bims-toxgon 2024-09-29 papers

bims-toxgon

Biomed News

on Toxoplasma gondii metabolism

Issue of 2024–09–29
twenty papers selected by
Lakesh Kumar, BITS Pilani

Toxoplasma acyl-CoA synthetase TgACS3 is crucial to channel host fatty acids in lipid droplets and for parasite propagation.
3-Benzylmenadiones and their Heteroaromatic Analogues Target the Apicoplast of Apicomplexa Parasites: Synthesis and Bioimaging Studies.
Evaluating the potential of non-immunosuppressive cyclosporin analogs for targeting Toxoplasma gondii cyclophilin: Insights from structural studies.
Toxoplasma gondii infection misdirects placental trophoblast lineage specification.
Oxygen-dependent regulation of F-box proteins in Toxoplasma gondii is mediated by Skp1 glycosylation.
Exploring similarities and differences between Toxoplasma gondii and Neospora caninum infections in dogs.
Heterologous immunization targeting the CST1 antigen confers better protection than ROP18 in mice.
Linking metabolism and histone acetylation dynamics by integrated metabolic flux analysis of Acetyl-CoA and histone acetylation sites.
Acetate drives ovarian cancer quiescence via ACSS2-mediated acetyl-CoA production.
Locked in Structure: Sestrin and GATOR-A Billion-Year Marriage.
Decoding the Role of Nuclear Sirtuins in Parkinson's Pathogenesis.
Metabolic regulation of cytoskeleton functions by HDAC6-catalyzed α-tubulin lactylation.
MmpL3, Wag31, and PlrA are involved in coordinating polar growth with peptidoglycan metabolism and nutrient availability.
tRNA lysidinylation is essential for the minimal translation system found in the apicoplast of Plasmodium falciparum.
The Role of Pyruvate Kinase M2 Posttranslational Modification in the Occurrence and Development of Hepatocellular Carcinoma.
Redox remodeling of central metabolism as a driving force for cellular protection, proliferation, differentiation, and dysfunction.
Pyruvate metabolism dictates fibroblast sensitivity to GLS1 inhibition during fibrogenesis.
The human touch: Utilizing AlphaFold 3 to analyze structures of endogenous metabolons.
Decreased sirtuin 4 levels promote cellular proliferation and invasion in papillary thyroid carcinoma.
The role of ATP citrate lyase in myelin formation and maintenance.

J Lipid Res. 2024 Sep 19. pii: S0022-2275(24)00150-0. [Epub ahead of print] 100645

Toxoplasma acyl-CoA synthetase TgACS3 is crucial to channel host fatty acids in lipid droplets and for parasite propagation.

Sheena Dass, Serena Shunmugam, Sarah Charital, Samuel Duley, Christophe-Sébastien Arnold, Nicholas J Katris, Pierre Cavaillès, Marie-France Cesbron-Delauw, Yoshiki Yamaryo-Botté, Cyrille Y Botté.

  Apicomplexa comprise important pathogenic parasitic protists that heavily depend on lipid acquisition to survive within their human host cells. Lipid synthesis relies on the incorporation of an essential combination of fatty acids (FAs) either generated by a metabolically adaptable de novo synthesis in the parasite or by scavenging from the host cell. The incorporation of FAs into membrane lipids depends on their obligate metabolic activation by specific enzyme groups, acyl-CoA synthetases (ACSs). Each ACS has its own specificity, so they can fulfill specific metabolic functions. Whilst such functionalities have been well studied in other eukaryotic models, their roles and importance in Apicomplexa is currently very limited, especially for Toxoplasma gondii. Here, we report the identification of 7 putative ACSs encoded by the genome of T. gondii (TgACS), which localize to different sub-cellular compartments of the parasite, suggesting exclusive functions. We show that the perinuclear/cytoplasmic TgACS3 regulates replication and growth of Toxoplasma tachyzoites. Conditional disruption of TgACS3 shows that the enzyme is required for parasite propagation and survival, especially under high host nutrient content. Lipidomic analysis of parasites lacking TgACS3 reveals its role in the activation of host-derived FAs that are used for i) parasite membrane phospholipid and ii) storage triacylglycerol (TAG) syntheses, allowing proper membrane biogenesis of parasite progenies. Altogether, our results reveal the role of TgACS3 as the bulk FA activator for membrane biogenesis allowing intracellular division and survival in T. gondii tachyzoites, further pointing at the importance of ACS and FA metabolism for the parasite.

Keywords:  Acyl-CoA synthetase; Apicomplexa; Lipid metabolism; Lipidomic; Toxoplasma gondii; fatty acid activation; host-parasite metabolic interactions; lipid-fatty acid fluxes; membrane biogenesis

DOI:  https://doi.org/10.1016/j.jlr.2024.100645
ACS Infect Dis. 2024 Sep 26.

