bims-toxgon 2024-02-25 papers

bims-toxgon

Biomed News

on Toxoplasma gondii metabolism

Issue of 2024–02–25
thirty-two papers selected by
Lakesh Kumar, BITS Pilani

Immunization with Live-Attenuated RHΔhad2a Strain Confers Partial Protective Immunity against Acute and Chronic Infection of Toxoplasma gondii in Mice.
Two Toxoplasma gondii putative pore-forming proteins, GRA47 and GRA72, influence small molecule permeability of the parasitophorous vacuole.
Calreticulin (CALR) promotes ionophore-induced microneme secretion in Toxoplasma gondii.
IFNs in host defence and parasite immune evasion during Toxoplasma gondii infections.
Regulation of Calcium entry by cyclic GMP signaling in Toxoplasma gondii.
Hypermigration of macrophages through the concerted action of GRA effectors on NF-κB/p38 signaling and host chromatin accessibility potentiates Toxoplasma dissemination.
First Description of Marinoquinoline Derivatives' Activity against Toxoplasma gondii.
The IRAK1/IRF5 axis initiates IL-12 response by dendritic cells and control of Toxoplasma gondii infection.
Parasite-induced IFN-γ regulates host defense via CD115 and mTOR-dependent mechanism of tissue-resident macrophage death.
Allosteric strand displacement isothermal amplification for the visual detection of Toxoplasma gondii in 30 minutes.
Application of gold immunochromatographic assay strip combined with digital evaluation for early detection of Toxoplasma gondii infection in multiple species.
Prevalence of Toxoplasma gondii in Wild American Mink (Neogale vison): The First Serological Study in Germany and Poland.
TORC1 is an essential regulator of nutrient-controlled proliferation and differentiation in Leishmania.
Bacteria employ lysine acetylation of transcriptional regulators to adapt gene expression to cellular metabolism.
mTORC1 activity oscillates throughout the cell cycle promoting mitotic entry and differentially influencing autophagy induction.
The bloodstream form of Trypanosoma brucei displays non-canonical gluconeogenesis.
ER remodeling via lipid metabolism.
Histone lysine acetyltransferase inhibitors: an emerging class of drugs for cancer therapy.
Sirtuins as Players in the Signal Transduction of Citrus Flavonoids.
SUMO Modifies GβL and Mediates mTOR Signaling.
Identification and characterization of nuclear localization signals in the circumsporozoite protein of Plasmodium falciparum.
The enzymes of serine synthesis pathway in cancer metastasis.
Acetyl-CoA synthetase (ACSS2) does not generate butyryl- and crotonyl-CoA.
Exploring the potential of EphA2 receptor signaling pathway: a comprehensive review in cancer treatment.
Going through changes - the role of autophagy during reprogramming and differentiation.
Association of changes in expression of HDAC and SIRT genes after drug treatment with cancer cell line sensitivity to kinase inhibitors.
Structural characterization of the ACDC domain from ApiAP2 proteins of the malaria parasite.
αKG-mediated carnitine synthesis promotes homologous recombination via histone acetylation.
Gene Expression of Ethanol and Acetate Metabolic Pathways in the Acinetobacter baumannii EmaSR Regulon.
Sirtuins Affect Cancer Stem Cells via Epigenetic Regulation of Autophagy.
Impact of Lysine Succinylation on the Biology of Fungi.
Protein succinylation: regulating metabolism and beyond.

Pathogens. 2024 Jan 28. pii: 121. [Epub ahead of print]13(2):

Immunization with Live-Attenuated RHΔhad2a Strain Confers Partial Protective Immunity against Acute and Chronic Infection of Toxoplasma gondii in Mice.

Hai-Sheng Zhang, Hui Cao, Chen-Xu Li, Zhi-Wei Zhang, Meng Wang, Xing-Quan Zhu, Xiao-Nan Zheng.

  Toxoplasmosis caused by Toxoplasma gondii is an important zoonosis of human and animal health significance. Current chemical therapeutics have side effects, and no commercially available vaccine is licensed for the prevention of toxoplasmosis in humans and most animals. Developing a safe and effective vaccine with long-term protection against T. gondii infection is necessary to control toxoplasmosis. HAD2a is a key member of the haloacid dehalogenase (HAD) phosphatase family, which is essential for T. gondii daughter budding. However, the role of HAD2a in T. gondii virulence remains unknown. In this study, we successfully constructed the had2a gene knockout strain in the T. gondii-type I RH strain (RHΔhad2a) and determined its role in virulence and vaccination. These results demonstrate that HAD2a played an important role in parasite daughter budding and in vitro replication. Knockout of the had2a gene attenuated the virulence of the T. gondii-type I RH strain. Vaccination with RHΔhad2a tachyzoites induced a Th1-biased immune response, provided partial protection against acute T. gondii infection in mice by highly virulent tachyzoites of RH and PYS (ToxoDB#9, Chinese I) strains, and conferred strong protection against challenge infection by cysts and oocysts of the less virulent type II Pru strain. These results demonstrate that T. gondii had2a is important for its in vitro proliferation and virulence in mice and that RHΔhad2a may be used as a candidate strain to generate a multiple gene knockout live-attenuated strain or be collaboratively applied with other live-attenuated strains to confer more effective protection against T. gondii infection.

Keywords:  CRISPR-Cas9; HAD phosphatase; Toxoplasma gondii; live-attenuated strain; toxoplasmosis; vaccine

DOI:  https://doi.org/10.3390/pathogens13020121
mBio. 2024 Feb 21. e0308123

Two Toxoplasma gondii putative pore-forming proteins, GRA47 and GRA72, influence small molecule permeability of the parasitophorous vacuole.

Mebratu A Bitew, Pablo S Gaete, Christopher Swale, Parag Maru, Jorge E Contreras, Jeroen P J Saeij.

  Toxoplasma gondii, a medically important intracellular parasite, uses GRA proteins secreted from dense granule organelles to mediate nutrient flux across the parasitophorous vacuole membrane (PVM). GRA17 and GRA23 are known pore-forming proteins on the PVM involved in this process, but the roles of additional proteins have remained largely uncharacterized. We recently identified GRA72 as synthetically lethal with GRA17. Deleting GRA72 produced similar phenotypes to Δgra17 parasites, and computational predictions suggested it forms a pore. To understand how GRA72 functions, we performed immunoprecipitation experiments and identified GRA47 as an interactor of GRA72. Deletion of GRA47 resulted in an aberrant "bubble vacuole" morphology with reduced small molecule permeability, mirroring the phenotype observed in GRA17 and GRA72 knockouts. Structural predictions indicated that GRA47 and GRA72 form heptameric and hexameric pores, respectively, with conserved histidine residues lining the pore. Mutational analysis highlighted the critical role of these histidines for protein functionality. Validation through electrophysiology confirmed alterations in membrane conductance, corroborating their pore-forming capabilities. Furthermore, Δgra47 parasites and parasites expressing GRA47 with a histidine mutation had reduced in vitro proliferation and attenuated virulence in mice. Our findings show the important roles of GRA47 and GRA72 in regulating PVM permeability, thereby expanding the repertoire of potential therapeutic targets against Toxoplasma infections.IMPORTANCEToxoplasma gondii is a parasite that poses significant health risks to those with impaired immunity. It replicates inside host cells shielded by the PVM, which controls nutrient and waste exchange with the host. GRA72, previously identified as essential in the absence of the GRA17 nutrient channel, is implicated in forming an alternative nutrient channel. Here we found that GRA47 associates with GRA72 and is also important for the PVM's permeability to small molecules. Removal of GRA47 leads to distorted vacuoles and impairs small molecule transport across the PVM, resembling the effects of GRA17 and GRA72 deletions. Structural models suggest GRA47 and GRA72 form distinct pore structures, with a pore-lining histidine critical to their function. Toxoplasma strains lacking GRA47 or those with a histidine mutation have impaired growth and reduced virulence in mice, highlighting these proteins as potential targets for new treatments against toxoplasmosis.

