their apo states. A network of interactions related to the substitutions in Domain II influence structural flexibility in these GDHs. Remarkably, we have identified a distant substitution (R246 to S) in Domain II, as a part of this network, which can impact the mouth opening and converts non-cooperative AtGDH into a cooperative enzyme. Our study demonstrates that remote residues can influence structural and kinetic properties in homologous GDHs.

Keywords:  allostery; cryo‐EM; crystal structure; glutamate dehydrogenase; homotropic cooperativity; intrinsic domain dynamics; kinetic properties; remote residue substitution

DOI:  https://doi.org/10.1002/pro.70038
J Bioenerg Biomembr. 2025 Feb 15.

Mitochondrial glutamic-oxaloacetic transaminase (GOT2) in the growth of C2C12 myoblasts.

Ritu Som, Brian D Fink, Adam J Rauckhorst, Eric B Taylor, William I Sivitz.

  Glutamine is well recognized as critical to the growth of most cell types. Within mitochondria glutamine is converted to glutamate by glutaminase. Oxaloacetate and glutamate then react to form alpha-ketoglutarate (α-KG) and aspartate catalyzed by glutamic-oxaloacetic transaminase (GOT2) or directly converted to α-KG by glutamate dehydrogenase (GDH). We investigated the role of GOT2 in mediating glutamate metabolism and cell growth in undifferentiated C2C12 cells. CRISPR mediated GOT2 knockout (KO) impaired cell growth, partially overcome by higher concentrations of glutamine. Mitochondrial respiration did not differ between KO and wildtype (WT) cells. Metabolite profiling showed that GOT2KO decreased aspartate by about 50% in KO versus WT cells. In contrast, α-KG increased. Metabolites reflecting the pentose phosphate pathway were significantly increased in KO cells. Metabolic pathway analyses revealed alteration of the TCA cycle, the pentose phosphate pathway, and amino acid metabolism. Glutamine 13C-tracing revealed decreased generation of aspartate, increased ribulose phosphate and evidence for reductive carboxylation of α-KG to isocitrate in KO cells. GDH expression was detected in C2C12 cells but did not differ between WT and GOT2KO mitochondria. GDH is not or barely expressed in adult muscle, however, we observed clear expression in pre-weanling mice. Cytosolic glutamic-oxaloacetic transaminase, GOT1, expression did not differ between GOT2KO and WT cells. In summary, GOT2 is necessary for glutamate flux and generation of downstream metabolites needed for the growth of C2C12 myoblasts. Although respiration did not differ, lack of aspartate and other compounds needed for cell proliferation may have been major factors impairing growth.

Keywords:  Glutamate; Glutamate dehydrogenase; Glutamic oxaloacetic transaminase; Mitochondria; Myoblasts; Skeletal muscle

DOI:  https://doi.org/10.1007/s10863-025-10053-2
Mikrochim Acta. 2025 Feb 21. 192(3): 180

SFP6 fluorescent probes for imaging SAM dynamics in living cells.

Shuhui Zhang, Jinghui Li, Gengsheng Cao.

  A genetically encoded probe, SFP6 (S-adenosyl-L-methionine fluorescent probe), based on the principle of fluorescence resonance energy transfer (FRET) was developed. The SFP6 probe dynamically visualizes changes in S-adenosyl-L-methionine (SAM) levels in living cells with high spatiotemporal resolution. The results demonstrated that SFP6 exhibits high sensitivity to SAM, can be stably expressed in various mammalian cells, and has excellent biocompatibility. The probe accurately monitors SAM levels and detects changes caused by both endogenous and exogenous factors. In summary, we have developed a fluorescent probe that can monitor changes in SAM levels with single-cell and time resolution. Dynamic changes in SAM levels are linked to various methylation modifications in cells. Therefore, monitoring intracellular SAM concentrations offers the possibility to study physiological and biochemical processes in real-time, such as gene expression and metabolism, related to methylation modifications.

Keywords:  FRET; Live cell imaging; SAM; SFP6 fluorescent probe

DOI:  https://doi.org/10.1007/s00604-025-07039-7