3-Benzylmenadiones and their Heteroaromatic Analogues Target the Apicoplast of Apicomplexa Parasites: Synthesis and Bioimaging Studies.

Baptiste Dupouy, Maxime Donzel, Matthieu Roignant, Sarah Charital, Rodrigue Keumoe, Yoshiki Yamaryo-Botté, Alexander Feckler, Mirco Bundschuh, Yann Bordat, Matthias Rottmann, Pascal Mäser, Cyrille Y Botté, Stéphanie A Blandin, Sébastien Besteiro, Elisabeth Davioud-Charvet.

  The apicoplast is an essential organelle for the viability of apicomplexan parasites Plasmodium falciparum or Toxoplasma gondii, which has been proposed as a suitable drug target for the development of new antiplasmodial drug-candidates. Plasmodione, an antimalarial redox-active lead drug is active at low nM concentrations on several blood stages of Plasmodiumsuch as early rings and gametocytes. Nevertheless, its precise biological targets remain unknown. Here, we described the synthesis and the evaluation of new heteroaromatic analogues of plasmodione, active on asexual blood P. falciparum stages and T. gondii tachyzoites. Using a bioimaging-based analysis, we followed the morphological alterations of T. gondii tachyzoites and revealed a specific loss of the apicoplast upon drug treatment. Lipidomic and fluxomic analyses determined that drug treatment severely impacts apicoplast-hosted FASII activity in T. gondii tachyzoites, further supporting that the apicoplast is a primary target of plasmodione analogues. To follow the drug localization, "clickable" analogues of plasmodione were designed as tools for fluorescence imaging through a Cu(I)-catalyzed azide-alkyne cycloaddition reaction. Short-time incubation of two probes with P. falciparum trophozoites and T. gondii tachyzoites showed that the clicked products localize within, or in the vicinity of, the apicoplast of both Apicomplexa parasites. In P. falciparum, the fluorescence signal was also associated with the mitochondrion, suggesting that bioactivation and activity of plasmodione and related analogues are potentially associated with these two organelles in malaria parasites.

Keywords:  CuAAC-based imaging; Plasmodium falciparum; Toxoplasma gondii; apicoplast; lipidomics; plasmodione

DOI:  https://doi.org/10.1021/acsinfecdis.4c00304
Protein Sci. 2024 Oct;33(10): e5157

Evaluating the potential of non-immunosuppressive cyclosporin analogs for targeting Toxoplasma gondii cyclophilin: Insights from structural studies.

Filippo Favretto, Eva Jiménez-Faraco, Gianluca Catucci, Adele Di Matteo, Carlo Travaglini-Allocatelli, Sheila J Sadeghi, Paola Dominici, Juan A Hermoso, Alessandra Astegno.

  Toxoplasmosis persists as a prevalent disease, facing challenges from parasite resistance and treatment side effects. Consequently, identifying new drugs by exploring novel protein targets is essential for effective intervention. Cyclosporin A (CsA) possesses antiparasitic activity against Toxoplasma gondii, with cyclophilins identified as possible targets. However, CsA immunosuppressive nature hinders its use as an antitoxoplasmosis agent. Here, we evaluate the potential of three CsA derivatives devoid of immunosuppressive activity, namely, NIM811, Alisporivir, and dihydrocyclosporin A to target a previously characterized cyclophilin from Toxoplasma gondii (TgCyp23). We determined the X-ray crystal structures of TgCyp23 in complex with the three analogs and elucidated their binding and inhibitory properties. The high resolution of the structures revealed the precise positioning of ligands within the TgCyp23 binding site and the details of protein-ligand interactions. A comparison with the established ternary structure involving calcineurin indicates that substitutions at position 4 in CsA derivatives prevent calcineurin binding. This finding provides a molecular explanation for why CsA analogs can target Toxoplasma cyclophilins without compromising the human immune response.

Keywords:  Toxoplasma gondii; X‐ray crystal structure; cyclophilin inhibitors; cyclophilins; cyclosporin A; peptidyl‐prolyl isomerases

DOI:  https://doi.org/10.1002/pro.5157
bioRxiv. 2024 Sep 12. pii: 2024.09.10.612241. [Epub ahead of print]

Toxoplasma gondii infection misdirects placental trophoblast lineage specification.

Leah F Cabo, Liheng Yang, Mingze Gao, Rafaela J da Silva, NyJaee N Washington, Sarah M Reilly, Christina J Megli, Carolyn B Coyne, Jon P Boyle.