Keywords:  GRA47; GRA72; PVM permeability; Toxoplasma gondii; Xenopus oocytes; pore; pore-lining histidine

DOI:  https://doi.org/10.1128/mbio.03081-23
Parasitol Res. 2024 Feb 21. 123(2): 139

Calreticulin (CALR) promotes ionophore-induced microneme secretion in Toxoplasma gondii.

Zhili Shan, Xingju Song, Xu Yang, Yangfei Xue, Yayun Wu, Xianmei Wang, Jing Liu, Qun Liu.

  The flow of calcium ions (Ca2+) is involved in numerous vital activities of Toxoplasma gondii. Calreticulin is a type of Ca2+-binding protein in the endoplasmic reticulum (ER) that is involved in Ca2+ signaling pathway regulation, Ca2+ storage, and protein folding. In this work, the calreticulin (CALR), a protein predicted to possess a conserved domain of calreticulin in T. gondii, was characterized. The CALR localized in the ER. Using reverse genetics, we discovered that CALR is not necessary for the lytic cycle, including invasion and replication. However, depletion of CALR affected microneme secretion triggered by A23187, which is a Ca2+ ionophore used to increase cytoplasmic Ca2+ concentration. Furthermore, we discovered that CALR influences Ca2+ release. Transcriptomic comparison between Δcalr and Δku80 parasites showed that 226 genes in the Δcalr parasites were significantly downregulated (p < 0.05). The cellular biological functions of the downregulated genes were mainly involved in calmodulin-dependent protein kinase pathways. Furthermore, in the absence of CALR, tachyzoites were still able to cause acute infection in mice. These results imply that by influencing ER Ca2+ release content, CALR may further impair the ionophore-induced secretion of the parasite. However, this protein is not required for the completion of the parasite's lytic cycle or for the acute virulence of the parasite.

Keywords:  Calreticulin; Ionophore-induced secretion; Microneme proteins; Toxoplasma gondii

DOI:  https://doi.org/10.1007/s00436-024-08162-9
Front Immunol. 2024 ;15 1356216

IFNs in host defence and parasite immune evasion during Toxoplasma gondii infections.

Carsten G K Lüder.

  Interferons (IFNs) are a family of cytokines with diverse functions in host resistance to pathogens and in immune regulation. Type II IFN, i.e. IFN-γ, is widely recognized as a major mediator of resistance to intracellular pathogens, including the protozoan Toxoplasma gondii. More recently, IFN-α/β, i.e. type I IFNs, and IFN-λ (type III IFN) have been identified to also play important roles during T. gondii infections. This parasite is a widespread pathogen of humans and animals, and it is a model organism to study cell-mediated immune responses to intracellular infection. Its success depends, among other factors, on the ability to counteract the IFN system, both at the level of IFN-mediated gene expression and at the level of IFN-regulated effector molecules. Here, I review recent advances in our understanding of the molecular mechanisms underlying IFN-mediated host resistance and immune regulation during T. gondii infections. I also discuss those mechanisms that T. gondii has evolved to efficiently evade IFN-mediated immunity. Knowledge of these fascinating host-parasite interactions and their underlying signalling machineries is crucial for a deeper understanding of the pathogenesis of toxoplasmosis, and it might also identify potential targets of parasite-directed or host-directed supportive therapies to combat the parasite more effectively.

Keywords:  Toxoplasma gondii; host defence; immune evasion; immunity; interferons; parasite effectors; signalling

DOI:  https://doi.org/10.3389/fimmu.2024.1356216
J Biol Chem. 2024 Feb 19. pii: S0021-9258(24)00147-9. [Epub ahead of print] 105771

Regulation of Calcium entry by cyclic GMP signaling in Toxoplasma gondii.

Miryam A Hortua Triana, Karla M Márquez-Nogueras, Mojtaba Sedigh Fazli, Shannon Quinn, Silvia N J Moreno.

  Ca2+ signaling impacts almost every aspect of cellular life. Ca2+ signals are generated through the opening of ion channels that permit the flow of Ca2+ down an electrochemical gradient. Cytosolic Ca2+ fluctuations can be generated through Ca2+ entry from the extracellular milieu or release from intracellular stores. In Toxoplasma gondii, Ca2+ ions play critical roles in several essential functions for the parasite like invasion of host cells, motility and egress. Plasma membrane Ca2+ entry in T. gondii was previously shown to be activated by cytosolic calcium and inhibited by the voltage-operated Ca2+ channel blocker nifedipine. However, Ca2+ entry in T. gondii did not show the classical characteristics of store regulation. In this work, we characterized the mechanism by which cytosolic Ca2+ regulates plasma membrane Ca2+ entry in extracellular T. gondii tachyzoites loaded with the Ca2+ indicator Fura 2. We compared the inhibition by nifedipine with the effect of the broad spectrum TRP channel inhibitor, anthranilic acid or ACA and we find that both inhibitors act on different Ca2+ entry activities. We demonstrate, using pharmacological and genetic tools, that an intracellular signaling pathway engaging cyclicGMP (cGMP), protein kinase G (PKG), Ca2+ and the phosphatidyl inositol phospholipase C (PI-PLC) affects Ca2+ entry and we present a model for crosstalk between cGMP and cytosolic Ca2+ for the activation of T. gondii's lytic cycle traits.

Keywords:  Calcium; Toxoplasma gondii; cGMP; egress; invasion; protein kinase G; signaling

DOI:  https://doi.org/10.1016/j.jbc.2024.105771
bioRxiv. 2024 Feb 06. pii: 2024.02.06.579146. [Epub ahead of print]

Hypermigration of macrophages through the concerted action of GRA effectors on NF-κB/p38 signaling and host chromatin accessibility potentiates Toxoplasma dissemination.

Arne L Ten Hoeve, Matias E Rodriguez, Martin Säflund, Valentine Michel, Lucas Magimel, Albert Ripoll, Tianxiong Yu, Mohamed-Ali Hakimi, Jeroen P J Saeij, Deniz M Ozata, Antonio Barragan.

Mononuclear phagocytes facilitate the dissemination of the obligate intracellular parasite Toxoplasma gondii . Here, we report how a set of secreted parasite effector proteins from dense granule organelles (GRA) orchestrates dendritic cell-like chemotactic and pro-inflammatory activation of parasitized macrophages. These effects enabled efficient dissemination of the type II T. gondii lineage, a highly prevalent genotype in humans. We identify novel functions for effectors GRA15 and GRA24 in promoting CCR7-mediated macrophage chemotaxis by acting on NF-κB and p38 MAPK signaling pathways, respectively, with contributions of GRA16/18 and counter-regulation by effector TEEGR. Further, GRA28 boosted chromatin accessibility and GRA15/24/NF-κB-dependent transcription at the Ccr7 gene locus in primary macrophages. In vivo , adoptively transferred macrophages infected with wild-type T. gondii outcompeted macrophages infected with a GRA15/24 double mutant in migrating to secondary organs in mice. The data show that T. gondii , rather than being passively shuttled, actively promotes its dissemination by inducing a finely regulated pro-migratory state in parasitized human and murine phagocytes via co-operating polymorphic GRA effectors.
Importance: Intracellular pathogens can hijack cellular functions of infected host cells to their advantage, for example, for intracellular survival and for dissemination. However, how microbes orchestrate the hijacking of complex cellular processes, such as host cell migration, remains poorly understood. As such, the common parasite Toxoplasma gondii actively invades immune cells of humans and other vertebrates and modifies their migratory properties. Here, we show that the concerted action of a number of secreted effector proteins from the parasite, principally GRA15 and GRA24, act on host cell signaling pathways to activate chemotaxis. Further, the protein effector GRA28 selectively acted on chromatin accessibility in the host cell nucleus to selectively boost host gene expression. The joint activities of effectors culminated in pro-migratory signaling within the infected phagocyte. We provide a molecular framework delineating how T. gondii can orchestrate a complex biological phenotype, such as the migratory activation of phagocytes to boost dissemination.