Pregnancy is a critical point of vulnerability to infection and other insults that could compromise proper fetal development. The placenta acts as a protective and nutrient-permeable barrier to most infectious agents, but a few are capable of bypassing its defenses. Remarkably little is known about how exposure to these select pathogens might impact ongoing placental development. Here we demonstrate that Toxoplasma gondii entirely misdirects the developmental program of trophoblast stem cells. Infection of progenitor cytotrophoblasts prevents fusion and differentiation to infection-resistant syncytiotrophoblast. Rather, T. gondii elicits a unique transcriptional identity that polarizes cytotrophoblasts to the infection-permissive extravillous trophoblast fate. Strong evidence of developmental disruption is found in multiple orthogonal models, including trophoblast stem cells, trophoblast organoids, and chorionic villi. Manipulation of cell fate by the parasite is most dramatic in trophoblast organoids, where we see robust outgrowth of HLA-G(+) extravillous trophoblasts. Collectively, these data show that Toxoplasma antagonizes differentiation of an infection-resistant cell type by inducing formation of an infection-permissive cell type, therefore potentiating its own transmission to the fetus.

DOI: https://doi.org/10.1101/2024.09.10.612241
J Biol Chem. 2024 Sep 20. pii: S0021-9258(24)02302-0. [Epub ahead of print] 107801

Oxygen-dependent regulation of F-box proteins in Toxoplasma gondii is mediated by Skp1 glycosylation.

Msano N Mandalasi, Elisabet Gas-Pascual, Carlos Gustavo Baptista, Bowen Deng, Hanke van der Wel, John A W Kruijtzer, Geert-Jan Boons, Ira J Blader, Christopher M West.

  A dynamic proteome is required for cellular adaption to changing environments including levels of O2, and the SKP1/CULLIN-1/F-box protein/RBX1 (SCF) family of E3 ubiquitin ligases contributes importantly to proteasome-mediated degradation. We examine, in the apicomplexan parasite Toxoplasma gondii, the influence on the interactome of SKP1 by its novel glycan attached to a hydroxyproline generated by PHYa, the likely ortholog of the HIFα PHD2 oxygen-sensor of human host cells. Strikingly, the representation of several putative F-box proteins (FBPs) is substantially reduced in PHYaΔ parasites grown in fibroblasts. One, FBXO13, is a predicted lysyl hydroxylase related to the human JmjD6 oncogene except for its F-box domain. The abundance of FBXO13, epitope-tagged at its genetic locus, was reduced in PHYaΔ parasites thus explaining its diminished presence in the SKP1 interactome. A similar effect was observed for FBXO14, a cytoplasmic protein of unknown function that may have co-evolved with PHYa in apicomplexans. Similar findings in glycosylation-mutant cells, rescue by proteasomal inhibitors, and unchanged transcript levels, suggested the involvement of the SCF in their degradation. The effect was selective, because FBXO1 was not affected by loss of PHYa. These findings are physiologically significant because the effects were phenocopied in parasites reared at 0.5% O2. Modest impact on steady-state SKP1 modification levels suggests that effects are mediated during a lag phase in hydroxylation of nascent SKP1. The dependence of FBP abundance on O2-dependent SKP1 modification likely contributes to the reduced virulence of PHYaΔ parasites owing to impaired ability to sense O2 as an environmental signal.

Keywords:  F-box protein; O-linked glycan; Toxoplasma; glycosyltransferase; hydroxyproline; oxygen-sensing; prolyl hydroxylase; ubiquitin ligase

DOI:  https://doi.org/10.1016/j.jbc.2024.107801
Vet Res Commun. 2024 Sep 25.

Exploring similarities and differences between Toxoplasma gondii and Neospora caninum infections in dogs.

Giulia Morganti, Giulia Rigamonti, Leonardo Brustenga, Valentina Calgaro, Giovanni Angeli, Iolanda Moretta, Manuela Diaferia, Fabrizia Veronesi.

  Toxoplasma gondii and Neospora caninum infections in dogs are predominantly manifest asymptomatic. However, these infections can also present highly varied and potentially severe clinical signs. This is due to the parasites' ability to replicate in a number of cell types within the host organism, with N. caninum exhibiting a particular tropism for the central and peripheral nervous systems, and T. gondii targeting the central nervous system and musculature. In clinical practice, toxoplasmosis and neosporosis are often considered to be closely related diseases, despite their distinct epidemiological, clinical, diagnostic, and therapeutic characteristics. The present review analyses the similarities and differences between these two protozoan infections, since an accurate and timely aetiological diagnosis is essential for establishing effective therapeutic protocols and control strategies.

Keywords:   Neospora caninum ; Toxoplasma gondii ; Differences; Dog; Similarities

DOI:  https://doi.org/10.1007/s11259-024-10549-z
Nanomedicine (Lond). 2024 Sep 25. 1-10

Heterologous immunization targeting the CST1 antigen confers better protection than ROP18 in mice.

Gi-Deok Eom, Ki Back Chu, Jie Mao, Keon-Woong Yoon, Hae-Ji Kang, Eun-Kyung Moon, Sung Soo Kim, Fu-Shi Quan.