DOI: https://doi.org/10.1101/2024.02.06.579146
Pharmaceutics. 2024 Feb 10. pii: 262. [Epub ahead of print]16(2):

First Description of Marinoquinoline Derivatives' Activity against Toxoplasma gondii.

Luiza Tamie Hirata Diethelm, Amanda Bruno da Silva Bellini Ramos, Giovanna Braga de Lorena, Bruna Inácio Trajano, Rafael Dias do Espírito Santo, Renata Priscila Barros de Menezes, Marcus Tullius Scotti, Fabio Antonio Colombo, Marcos José Marques, Carlos Roque Duarte Correia, Juliana Quero Reimão.

  Toxoplasmosis is a globally prevalent zoonotic disease with significant clinical implications, including neurotoxoplasmosis, a leading cause of cerebral lesions in AIDS patients. The current pharmacological treatments for toxoplasmosis face clinical limitations, necessitating the urgent development of new therapeutics. Natural sources have yielded diverse bioactive compounds, serving as the foundation for clinically used derivatives. The exploration of marine bacteria-derived natural products has led to marinoquinolines, which feature a pyrroloquinoline core and demonstrate in vitro and in vivo anti-Plasmodium activity. This study investigates the in vitro anti-Toxoplasma gondii potential of six marinoquinoline derivatives. Additionally, it conducts absorption, distribution, metabolism, excretion, and toxicity (ADMET) predictions, and evaluates the in vivo efficacy of one selected compound. The compounds displayed half-maximal effective concentration (EC50) values between 1.31 and 3.78 µM and half-maximal cytotoxic concentration (CC50) values ranging from 4.16 to 30.51 µM, resulting in selectivity indices (SI) from 3.18 to 20.85. MQ-1 exhibiting the highest in vitro SI, significantly reduced tachyzoite numbers in the peritoneum of RH-infected Swiss mice when it was orally administered at 12.5 mg/kg/day for eight consecutive days. Also, MQ-1 significantly reduced the cerebral parasite burden in chronically ME49 infected C57BL/6 mice when it was orally administered at 25 mg/kg/day for 10 consecutive days. These findings underscore the promising anti-T. gondii activity of marinoquinolines and their potential as novel therapeutic agents against this disease.

Keywords:  anti-T. gondii activity; in vitro evaluation; in vivo efficacy; marinoquinolines; neurotoxoplasmosis; therapeutics; toxoplasmosis

DOI:  https://doi.org/10.3390/pharmaceutics16020262
Cell Rep. 2024 Feb 15. pii: S2211-1247(24)00123-2. [Epub ahead of print]43(2): 113795

The IRAK1/IRF5 axis initiates IL-12 response by dendritic cells and control of Toxoplasma gondii infection.

Milton Pereira, Theresa Ramalho, Warrison A Andrade, Danielle F Durso, Maria C Souza, Katherine A Fitzgerald, Douglas T Golenbock, Neal Silverman, Ricardo T Gazzinelli.

  Activation of endosomal Toll-like receptor (TLR) 7, TLR9, and TLR11/12 is a key event in the resistance against the parasite Toxoplasma gondii. Endosomal TLR engagement leads to expression of interleukin (IL)-12 via the myddosome, a protein complex containing MyD88 and IL-1 receptor-associated kinase (IRAK) 4 in addition to IRAK1 or IRAK2. In murine macrophages, IRAK2 is essential for IL-12 production via endosomal TLRs but, surprisingly, Irak2-/- mice are only slightly susceptible to T. gondii infection, similar to Irak1-/- mice. Here, we report that upon T. gondii infection IL-12 production by different cell populations requires either IRAK1 or IRAK2, with conventional dendritic cells (DCs) requiring IRAK1 and monocyte-derived DCs (MO-DCs) requiring IRAK2. In both populations, we identify interferon regulatory factor 5 as the main transcription factor driving the myddosome-dependent IL-12 production during T. gondii infection. Consistent with a redundant role of DCs and MO-DCs, mutations that affect IL-12 production in both cell populations show high susceptibility to infection in vivo.

Keywords:  CP: Immunology; CP: Microbiology; IL-12; IRAK; IRF5; Toll-like receptors; Toxoplasma gondii; cell signaling; dendritic cells; innate immunity; monocytes; myddosome

DOI:  https://doi.org/10.1016/j.celrep.2024.113795
PLoS Pathog. 2024 Feb 20. 20(2): e1011502

Parasite-induced IFN-γ regulates host defense via CD115 and mTOR-dependent mechanism of tissue-resident macrophage death.

Andrew T Martin, Shilpi Giri, Alexandra Safronova, Sophia I Eliseeva, Samantha F Kwok, Felix Yarovinsky.

Host resistance to a common protozoan parasite Toxoplasma gondii relies on a coordinated immune response involving multiple cell types, including macrophages. Embryonically seeded tissue-resident macrophages (TRMs) play a critical role in maintaining tissue homeostasis, but their role in parasite clearance is poorly understood. In this study, we uncovered a crucial aspect of host defense against T. gondii mediated by TRMs. Through the use of neutralizing antibodies and conditional IFN-γ receptor-deficient mice, we demonstrated that IFN-γ directly mediated the elimination of TRMs. Mechanistically, IFN-γ stimulation in vivo rendered macrophages unresponsive to macrophage colony-stimulating factor (M-CSF) and inactivated mTOR signaling by causing the shedding of CD115 (CSFR1), the receptor for M-CSF. Further experiments revealed the essential role of macrophage IFN-γ responsiveness in host resistance to T. gondii. The elimination of peritoneal TRMs emerged as an additional host defense mechanism aimed at limiting the parasite's reservoir. The identified mechanism, involving IFN-γ-induced suppression of CD115-dependent mTOR signaling in macrophages, provides insights into the adaptation of macrophage subsets during infection and highlights a crucial aspect of host defense against intracellular pathogens.

DOI: https://doi.org/10.1371/journal.ppat.1011502
Biosens Bioelectron. 2024 Feb 13. pii: S0956-5663(24)00137-4. [Epub ahead of print]251 116132

Allosteric strand displacement isothermal amplification for the visual detection of Toxoplasma gondii in 30 minutes.

Lei Wang, Zhaoxuan Zhang, Yanger Liu, Shenghao Lin, Wenqiang Zhang, Longjiao Zhu, Wentao Xu, Yulan Dong.

  Early detection of Toxoplasma gondii (T. gondii) is critical due to a lack of effective treatment for toxoplasmosis.This study established a simple, cost-effective, and rapid colorimetric detection method for T. gondii. The entire testing process, from sample collection to results, takes only 0.5 h. These characteristics fulfill the demands of researchers seeking rapid target detection with minimal equipment reliance. For genomic extraction, this study evaluated the ability of two filter papers to capture genomes. A rapid genomic extraction device combined with the two filter papers was designed to simplify the genomic extraction process, which was completed within 10 min and increased the detection sensitivity tenfold. The method utilized a simplified primer design for isothermal amplification, namely allosteric strand displacement (ASD), and employed an underutilized commercial color indicator, Bromothymol Blue (BTB), for signal output. Compared with other reported indicators, BTB exhibited a more pronounced color change, shifting from blue to yellow in positive samples, facilitating easier visual differentiation. The reaction was completed in 20 min with a limit of detection (LOD) as low as 0.014 T. gondii per microliter.