  Aim: To evaluate the protective efficacy induced by heterologous immunization with recombinant baculoviruses or virus-like particles targeting the CST1 and ROP18 antigens of Toxoplasma gondii.Materials & methods: Recombinant baculovirus and virus-like particle vaccines expressing T. gondii CST1 or ROP18 antigens were developed to evaluate protective immunity in mice upon challenge infection with 450 Toxoplasma gondii (ME49).Results: Immunization with CST1 or ROP18 vaccines induced similar levels of T. gondii-specific IgG and IgA responses. Compared with ROP 18, CST1 vaccine showed better antibody-secreting cell response, germinal center B cell activation, and significantly reduced brain cyst burden and body weight loss.Conclusion: Our findings suggest that CST1 heterologous immunization elicited better protection than ROP18, providing important insight into improving the toxoplasmosis vaccine design strategy.

Keywords:  CST1; ROP18; Toxoplasma gondii; heterologous immunization; recombinant baculovirus; virus-like particle

DOI:  https://doi.org/10.1080/17435889.2024.2403333
Mol Metab. 2024 Sep 19. pii: S2212-8778(24)00163-7. [Epub ahead of print] 102032

Linking metabolism and histone acetylation dynamics by integrated metabolic flux analysis of Acetyl-CoA and histone acetylation sites.

Anna-Sophia Egger, Eva Rauch, Suraj Sharma, Tobias Kipura, Madlen Hotze, Thomas Mair, Alina Hohenegg, Philipp Kobler, Ines Heiland, Marcel Kwiatkowski.

  Histone acetylation is an important epigenetic modification that regulates various biological processes and cell homeostasis. Acetyl-CoA, a hub molecule of metabolism, is the substrate for histone acetylation, thus linking metabolism with epigenetic regulation. However, still relatively little is known about the dynamics of histone acetylation and its dependence on metabolic processes, due to the lack of integrated methods that can capture site-specific histone acetylation and deacetylation reactions together with the dynamics of acetyl-CoA synthesis. In this study, we present a novel proteo-metabo-flux approach that combines mass spectrometry-based metabolic flux analysis of acetyl-CoA and histone acetylation with computational modelling. We developed a mathematical model to describe metabolic label incorporation into acetyl-CoA and histone acetylation based on experimentally measured relative abundances. We demonstrate that our approach is able to determine acetyl-CoA synthesis dynamics and site-specific histone acetylation and deacetylation reaction rate constants, and that consideration of the metabolically labelled acetyl-CoA fraction is essential for accurate determination of histone acetylation dynamics. Furthermore, we show that without correction, changes in metabolic fluxes would be misinterpreted as changes in histone acetylation dynamics, whereas our proteo-metabo-flux approach allows to distinguish between the two processes.

Keywords:  Computational modelling; Epigenetics; Histone modifications; LC-MS; Metabolic flux analysis; Metabolism

DOI:  https://doi.org/10.1016/j.molmet.2024.102032
Mol Metab. 2024 Sep 19. pii: S2212-8778(24)00162-5. [Epub ahead of print]89 102031

Acetate drives ovarian cancer quiescence via ACSS2-mediated acetyl-CoA production.

Allison C Sharrow, Emily Megill, Amanda J Chen, Afifa Farooqi, Naveen Kumar Tangudu, Apoorva Uboveja, Stacy McGonigal, Nadine Hempel, Nathaniel W Snyder, Ronald J Buckanovich, Katherine M Aird.

  Quiescence is a reversible cell cycle exit traditionally thought to be associated with a metabolically inactive state. Recent work in muscle cells indicates that metabolic reprogramming is associated with quiescence. Whether metabolic changes occur in cancer to drive quiescence is unclear. Using a multi-omics approach, we found that the metabolic enzyme ACSS2, which converts acetate into acetyl-CoA, is both highly upregulated in quiescent ovarian cancer cells and required for their survival. Indeed, quiescent ovarian cancer cells have increased levels of acetate-derived acetyl-CoA, confirming increased ACSS2 activity in these cells. Furthermore, either inducing ACSS2 expression or supplementing cells with acetate was sufficient to induce a reversible quiescent cell cycle exit. RNA-Seq of acetate treated cells confirmed negative enrichment in multiple cell cycle pathways as well as enrichment of genes in a published G0 gene signature. Finally, analysis of patient data showed that ACSS2 expression is upregulated in tumor cells from ascites, which are thought to be more quiescent, compared to matched primary tumors. Additionally, high ACSS2 expression is associated with platinum resistance and worse outcomes. Together, this study points to a previously unrecognized ACSS2-mediated metabolic reprogramming that drives quiescence in ovarian cancer. As chemotherapies to treat ovarian cancer, such as platinum, have increased efficacy in highly proliferative cells, our data give rise to the intriguing question that metabolically-driven quiescence may affect therapeutic response.