Keywords:  Allosteric strand displacement; New color indicator; Rapid extraction and enrichment of genomes; Toxoplasma gondii

DOI:  https://doi.org/10.1016/j.bios.2024.116132
Parasit Vectors. 2024 Feb 22. 17(1): 81

Application of gold immunochromatographic assay strip combined with digital evaluation for early detection of Toxoplasma gondii infection in multiple species.

Jiyuan Fan, Hao Sun, Jiawen Fang, Yafan Gao, Haojie Ding, Bin Zheng, Qingming Kong, Xunhui Zhuo, Shaohong Lu.

   BACKGROUND: Timely diagnosis of Toxoplasma gondii infection is necessary to prevent and control toxoplasmosis transmission. The gold immunochromatographic assay (GICA) is a means of rapidly detecting pathogen in samples. GICA-based diagnostic methods have been developed to accurately detect pathogens with high sensitivity and specificity, and their application in T. gondii diagnosis is expected to yield good results.
METHODS: Colloidal gold test strips were produced using T. gondii C-terminal truncated apical membrane antigen 1 (AMA1C). Colloidal gold-AMA1C and colloidal gold-murine protein conjugate were synthesized under optimal conditions. A nitrocellulose membrane was treated with AMA1C and goat anti-mouse antibody as the test line and control line, respectively. In total, 90 cat serum samples were tested using AMA1C-GICA and a commercial enzyme linked immunosorbent assay (ELISA) kit. The GICA results were digitally displayed using a portable colloidal gold immunochromatographic test strip analyzer (HMREADER). The sensitivity, specificity, and stability of AMA1C-GICA were assessed, and this was then used to examine clinical samples, including 203 human sera, 266 cat sera, and 81 dog sera.
RESULTS: AMA1C-GICA had a detection threshold of 1:32 for T. gondii-positive serum. The GICA strips specifically detected T. gondii antibodies and exhibited no reactivity with Plasmodium vivax, Paragonimus kellicotti, Schistosoma japonicum, Clonorchis sinensis, and Schistosoma mansoni. Consequently, 15 (16.7%) positive samples were detected using the AMA1C-GICA and commercial ELISA kits for each of the assays. The receiver-operating characteristic curve showed that GICA had a relative sensitivity of 85.3% and specificity of 92%, with an area under the curve of 98%. After analyzing clinical samples using HMREADER, 1.2%-23.4% of these samples were found to be positive for T. gondii.
CONCLUSIONS: This study presents a novel assay that enables timely and efficient detection of serum antibodies against T. gondii, thereby allowing for its early clinical diagnosis. Furthermore, the integration of digital detection using HMREADER can enhance the implementation of GICA.

Keywords:  AMA1C; Digital application; Early detection; GICA; T. gondii

DOI:  https://doi.org/10.1186/s13071-024-06180-1
Pathogens. 2024 Feb 07. pii: 153. [Epub ahead of print]13(2):

Prevalence of Toxoplasma gondii in Wild American Mink (Neogale vison): The First Serological Study in Germany and Poland.

Mike Heddergott, Jutta Pikalo, Franz Müller, Natalia Osten-Sacken, Peter Steinbach.

  Toxoplasma gondii is an obligate intracellular protozoan that causes toxoplasmosis in warm-blooded animals. Although most infections in humans and animals are subclinical, an infection can nevertheless be fatal. One of the important characteristics in the epidemiology of this parasite is waterborne transmission. The American mink (Neogale vison), a mammal closely adapted to freshwater ecosystems, is a potential sentinel for T. gondii. We analysed meat juice from the heart of 194 wild minks collected between 2019 and 2022 in five study areas from Germany and Poland and tested for the presence of antibodies against T. gondii. The analysis was performed using a commercial enzyme-linked immunosorbent assay test (ELISA). Antibodies were detected in 45.36% (88/194, 95% confidence interval (CI): 38.39-52.41%) of the analysed animals. While the prevalence values ranged from 37.50% to 49.30%, there was no significant difference in seroprevalence between the study areas. Juveniles were less likely to carry T. gondii antibodies than adults (odds ratio: 0.216), whereas there was no significant difference in prevalence between the sexes (odds ratio: 0.933). The results of our study show that contact with T. gondii is widespread in minks, and the parasite is common in inland freshwater ecosystems in Germany and Poland. This indicates that watercourses play an important role in the spread of T. gondii oocysts.

Keywords:  American mink; Neogale vison; Toxoplasma gondii; enzyme-linked immunosorbent assay (ELISA); invasive species; wildlife; zoonosis

DOI:  https://doi.org/10.3390/pathogens13020153
EMBO Rep. 2024 Feb 23.

TORC1 is an essential regulator of nutrient-controlled proliferation and differentiation in Leishmania.

Elmarie Myburgh, Vincent Geoghegan, Eliza Vc Alves-Ferreira, Y Romina Nievas, Jaspreet S Grewal, Elaine Brown, Karen McLuskey, Jeremy C Mottram.

  Leishmania parasites undergo differentiation between various proliferating and non-dividing forms to adapt to changing host environments. The mechanisms that link environmental cues with the parasite's developmental changes remain elusive. Here, we report that Leishmania TORC1 is a key environmental sensor for parasite proliferation and differentiation in the sand fly-stage promastigotes and for replication of mammalian-stage amastigotes. We show that Leishmania RPTOR1, interacts with TOR1 and LST8, and identify new parasite-specific proteins that interact in this complex. We investigate TORC1 function by conditional deletion of RPTOR1, where under nutrient-rich conditions RPTOR1 depletion results in decreased protein synthesis and growth, G1 cell cycle arrest and premature differentiation from proliferative promastigotes to non-dividing mammalian-infective metacyclic forms. These parasites are unable to respond to nutrients to differentiate into proliferative retroleptomonads, which are required for their blood-meal induced amplification in sand flies and enhanced mammalian infectivity. We additionally show that RPTOR1-/- metacyclic promastigotes develop into amastigotes but do not proliferate in the mammalian host to cause pathology. RPTOR1-dependent TORC1 functionality represents a critical mechanism for driving parasite growth and proliferation.

Keywords:  Differentiation; Leishmania; Proliferation; RPTOR1; TORC1

DOI:  https://doi.org/10.1038/s44319-024-00084-y
Nat Commun. 2024 Feb 23. 15(1): 1674

Bacteria employ lysine acetylation of transcriptional regulators to adapt gene expression to cellular metabolism.

Magdalena Kremer, Sabrina Schulze, Nadja Eisenbruch, Felix Nagel, Robert Vogt, Leona Berndt, Babett Dörre, Gottfried J Palm, Jens Hoppen, Britta Girbardt, Dirk Albrecht, Susanne Sievers, Mihaela Delcea, Ulrich Baumann, Karin Schnetz, Michael Lammers.