Keywords:  ACSS2; Cell cycle; G0 phase; Metabolism

DOI:  https://doi.org/10.1016/j.molmet.2024.102031
Cells. 2024 Sep 21. pii: 1587. [Epub ahead of print]13(18):

Locked in Structure: Sestrin and GATOR-A Billion-Year Marriage.

Alexander Haidurov, Andrei V Budanov.

  Sestrins are a conserved family of stress-responsive proteins that play a crucial role in cellular metabolism, stress response, and ageing. Vertebrates have three Sestrin genes (SESN1, SESN2, and SESN3), while invertebrates encode only one. Initially identified as antioxidant proteins that regulate cell viability, Sestrins are now recognised as crucial inhibitors of the mechanistic target of rapamycin complex 1 kinase (mTORC1), a central regulator of anabolism, cell growth, and autophagy. Sestrins suppress mTORC1 through an inhibitory interaction with the GATOR2 protein complex, which, in concert with GATOR1, signals to inhibit the lysosomal docking of mTORC1. A leucine-binding pocket (LBP) is found in most vertebrate Sestrins, and when bound with leucine, Sestrins do not bind GATOR2, prompting mTORC1 activation. This review examines the evolutionary conservation of Sestrins and their functional motifs, focusing on their origins and development. We highlight that the most conserved regions of Sestrins are those involved in GATOR2 binding, and while analogues of Sestrins exist in prokaryotes, the unique feature of eukaryotic Sestrins is their structural presentation of GATOR2-binding motifs.

Keywords:  GATOR2; SESN2; conservation; mTORC1

DOI:  https://doi.org/10.3390/cells13181587
ACS Chem Neurosci. 2024 Sep 27.

Decoding the Role of Nuclear Sirtuins in Parkinson's Pathogenesis.

Dishank Patel, Ritu Soni, Jigna Shah.

  Parkinson's disease (PD) is the second most prevailing degenerative disease that deals with dopaminergic neuronal loss and deficiency of dopamine in SNpc and striatum. Manifestations primarily include motor symptoms like tremor, rigidity, and akinesia/dyskinesia along with some nonmotor symptoms like GI and olfactory dysfunction. α-Synuclein pathogenesis is the major cause behind progression of PD; however there are many underlying molecular mechanisms behind the pathophysiology of PD. Sirtuins are small molecular deacetylases that have an imperative role in pathology of such neurodegenerative disorders like PD. Sirtuins are majorly classified according to their location; nuclear (SIRT1,7,6), mitochondrial sirtuins (SIRT3-5), and cytosolic (SIRT2). These actively take part in pathological development and possess independent actions. In this review, the role of nuclear sirtuins is individualistically explored for better understanding of PD pathology and development of advanced therapeutics targeting sirtuins.

Keywords:  Parkinson’s disease; SIRT1; SIRT6; SIRT7; nuclear sirtuins; sirtuins

DOI:  https://doi.org/10.1021/acschemneuro.4c00507
Nat Commun. 2024 Sep 27. 15(1): 8377

Metabolic regulation of cytoskeleton functions by HDAC6-catalyzed α-tubulin lactylation.

Shuangshuang Sun, Zhe Xu, Liying He, Yihui Shen, Yuqing Yan, Xubing Lv, Xujing Zhu, Wei Li, Wei-Ya Tian, Yongjun Zheng, Sen Lin, Yadong Sun, Lei Li.

Posttranslational modifications (PTMs) of tubulin, termed the "tubulin code", play important roles in regulating microtubule functions within subcellular compartments for specialized cellular activities. While numerous tubulin PTMs have been identified, a comprehensive understanding of the complete repertoire is still underway. In this study, we report that α-tubulin lactylation is catalyzed by HDAC6 by using lactate to increase microtubule dynamics in neurons. We identify lactylation on lysine 40 of α-tubulin in the soluble tubulin dimers. Notably, lactylated α-tubulin enhances microtubule dynamics and facilitates neurite outgrowth and branching in cultured hippocampal neurons. Moreover, we discover an unexpected function of HDAC6, acting as the primary lactyltransferase to catalyze α-tubulin lactylation. HDAC6-catalyzed lactylation is a reversible process, dependent on lactate concentrations. Intracellular lactate concentration triggers HDAC6 to lactylate α-tubulin, a process dependent on its deacetylase activity. Additionally, the lactyltransferase activity may be conserved in HDAC family proteins. Our study reveals the primary role of HDAC6 in regulating α-tubulin lactylation, establishing a link between cell metabolism and cytoskeleton functions.

DOI: https://doi.org/10.1038/s41467-024-52729-0
J Bacteriol. 2024 Sep 25. e0020424

MmpL3, Wag31, and PlrA are involved in coordinating polar growth with peptidoglycan metabolism and nutrient availability.

Neda Habibi Arejan, Desiree R Czapski, Joseph A Buonomo, Cara C Boutte.