The Escherichia coli TetR-related transcriptional regulator RutR is involved in the coordination of pyrimidine and purine metabolism. Here we report that lysine acetylation modulates RutR function. Applying the genetic code expansion concept, we produced site-specifically lysine-acetylated RutR proteins. The crystal structure of lysine-acetylated RutR reveals how acetylation switches off RutR-DNA-binding. We apply the genetic code expansion concept in E. coli in vivo revealing the consequences of RutR acetylation on the transcriptional level. We propose a model in which RutR acetylation follows different kinetic profiles either reacting non-enzymatically with acetyl-phosphate or enzymatically catalysed by the lysine acetyltransferases PatZ/YfiQ and YiaC. The NAD+-dependent sirtuin deacetylase CobB reverses enzymatic and non-enzymatic acetylation of RutR playing a dual regulatory and detoxifying role. By detecting cellular acetyl-CoA, NAD+ and acetyl-phosphate, bacteria apply lysine acetylation of transcriptional regulators to sense the cellular metabolic state directly adjusting gene expression to changing environmental conditions.

DOI: https://doi.org/10.1038/s41467-024-46039-8
bioRxiv. 2024 Feb 07. pii: 2024.02.06.579216. [Epub ahead of print]

mTORC1 activity oscillates throughout the cell cycle promoting mitotic entry and differentially influencing autophagy induction.

Jay N Joshi, Ariel D Lerner, Frank Scallo, Alexandra N Grumet, Paul Matteson, James H Millonig, Alexander J Valvezan.

Mechanistic Target of Rapamycin Complex 1 (mTORC1) is a master metabolic regulator that stimulates anabolic cell growth while suppressing catabolic processes such as autophagy. mTORC1 is active in most, if not all, proliferating eukaryotic cells. However, it remains unclear whether and how mTORC1 activity changes from one cell cycle phase to another. Here we tracked mTORC1 activity through the complete cell cycle and uncover oscillations in its activity. We find that mTORC1 activity peaks in S and G2, and is lowest in mitosis and G1. We further demonstrate that multiple mechanisms are involved in controlling this oscillation. The interphase oscillation is mediated through the TSC complex, an upstream negative regulator of mTORC1, but is independent of major known regulatory inputs to the TSC complex, including Akt, Mek/Erk, and CDK4/6 signaling. By contrast, suppression of mTORC1 activity in mitosis does not require the TSC complex, and instead involves CDK1-dependent control of the subcellular localization of mTORC1 itself. Functionally, we find that in addition to its well-established role in promoting progression through G1, mTORC1 also promotes progression through S and G2, and is important for satisfying the Wee1- and Chk1-dependent G2/M checkpoint to allow entry into mitosis. We also find that low mTORC1 activity in G1 sensitizes cells to autophagy induction in response to partial mTORC1 inhibition or reduced nutrient levels. Together these findings demonstrate that mTORC1 is differentially regulated throughout the cell cycle, with important phase-specific functional consequences in proliferating cells.

DOI: https://doi.org/10.1101/2024.02.06.579216
PLoS Negl Trop Dis. 2024 Feb 23. 18(2): e0012007

The bloodstream form of Trypanosoma brucei displays non-canonical gluconeogenesis.

Julie Kovářová, Martin Moos, Michael P Barrett, David Horn, Alena Zíková.

Trypanosoma brucei is a causative agent of the Human and Animal African Trypanosomiases. The mammalian stage parasites infect various tissues and organs including the bloodstream, central nervous system, skin, adipose tissue and lungs. They rely on ATP produced in glycolysis, consuming large amounts of glucose, which is readily available in the mammalian host. In addition to glucose, glycerol can also be used as a source of carbon and ATP and as a substrate for gluconeogenesis. However, the physiological relevance of glycerol-fed gluconeogenesis for the mammalian-infective life cycle forms remains elusive. To demonstrate its (in)dispensability, first we must identify the enzyme(s) of the pathway. Loss of the canonical gluconeogenic enzyme, fructose-1,6-bisphosphatase, does not abolish the process hence at least one other enzyme must participate in gluconeogenesis in trypanosomes. Using a combination of CRISPR/Cas9 gene editing and RNA interference, we generated mutants for four enzymes potentially capable of contributing to gluconeogenesis: fructose-1,6-bisphoshatase, sedoheptulose-1,7-bisphosphatase, phosphofructokinase and transaldolase, alone or in various combinations. Metabolomic analyses revealed that flux through gluconeogenesis was maintained irrespective of which of these genes were lost. Our data render unlikely a previously hypothesised role of a reverse phosphofructokinase reaction in gluconeogenesis and preclude the participation of a novel biochemical pathway involving transaldolase in the process. The sustained metabolic flux in gluconeogenesis in our mutants, including a triple-null strain, indicates the presence of a unique enzyme participating in gluconeogenesis. Additionally, the data provide new insights into gluconeogenesis and the pentose phosphate pathway, and improve the current understanding of carbon metabolism of the mammalian-infective stages of T. brucei.

DOI: https://doi.org/10.1371/journal.pntd.0012007
Trends Cell Biol. 2024 Feb 22. pii: S0962-8924(24)00023-0. [Epub ahead of print]

ER remodeling via lipid metabolism.

Wonyul Jang, Volker Haucke.

  Unlike most other organelles found in multiple copies, the endoplasmic reticulum (ER) is a unique singular organelle within eukaryotic cells. Despite its continuous membrane structure, encompassing more than half of the cellular endomembrane system, the ER is subdivided into specialized sub-compartments, including morphological, membrane contact site (MCS), and de novo organelle biogenesis domains. In this review, we discuss recent emerging evidence indicating that, in response to nutrient stress, cells undergo a reorganization of these sub-compartmental ER domains through two main mechanisms: non-destructive remodeling of morphological ER domains via regulation of MCS and organelle hitchhiking, and destructive remodeling of specialized domains by ER-phagy. We further highlight and propose a critical role of membrane lipid metabolism in this ER remodeling during starvation.

Keywords:  endoplasmic reticulum; hitchhiking; lipids; membrane contact sites; membrane remodeling; metabolism; nutrient stress

DOI:  https://doi.org/10.1016/j.tcb.2024.01.011
Trends Pharmacol Sci. 2024 Feb 20. pii: S0165-6147(24)00023-3. [Epub ahead of print]

Histone lysine acetyltransferase inhibitors: an emerging class of drugs for cancer therapy.

Jeffrey White, Frederick A Derheimer, Kristen Jensen-Pergakes, Shawn O'Connell, Shikhar Sharma, Noah Spiegel, Thomas A Paul.

  Lysine acetyltransferases (KATs) are a family of epigenetic enzymes involved in the regulation of gene expression; they represent a promising class of emerging drug targets. The frequent molecular dysregulation of these enzymes, as well as their mechanistic links to biological functions that are crucial to cancer, have led to exploration around the development of small-molecule inhibitors against KATs. Despite early challenges, recent advances have led to the development of potent and selective enzymatic and bromodomain (BRD) KAT inhibitors. In this review we discuss the discovery and development of new KAT inhibitors and their application as oncology therapeutics. Additionally, new chemically induced proximity approaches are presented, offering opportunities for unique target selectivity profiles and tissue-specific targeting of KATs. Emerging clinical data for CREB binding protein (CREBBP)/EP300 BRD inhibitors and KAT6 catalytic inhibitors indicate the promise of this target class in cancer therapeutics.

Keywords:  CREBBP; EP300; KAT2A; KAT6A; KAT7; lysine acetyltransferases

DOI:  https://doi.org/10.1016/j.tips.2024.01.010
Int J Mol Sci. 2024 Feb 06. pii: 1956. [Epub ahead of print]25(4):

Sirtuins as Players in the Signal Transduction of Citrus Flavonoids.

Giovanni Enrico Lombardo, Caterina Russo, Alessandro Maugeri, Michele Navarra.