  Cell growth in mycobacteria involves cell wall expansion that is restricted to the cell poles. The DivIVA homolog Wag31 is required for this process, but the molecular mechanism and protein partners of Wag31 have not been described. In this study of Mycobacterium smegmatis, we identify a connection between wag31 and trehalose monomycolate (TMM) transporter mmpl3 in a suppressor screen and show that Wag31 and polar regulator PlrA are required for MmpL3's polar localization. In addition, the localization of PlrA and MmpL3 is responsive to nutrient and energy deprivation and inhibition of peptidoglycan metabolism. We show that inhibition of MmpL3 causes delocalized cell wall metabolism but does not delocalize MmpL3 itself. We found that cells with an MmpL3 C-terminal truncation, which is defective for localization, have only minor defects in polar growth but are impaired in their ability to downregulate cell wall metabolism under stress. Our work suggests that, in addition to its established function in TMM transport, MmpL3 has a second function in regulating global cell wall metabolism in response to stress. Our data are consistent with a model in which the presence of TMMs in the periplasm stimulates polar elongation and in which the connection between Wag31, PlrA, and the C-terminus of MmpL3 is involved in detecting and responding to stress in order to coordinate the synthesis of the different layers of the mycobacterial cell wall in changing conditions.
IMPORTANCE: This study is performed in Mycobacterium smegmatis, which is used as a model to understand the basic physiology of pathogenic mycobacteria such as Mycobacterium tuberculosis. In this work, we examine the function and regulation of three proteins involved in regulating cell wall elongation in mycobacterial cells, which occurs at the cell tips or poles. We find that Wag31, a regulator of polar elongation, works partly through the regulation of MmpL3, a transporter of cell wall constituents and an important drug target. Our work suggests that, beyond its transport function, MmpL3 has another function in controlling cell wall synthesis broadly in response to stress.

Keywords:  DivIVA; MmpL3; Mycobacterium; PlrA; TMMs; murJ; mycolic acid; peptidoglycan; polar growth regulation; stress

DOI:  https://doi.org/10.1128/jb.00204-24
bioRxiv. 2024 Sep 14. pii: 2024.09.13.612944. [Epub ahead of print]

tRNA lysidinylation is essential for the minimal translation system found in the apicoplast of Plasmodium falciparum.

Rubayet Elahi, Sean T Prigge.

For decades, researchers have sought to define minimal genomes to elucidate the fundamental principles of life and advance biotechnology. tRNAs, essential components of this machinery, decode mRNA codons into amino acids. The apicoplast of malaria parasites encodes 25 tRNA isotypes in its organellar genome - the lowest number found in known translation systems. Efficient translation in such minimal systems depends heavily on post-transcriptional tRNA modifications, especially at the wobble anticodon position. Lysidine modification at the wobble position (C34) of tRNA CAU distinguishes between methionine (AUG) and isoleucine (AUA) codons, altering the amino acid delivered by this tRNA and ensuring accurate protein synthesis. Lysidine is formed by the enzyme tRNA isoleucine lysidine synthetase (TilS) and is nearly ubiquitous in bacteria and essential for cellular viability. We identified a TilS ortholog ( Pf TilS) located in the apicoplast of Plasmodium falciparum parasites. By complementing Pf TilS with a bacterial ortholog, we demonstrated that the lysidinylation activity of Pf TilS is critical for parasite survival and apicoplast maintenance, likely due to its impact on apicoplast protein translation. Our findings represent the first characterization of TilS in an endosymbiotic organelle, advancing eukaryotic organelle research and our understanding of minimal translational machinery. Due to the absence of lysidine modifications in humans, this research also exposes a potential vulnerability in malaria parasites that could be targeted by antimalarial strategies.
Significance: In recent decades, synthetic biologists have sought the minimal cellular components required for life, focusing on simpler systems for easier modeling. The apicoplast organelle of malaria parasites, with only 25 tRNA isotypes, contains the smallest known complete tRNA set, even smaller than in synthetic organisms. This makes it an ideal model for studying minimal translational machinery, where tRNAs depend on post-transcriptional modifications for efficient protein translation. A key modification, lysidine, is crucial for decoding isoleucine and methionine. This study describes a tRNA-isoleucine lysidine synthetase (TilS) enzyme, essential for apicoplast protein translation. These findings have implications for understanding eukaryotic organelles and minimal translation machinery. Additionally, the absence of lysidine in humans suggests a potential target for antimalarial strategies.

DOI: https://doi.org/10.1101/2024.09.13.612944
Cell Biochem Funct. 2024 Sep;42(7): e4125

The Role of Pyruvate Kinase M2 Posttranslational Modification in the Occurrence and Development of Hepatocellular Carcinoma.

Zhao Chunlian, Wan Qi, Zhao Rui.