  Sirtuins (SIRTs) belong to the family of nicotine adenine dinucleotide (NAD+)-dependent class III histone deacetylases, which come into play in the regulation of epigenetic processes through the deacetylation of histones and other substrates. The human genome encodes for seven homologs (SIRT1-7), which are localized into the nucleus, cytoplasm, and mitochondria, with different enzymatic activities and regulatory mechanisms. Indeed, SIRTs are involved in different physio-pathological processes responsible for the onset of several human illnesses, such as cardiovascular and neurodegenerative diseases, obesity and diabetes, age-related disorders, and cancer. Nowadays, it is well-known that Citrus fruits, typical of the Mediterranean diet, are an important source of bioactive compounds, such as polyphenols. Among these, flavonoids are recognized as potential agents endowed with a wide range of beneficial properties, including antioxidant, anti-inflammatory, hypolipidemic, and antitumoral ones. On these bases, we offer a comprehensive overview on biological effects exerted by Citrus flavonoids via targeting SIRTs, which acted as modulator of several signaling pathways. According to the reported studies, Citrus flavonoids appear to be promising SIRT modulators in many different pathologies, a role which might be potentially evaluated in future therapies, along with encouraging the study of those SIRT members which still lack proper evidence on their support.

Keywords:  Citrus fruits; SIRT1; SIRT2; SIRT3; SIRT4; SIRT5; SIRT6; SIRT7; flavonoids; polyphenols

DOI:  https://doi.org/10.3390/ijms25041956
J Biol Chem. 2024 Feb 21. pii: S0021-9258(24)00154-6. [Epub ahead of print] 105778

SUMO Modifies GβL and Mediates mTOR Signaling.

Sophia Louise Lucille Park, Uri Nimrod Ramírez-Jarquín, Neelam Shahani, Oscar Rivera, Manish Sharma, Preksha Sandipkumar Joshi, Aayushi Hansalia, Sunayana Dagar, Francis P McManus, Pierre Thibault, Srinivasa Subramaniam.

  The mechanistic target of rapamycin (mTOR) signaling is influenced by multiple regulatory proteins and post-translational modifications, however, underlying mechanisms remain unclear. Here, we report a novel role of small ubiquitin-like modifier (SUMO) in mTOR complex assembly and activity. By investigating the SUMOylation status of core mTOR components, we observed that the regulatory subunit, GβL, is modified by SUMO1, 2, and 3 isoforms. Using mutagenesis and mass spectrometry, we identified that GβL is SUMOylated at lysine sites K86, K215, K245, K261 and K305. We found that SUMO depletion reduces mTOR-Raptor and mTOR-Rictor complex formation and diminishes nutrient-induced mTOR signaling. Reconstitution with WT GβL but not SUMOylation defective KR mutant GβL promote mTOR signaling in GβL-depleted cells. Furthermore, we found that amino acids-induced mTORC1 signaling is diminished in the SUMO1-KO mice. Taken together, we report for the very first time that SUMO modifies GβL, influences the assembly of mTOR protein complexes, and regulates mTOR activity.

Keywords:  Amino acid stimulation; Kinase signaling; Lysine-site regulation; Nutrient signaling; Post-translational modification; Protein-protein interaction; SUMO isoforms; SUMO mechanism; Sumo interactive motif (SIM)

DOI:  https://doi.org/10.1016/j.jbc.2024.105778
FEBS Lett. 2024 Feb 18.

Identification and characterization of nuclear localization signals in the circumsporozoite protein of Plasmodium falciparum.

Akshaykumar Nanaji Shrikondawar, Kiranmai Chennoju, Debasish Kumar Ghosh, Akash Ranjan.

  Secretory proteins of Plasmodium exhibit differential spatial and functional activity within the host cell nucleus. However, the nuclear localization signals (NLSs) for these proteins remain largely uncharacterized. In this study, we have identified and characterized two NLSs in the circumsporozoite protein of Plasmodium falciparum (Pf-CSP). Both NLSs in the Pf-CSP contain clusters of lysine and arginine residues essential for specific interactions with the conserved tryptophan and asparagine residues of importin-α, facilitating nuclear translocation of Pf-CSP. While the two NLSs of Pf-CSP function independently and are both crucial for nuclear localization, a single NLS of Pf-CSP leads to weak nuclear localization. These findings shed light on the mechanism of nuclear penetrability of secretory proteins of Plasmodium proteins.

Keywords:  Plasmodium falciparum; circumsporozoite protein; nuclear localization signal; nuclear translocation

DOI:  https://doi.org/10.1002/1873-3468.14829
Biochim Biophys Acta Mol Cell Res. 2024 Feb 19. pii: S0167-4889(24)00040-5. [Epub ahead of print]1871(4): 119697

The enzymes of serine synthesis pathway in cancer metastasis.

Lei Li, Yuting Qin, Yuping Chen.

  Metastasis, the major cause of cancer mortality, requires cancer cells to reprogram their metabolism to adapt to and thrive in different environments, thereby leaving metastatic cells metabolic characteristics different from their parental cells. Mounting research has revealed that the de novo serine synthesis pathway (SSP), a glycolytic branching pathway that consumes glucose carbons for serine makeup and α-ketoglutarate generation and thus supports the proliferation, survival, and motility of cancer cells, is one such reprogrammed metabolic pathway. During different metastatic cascades, the SSP enzyme proteins or their enzymatic activity are both dynamically altered; manipulating their expression or catalytic activity could effectively prevent the progression of cancer metastasis; and the SSP enzymatic proteins could even conduce to metastasis via their nonenzymatic functions. In this article we overview the SSP dynamics during cancer metastasis and put the focuses on the regulatory role of the SSP in metastasis and the underlying mechanisms that mainly involve cellular anabolism/catabolism, redox balance, and epigenetics, aiming to provide a theoretical basis for the development of therapeutic strategies for targeting metastatic lesions.

Keywords:  Metabolic reprogramming; Metastasis; PHGDH; Serine biosynthesis; Serine metabolism

DOI:  https://doi.org/10.1016/j.bbamcr.2024.119697
Mol Metab. 2024 Feb 16. pii: S2212-8778(24)00034-6. [Epub ahead of print] 101903

Acetyl-CoA synthetase (ACSS2) does not generate butyryl- and crotonyl-CoA.

Nour Zeaiter, Laura Belot, Valérie Cunin, Roland Abi Nahed, Malgorzata Tokarska-Schlattner, Audrey Le Gouellec, Carlo Petosa, Saadi Khochbin, Uwe Schlattner.

  Acetyl and other acyl groups from different short-chain fatty acids (SCFA) competitively modify histones at various lysine sites. To fully understand the functional significance of such histone acylation, a key epigenetic mechanism, it is crucial to characterize the cellular sources of the corresponding acyl-CoA molecules required for the lysine modification. Like acetate, SCFAs such as propionate, butyrate and crotonate are thought to be the substrates used to generate the corresponding acyl-CoAs by enzymes known as acyl-CoA synthetases. The acetyl-CoA synthetase, ACSS2, which produces acetyl-CoA from acetate in the nucleocytoplasmic compartment, has been proposed to also mediate the synthesis of acyl-CoAs such as butyryl- and crotonyl-CoA from the corresponding SCFAs. This idea is now widely accepted and is sparking new research projects. However, based on our direct in vitro experiments with purified or recombinant enzymes and structural considerations, we demonstrate that ACSS2 is unable to mediate the generation of non-acetyl acyl-CoAs like butyryl- and crotonyl-CoA. It is therefore essential to re-examine published data and corresponding discussions in the light of this new finding.

Keywords:  Acyl-CoA; Epigenetics; Histone acylation; Protein acylation; Substrate specificity

DOI:  https://doi.org/10.1016/j.molmet.2024.101903
Mol Biol Rep. 2024 Feb 23. 51(1): 337

Exploring the potential of EphA2 receptor signaling pathway: a comprehensive review in cancer treatment.

Mohd Nehal, Jahanarah Khatoon, Salman Akhtar, Mohammad Kalim Ahmad Khan.

  The protein encoded by the ephrin type-A receptor 2 (EphA2) gene is a member of the ephrin receptor subfamily of the receptor tyrosine kinase family (RTKs). Eph receptors play a significant role in various biological processes, particularly cancer progression, development, and pathogenesis. They have been observed to regulate cancer cell growth, migration, invasion, tumor development, invasiveness, angiogenesis, and metastasis. To target EphA2 activity, various molecular, genetic, biochemical, and pharmacological strategies have been extensively tested in laboratory cultures and animal models. Notably, drugs, such as dasatinib, initially designed to target the kinase family, have demonstrated an additional capability to target EphA2 activity. Additionally, a novel monoclonal antibody named EA5 has emerged as a promising option to counteract the effects of EphA2 overexpression and restore tamoxifen sensitivity in EphA2-transfected MCF-7 cells during in vitro experiments. This antibody mimicked the binding of Ephrin A to EphA2. These methods offer potential avenues for inhibiting EphA2 activity, which could significantly decelerate breast cancer progression and restore sensitivity to certain drugs. This review article comprehensively covers EphA2's involvement in multiple malignancies, including ovarian, colorectal, breast, lung, glioma, and melanoma. Furthermore, we discuss the structure of EphA2, the Eph-Ephrin signaling pathway, various EphA2 inhibitors, and the mechanisms of EphA2 degradation. This article provides an extensive overview of EphA2's vital role in different types of cancers and outlines potential therapeutic approaches to target EphA2, shedding light on the underlying molecular mechanisms that make it an attractive target for cancer treatment.

Keywords:  Cancer; EphA2; Kinase inhibitors; RTKs; Targeted therapy

DOI:  https://doi.org/10.1007/s11033-024-09298-8
J Cell Sci. 2024 Feb 15. pii: jcs261655. [Epub ahead of print]137(4):

Going through changes - the role of autophagy during reprogramming and differentiation.

Morten Petersen, Elise Ebstrup, Eleazar Rodriguez.

  Somatic cell reprogramming is a complex feature that allows differentiated cells to undergo fate changes into different cell types. This process, which is conserved between plants and animals, is often achieved via dedifferentiation into pluripotent stem cells, which have the ability to generate all other types of cells and tissues of a given organism. Cellular reprogramming is thus a complex process that requires extensive modification at the epigenetic and transcriptional level, unlocking cellular programs that allow cells to acquire pluripotency. In addition to alterations in the gene expression profile, cellular reprogramming requires rearrangement of the proteome, organelles and metabolism, but these changes are comparatively less studied. In this context, autophagy, a cellular catabolic process that participates in the recycling of intracellular constituents, has the capacity to affect different aspects of cellular reprogramming, including the removal of protein signatures that might hamper reprogramming, mitophagy associated with metabolic reprogramming, and the supply of energy and metabolic building blocks to cells that undergo fate changes. In this Review, we discuss advances in our understanding of the role of autophagy during cellular reprogramming by drawing comparisons between plant and animal studies, as well as highlighting aspects of the topic that warrant further research.

Keywords:  Autophagy; Plant science; Regeneration; Reprogramming; Somatic reprogramming; Stem cells

DOI:  https://doi.org/10.1242/jcs.261655
Epigenetics. 2024 Dec;19(1): 2309824

Association of changes in expression of HDAC and SIRT genes after drug treatment with cancer cell line sensitivity to kinase inhibitors.

Julia Krushkal, Yingdong Zhao, Kyle Roney, Weimin Zhu, Alan Brooks, Deborah Wilsker, Ralph E Parchment, Lisa M McShane, James H Doroshow.

  Histone deacetylases (HDACs) and sirtuins (SIRTs) are important epigenetic regulators of cancer pathways. There is a limited understanding of how transcriptional regulation of their genes is affected by chemotherapeutic agents, and how such transcriptional changes affect tumour sensitivity to drug treatment. We investigated the concerted transcriptional response of HDAC and SIRT genes to 15 approved antitumor agents in the NCI-60 cancer cell line panel. Antitumor agents with diverse mechanisms of action induced upregulation or downregulation of multiple HDAC and SIRT genes. HDAC5 was upregulated by dasatinib and erlotinib in the majority of the cell lines. Tumour cell line sensitivity to kinase inhibitors was associated with upregulation of HDAC5, HDAC1, and several SIRT genes. We confirmed changes in HDAC and SIRT expression in independent datasets. We also experimentally validated the upregulation of HDAC5 mRNA and protein expression by dasatinib in the highly sensitive IGROV1 cell line. HDAC5 was not upregulated in the UACC-257 cell line resistant to dasatinib. The effects of cancer drug treatment on expression of HDAC and SIRT genes may influence chemosensitivity and may need to be considered during chemotherapy.

Keywords:  HDAC5; Histone deacetylase; chemosensitivity; dasatinib; sirtuin

DOI:  https://doi.org/10.1080/15592294.2024.2309824
bioRxiv. 2024 Feb 10. pii: 2024.02.09.579679. [Epub ahead of print]

Structural characterization of the ACDC domain from ApiAP2 proteins of the malaria parasite.

Marine Le Berre, Thibault Tubiana, Philippa Reuterswärd Waldner, Noureddine Lazar, Ines Li de la Sierra, Joana Mendonca Santos, Manuel Llinás, Sylvie Nessler.

The Apicomplexan AP2 (ApiAP2) proteins are the best characterized family of DNA-binding proteins in the malaria parasite. Apart from the AP2 DNA-binding domain, there is little sequence similarity between ApiAP2 proteins and no other functional domains have been extensively characterized. One protein domain, which is present in a subset of the ApiAP2 proteins, is the conserved AP2-coincident domain mostly at the C-terminus (ACDC domain). Here we solved for the first time the crystal structure of the ACDC domain from two distinct Plasmodium falciparum ApiAP2 proteins and one orthologue from P. vivax , revealing a non-canonical four-helix bundle. Despite little sequence conservation between the ACDC domains from the two proteins, the structures are remarkably similar and do not resemble that of any other known protein domains. Due to their unique protein architecture and lack of homologues in the human genome, we performed in silico docking calculations against a library of known antimalarial compounds and we identified a small molecule that can potentially bind to any Apicomplexan ACDC domain within a pocket highly conserved amongst ApiAP2 proteins. Inhibitors based on this compound would disrupt the function of the ACDC domain and thus of the ApiAP2 proteins containing it, providing a new therapeutic window for targeting the malaria parasite and other Apicomplexans.

DOI: https://doi.org/10.1101/2024.02.09.579679
bioRxiv. 2024 Feb 11. pii: 2024.02.06.578742. [Epub ahead of print]

αKG-mediated carnitine synthesis promotes homologous recombination via histone acetylation.

Apoorva Uboveja, Zhentai Huang, Raquel Buj, Amandine Amalric, Hui Wang, Naveen Kumar Tangudu, Aidan R Cole, Emily Megill, Daniel Kantner, Adam Chatoff, Hafsah Ahmad, Mariola M Marcinkiewicz, Julie A Disharoon, Sarah Graff, Erika S Dahl, Nadine Hempel, Wayne Stallaert, Simone Sidoli, Benjamin G Bitler, David T Long, Nathaniel W Snyder, Katherine M Aird.