  Hepatocellular carcinoma (HCC) is one of the deadly malignant tumors that directly leads to the death of nearly one million people worldwide every year, causing a serious burden on society. In the presence of sufficient oxygen, HCC cells rapidly generate energy through aerobic glycolysis, which promotes tumor cell proliferation, immune evasion, metastasis, angiogenesis, and drug resistance. Pyruvate kinase M2 (PKM2) is a key rate-limiting enzyme in glycolysis. In recent years, studies have found that PKM2 not only exerts pyruvate kinase activity in the process of glucose metabolism, but also exerts protein kinase activity in non-metabolic pathways to affect tumor cell processes, and its activity is flexibly regulated by various posttranslational modifications such as acetylation, phosphorylation, lactylation, ubiquitination, SUMOylation, and so forth. This review summarizes the role of posttranslational modifications of PKM2-related sites in the development of HCC.

Keywords:  glycolysis; hepatocellular carcinoma; posttranslational modification; pyruvate kinase M2; tumorigenesis and development

DOI:  https://doi.org/10.1002/cbf.4125
Free Radic Res. 2024 Sep 24. 1-24

Redox remodeling of central metabolism as a driving force for cellular protection, proliferation, differentiation, and dysfunction.

Junichi Fujii.

  The production of reactive oxygen species (ROS) is elevated via metabolic hyperactivation in response to a variety of stimuli such as growth factors and inflammation. Tolerable amounts of ROS moderately inactivate enzymes via oxidative modification, which can be reversed back to the native form in a redox-dependent manner. The excessive production of ROS, however, causes cell dysfunction and death. Redox-reactive enzymes are present in primary metabolic pathways such as glycolysis and the tricarboxylic acid cycle, and these act as floodgates for carbon flux. Oxidation of a specific form of cysteine inhibits glyceraldehyde-3-phosphate dehydrogenase, which is reversible, and causes an accumulation of upstream intermediary compounds that increases the flux of glucose-6-phosphate to the pentose phosphate pathway. These reactions increase the NADPH and ribose-5-phosphate that are available for reductive reactions and nucleotide synthesis, respectively. On the other hand, oxidative inactivation of mitochondrial aconitase increases citrate, which is then recruited to synthesize fatty acids in the cytoplasm. Decreases in the use of carbohydrate for ATP production can be compensated via amino acid catabolism, and this metabolic change makes nitrogen available for nucleic acid synthesis. Coupling of the urea cycle also converts nitrogen to urea and polyamine, the latter of which supports cell growth. This metabolic remodeling stimulates the proliferation of tumor cells and fibrosis in oxidatively damaged tissues. Oxidative modification of these enzymes is generally reversible in the early stages of oxidizing reactions, which suggests that early treatment with appropriate antioxidants promotes the maintenance of natural metabolism.

Keywords:  Glycolysis; TCA cycle; metabolic remodeling; reactive sulfhydryl; urea cycle

DOI:  https://doi.org/10.1080/10715762.2024.2407147
JCI Insight. 2024 Aug 13. pii: e178453. [Epub ahead of print]9(18):

Pyruvate metabolism dictates fibroblast sensitivity to GLS1 inhibition during fibrogenesis.

Greg Contento, Jo-Anne Am Wilson, Brintha Selvarajah, Manuela Platé, Delphine Guillotin, Valle Morales, Marcello Trevisani, Vanessa Pitozzi, Katiuscia Bianchi, Rachel C Chambers.

  Fibrosis is a chronic disease characterized by excessive extracellular matrix production, which leads to disruption of organ function. Fibroblasts are key effector cells of this process, responding chiefly to the pleiotropic cytokine transforming growth factor-β1 (TGF-β1), which promotes fibroblast to myofibroblast differentiation. We found that extracellular nutrient availability profoundly influenced the TGF-β1 transcriptome of primary human lung fibroblasts and that biosynthesis of amino acids emerged as a top enriched TGF-β1 transcriptional module. We subsequently uncovered a key role for pyruvate in influencing glutaminase (GLS1) inhibition during TGF-β1-induced fibrogenesis. In pyruvate-replete conditions, GLS1 inhibition was ineffective in blocking TGF-β1-induced fibrogenesis, as pyruvate can be used as the substrate for glutamate and alanine production via glutamate dehydrogenase (GDH) and glutamic-pyruvic transaminase 2 (GPT2), respectively. We further show that dual targeting of either GPT2 or GDH in combination with GLS1 inhibition was required to fully block TGF-β1-induced collagen synthesis. These findings embolden a therapeutic strategy aimed at additional targeting of mitochondrial pyruvate metabolism in the presence of a glutaminolysis inhibitor to interfere with the pathological deposition of collagen in the setting of pulmonary fibrosis and potentially other fibrotic conditions.