Homologous recombination (HR) deficiency enhances sensitivity to DNA damaging agents commonly used to treat cancer. In HR-proficient cancers, metabolic mechanisms driving response or resistance to DNA damaging agents remain unclear. Here we identified that depletion of alpha-ketoglutarate (αKG) sensitizes HR-proficient cells to DNA damaging agents by metabolic regulation of histone acetylation. αKG is required for the activity of αKG-dependent dioxygenases (αKGDDs), and prior work has shown that changes in αKGDD affect demethylases. Using a targeted CRISPR knockout library consisting of 64 αKGDDs, we discovered that Trimethyllysine Hydroxylase Epsilon (TMLHE), the first and rate-limiting enzyme in de novo carnitine synthesis, is necessary for proliferation of HR-proficient cells in the presence of DNA damaging agents. Unexpectedly, αKG-mediated TMLHE-dependent carnitine synthesis was required for histone acetylation, while histone methylation was affected but dispensable. The increase in histone acetylation via αKG-dependent carnitine synthesis promoted HR-mediated DNA repair through site- and substrate-specific histone acetylation. These data demonstrate for the first time that HR-proficiency is mediated through αKG directly influencing histone acetylation via carnitine synthesis and provide a metabolic avenue to induce HR-deficiency and sensitivity to DNA damaging agents.

DOI: https://doi.org/10.1101/2024.02.06.578742
Microorganisms. 2024 Feb 04. pii: 331. [Epub ahead of print]12(2):

Gene Expression of Ethanol and Acetate Metabolic Pathways in the Acinetobacter baumannii EmaSR Regulon.

Yu-Weng Huang, Hung-Yu Shu, Guang-Huey Lin.

   BACKGROUND: Previous studies have confirmed the involvement of EmaSR (ethanol metabolism a sensor/regulator) in the regulation of Acinetobacter baumannii ATCC 19606 ethanol and acetate metabolism. RNA-seq analysis further revealed that DJ41_568-571, DJ41_2796, DJ41_3218, and DJ41_3568 regulatory gene clusters potentially participate in ethanol and acetate metabolism under the control of EmaSR.
METHODS: This study fused the EmaSR regulon promoter segments with reporter genes and used fluorescence expression levels to determine whether EmaSR influences regulon expression in ethanol or acetate salt environments. The enzymatic function and kinetics of significantly regulated regulons were also studied.
RESULTS: The EmaSR regulons P2796 and P3218 exhibited > 2-fold increase in fluorescence expression in wild type compared to mutant strains in both ethanol and acetate environments, and PemaR demonstrated a comparable trend. Moreover, increases in DJ41_2796 concentration enhanced the conversion of acetate and succinyl-CoA into acetyl-CoA and succinate, suggesting that DJ41_2796 possesses acetate: succinyl-CoA transferase (ASCT) activity. The kcat/KM values for DJ41_2796 with potassium acetate, sodium acetate, and succinyl-CoA were 0.2131, 0.4547, and 20.4623 mM-1s-1, respectively.
CONCLUSIONS: In A. baumannii, EmaSR controls genes involved in ethanol and acetate metabolism, and the EmaSR regulon DJ41_2796 was found to possess ASCT activity.

Keywords:  Acinetobacter baumannii; acetate metabolism; acetate: succinyl-CoA transferase (ASCT); ethanol metabolism; two-component system

DOI:  https://doi.org/10.3390/microorganisms12020331
Biomedicines. 2024 Feb 07. pii: 386. [Epub ahead of print]12(2):

Sirtuins Affect Cancer Stem Cells via Epigenetic Regulation of Autophagy.

Ferenc Sipos, Györgyi Műzes.

  Sirtuins (SIRTs) are stress-responsive proteins that regulate several post-translational modifications, partly by acetylation, deacetylation, and affecting DNA methylation. As a result, they significantly regulate several cellular processes. In essence, they prolong lifespan and control the occurrence of spontaneous tumor growth. Members of the SIRT family have the ability to govern embryonic, hematopoietic, and other adult stem cells in certain tissues and cell types in distinct ways. Likewise, they can have both pro-tumor and anti-tumor effects on cancer stem cells, contingent upon the specific tissue from which they originate. The impact of autophagy on cancer stem cells, which varies depending on the specific circumstances, is a very intricate phenomenon that has significant significance for clinical and therapeutic purposes. SIRTs exert an impact on the autophagy process, whereas autophagy reciprocally affects the activity of certain SIRTs. The mechanism behind this connection in cancer stem cells remains poorly understood. This review presents the latest findings that position SIRTs at the point where cancer cells and autophagy interact. Our objective is to highlight the various roles of distinct SIRTs in cancer stem cell-related functions through autophagy. This would demonstrate their significance in the genesis and recurrence of cancer and offer a more precise understanding of their treatment possibilities in relation to autophagy.

Keywords:  DNA methylation; SIRT; acetylation; autophagy; cancer stem cells; deacetylation; epigenetics; sirtuins

DOI:  https://doi.org/10.3390/biomedicines12020386
Curr Issues Mol Biol. 2024 Jan 23. 46(2): 1020-1046

Impact of Lysine Succinylation on the Biology of Fungi.

John Adejor, Elisabeth Tumukunde, Guoqi Li, Hong Lin, Rui Xie, Shihua Wang.

  Post-translational modifications (PTMs) play a crucial role in protein functionality and the control of various cellular processes and secondary metabolites (SMs) in fungi. Lysine succinylation (Ksuc) is an emerging protein PTM characterized by the addition of a succinyl group to a lysine residue, which induces substantial alteration in the chemical and structural properties of the affected protein. This chemical alteration is reversible, dynamic in nature, and evolutionarily conserved. Recent investigations of numerous proteins that undergo significant succinylation have underscored the potential significance of Ksuc in various biological processes, encompassing normal physiological functions and the development of certain pathological processes and metabolites. This review aims to elucidate the molecular mechanisms underlying Ksuc and its diverse functions in fungi. Both conventional investigation techniques and predictive tools for identifying Ksuc sites were also considered. A more profound comprehension of Ksuc and its impact on the biology of fungi have the potential to unveil new insights into post-translational modification and may pave the way for innovative approaches that can be applied across various clinical contexts in the management of mycotoxins.

Keywords:  SIRT5; aflatoxin; fungi; post-translational modification; protein lysine acylation; succinyl-CoA; succinyltransferase

DOI:  https://doi.org/10.3390/cimb46020065
Front Nutr. 2024 ;11 1336057

Protein succinylation: regulating metabolism and beyond.

Xiaoli Hou, Yiqiu Chen, Xiao Li, Xianliang Gu, Weixia Dong, Jie Shi, Shaoping Ji.

  Modifications of protein post-translation are critical modulatory processes, which alters target protein biological activity，function and/or location, even involved in pathogenesis of some diseases. So far, there are at least 16 types of post-translation modifications identified, particularly through recent mass spectrometry analysis. Among them, succinylation (Ksuc) on protein lysine residues causes a variety of biological changes. Succinylation of proteins contributes to many cellular processes such as proliferation, growth, differentiation, metabolism and even tumorigenesis. Mechanically, Succinylation leads to conformation alteration of chromatin or remodeling. As a result, transcription/expression of target genes is changed accordingly. Recent research indicated that succinylation mainly contributes to metabolism modulations, from gene expression of metabolic enzymes to their activity modulation. In this review, we will conclude roles of succinylation in metabolic regulation of glucose, fat, amino acids and related metabolic disease launched by aberrant succinylation. Our goal is to stimulate extra attention to these still not well researched perhaps important succinylation modification on proteins and cell processes.

Keywords:  metabolism; post-translational modification; redox; regulation; succinylation

DOI:  https://doi.org/10.3389/fnut.2024.1336057