Keywords:  Cell biology; Collagens; Fibrosis; Glucose metabolism; Metabolism

DOI:  https://doi.org/10.1172/jci.insight.178453
Structure. 2024 Sep 17. pii: S0969-2126(24)00335-6. [Epub ahead of print]

The human touch: Utilizing AlphaFold 3 to analyze structures of endogenous metabolons.

Toni K Träger, Christian Tüting, Panagiotis L Kastritis.

Computational structural biology aims to accurately predict biomolecular complexes with AlphaFold 3 spearheading the field. However, challenges loom for structural analysis, especially when complex assemblies such as the pyruvate dehydrogenase complex (PDHc), which catalyzes the link reaction in cellular respiration, are studied. PDHc subcomplexes are challenging to predict, particularly interactions involving weaker, lower-affinity subcomplexes. Supervised modeling, i.e., integrative structural biology, will continue to play a role in fine-tuning this type of prediction (e.g., removing clashes, rebuilding loops/disordered regions, and redocking interfaces). 3D analysis of endogenous metabolic complexes continues to require, in addition to AI, precise and multi-faceted interrogation methods.

DOI: https://doi.org/10.1016/j.str.2024.08.018
Eur Thyroid J. 2024 Oct 01. pii: e240079. [Epub ahead of print]13(5):

Decreased sirtuin 4 levels promote cellular proliferation and invasion in papillary thyroid carcinoma.

Hyun-Jin Lee, Young-Sool Hah, So Young Cheon, Seong Jun Won, Chae Dong Yim, Somi Ryu, Seung-Jun Lee, Ji Hyun Seo, Jung Je Park.

   Objective: This study examined the effect of sirtuin 4 (SIRT4), a NAD+-dependent deacetylase, on the proliferation and progression of papillary thyroid carcinoma (PTC).
Methods: Data from The Cancer Genome Atlas (TCGA) were analyzed to identify SIRT4 expression in thyroid cancer. Subsequently, the correlation between SIRT4 expression and clinical characteristics was examined in 205 PTC tissue samples. In vitro assays using three human thyroid cancer cell lines (B-CPAP, TPC-1, and SNU-790) were conducted to assess the effects of regulated SIRT4 expression on cell growth, apoptosis, invasion, and migration. Furthermore, in vivo experiments were performed in a xenograft mouse model.
Results: Gene Expression Omnibus (GEO) and TCGA data indicated that SIRT4 expression is lower in thyroid cancer and SIRT4 downregulation is associated with poor overall survival. In PTC tissues, positive SIRT4 expression was associated with decreased extracapsular extension. In in vitro experiments using three human thyroid cancer cell lines, overexpression of SIRT4 decreased cell survival, clonogenic potential, and invasion and migratory capabilities, as well as inducing apoptosis and increasing reactive oxygen species levels. SIRT4 overexpression upregulated E-cadherin and downregulated N-cadherin, suggesting its potential involvement in the regulation of epithelial-mesenchymal transition. These findings were confirmed in vivo using a xenograft mouse model.
Conclusion: This study provides novel insight into the potential contribution of SIRT4 to the regulation of the pathological progression of PTC. The data suggest that SIRT4 plays a tumor-suppressive role in PTC by inhibiting growth, survival, and invasive potential. Future research should investigate the molecular mechanisms underlying these effects of SIRT4.

Keywords:  SIRT4 protein; epithelial–mesenchymal transition; papillary thyroid cancer; reactive oxygen species; thyroid cancer

DOI:  https://doi.org/10.1530/ETJ-24-0079
Glia. 2024 Sep 25.

The role of ATP citrate lyase in myelin formation and maintenance.

Andrew Schneider, Seongsik Won, Eric A Armstrong, Aaron J Cooper, Amulya Suresh, Rachell Rivera, Gregory Barrett-Wilt, John M Denu, Judith A Simcox, John Svaren.

  Formation of myelin by Schwann cells is tightly coupled to peripheral nervous system development and is important for neuronal function and long-term maintenance. Perturbation of myelin causes a number of specific disorders that are among the most prevalent diseases affecting the nervous system. Schwann cells synthesize myelin lipids de novo rather than relying on uptake of circulating lipids, yet one unresolved matter is how acetyl CoA, a central metabolite in lipid formation is generated during myelin formation and maintenance. Recent studies have shown that glucose-derived acetyl CoA itself is not required for myelination. However, the importance of mitochondrially-derived acetyl CoA has never been tested for myelination in vivo. Therefore, we have developed a Schwann cell-specific knockout of the ATP citrate lyase (Acly) gene to determine the importance of mitochondrial metabolism to supply acetyl CoA in nerve development. Intriguingly, the ACLY pathway is important for myelin maintenance rather than myelin formation. In addition, ACLY is required to maintain expression of a myelin-associated gene program and to inhibit activation of the latent Schwann cell injury program.

Keywords:  Schwann; acetyl CoA; lipid; lipidomic; myelin

DOI:  https://doi.org/10.1002/glia.24620