bims-toxgon 2022-12-25 papers

bims-toxgon

Biomed News

on Toxoplasma gondii metabolism

Issue of 2022–12–25
twenty papers selected by
Lakesh Kumar, BITS Pilani

Finding optimal drug target sites in parasite pathogens.
Host autophagy limits Toxoplasma gondii proliferation in the absence of IFN-γ by affecting the hijack of Rab11A-positive vesicles.
Regulation of Mitochondrial Energy Metabolism by Glutaredoxin 5 in the Apicomplexan Parasite Neospora caninum.
Recombinant Mouse Prolactin Confers Partial Protection Against Toxoplasma gondii Infection in a Pre-treated Experimental Murine Model.
A High-Throughput Amenable Dual Luciferase System for Measuring Toxoplasma gondii Bradyzoite Viability after Drug Treatment.
New insights into apicoplast metabolism in blood-stage malaria parasites.
Exploiting the Macrophage Production of IL-12 in Improvement of Vaccine Development against Toxoplasma gondii and Neospora caninum Infections.
Blocking AMPK β1 myristoylation enhances AMPK activity and protects mice from high-fat diet-induced obesity and hepatic steatosis.
Immunogenic evaluation of multi-epitope peptide-loaded PCPP microparticles as a vaccine candidate against Toxoplasma Gondii.
The multiple facets of acetyl-CoA metabolism: Energetics, biosynthesis, regulation, acylation and inborn errors.
mTOR Signaling Network in Cell Biology and Human Disease.
Changes in AMPK activity induces cellular senescence in human dental follicle cells.
PfATP4 inhibitors in the Medicines for Malaria Venture Malaria Box and Pathogen Box block the schizont-to-ring transition by inhibiting egress rather than invasion.
AMPK Promotes Larval Metamorphosis of Mytilus coruscus.
Targeting malaria parasites with novel derivatives of azithromycin.
Bioenergetics of pollen tube growth in Arabidopsis thaliana revealed by ratiometric genetically encoded biosensors.
mTORC1 regulates a lysosome-dependent adaptive shift in intracellular lipid species.
Corrections: Principles of Resonance Energy Transfer.
Role of the AMPK/SIRT1 pathway in non‑alcoholic fatty liver disease (Review).
Cryo-electron microscopy structures of human thyroid peroxidase (TPO) in complex with TPO antibodies.

Trends Parasitol. 2022 Dec 21. pii: S1471-4922(22)00308-7. [Epub ahead of print]

Finding optimal drug target sites in parasite pathogens.

Matthew Bogyo.

  Benns et al. have recently combined a chemoproteomic profiling method with a CRISPR-based gene-editing method to identify chemically targetable residues essential for fitness in the parasite Toxoplasma gondii. The result is a strategy that enables rapid discovery of new drug targets to combat T. gondii and other related parasites.

Keywords:  CRISPR-mediated gene editing; Plasmodium falciparum; Toxoplasma gondii; activity-based protein profiling; chemoproteomics; cysteine reactivity

DOI:  https://doi.org/10.1016/j.pt.2022.12.003
Front Microbiol. 2022 ;13 1052779

Host autophagy limits Toxoplasma gondii proliferation in the absence of IFN-γ by affecting the hijack of Rab11A-positive vesicles.

Lingtao Pan, Yimin Yang, Xueqiu Chen, Mingxiu Zhao, Chaoqun Yao, Kaiyin Sheng, Yi Yang, Guangxu Ma, Aifang Du.

   Introduction: Autophagy has been recognized as a bona fide immunological process. Evidence has shown that this process in IFN-γ stimulated cells controls Toxoplasma gondii proliferation or eliminates its infection. However, little is known about the effect of T. gondii infection on the host cell autophagy in the absence of IFN-γ.
Methods: Multiple autophagy detection methods and CRISPR/CAS9 technology were used to study T. gondii-induced autophagy in HeLa and several other mammalian cell lines.
Results: Here, we report increased LC3 II, autophagosome-like membrane structures, enhanced autophagic flux, and decreased lysosomes in a range of mammalian cell lines without IFN-γ treatment after T. gondii infection. Specifically, disruption of host atg5 (a necessary gene for autophagy) in HeLa cells promoted the intracellular replication of T. gondii, with the transcript level of rab11a increased, compared with that in wild-type cells. Further, after T. gondii infection, the abundance of Rab11A remained stable in wild-type HeLa cells but decreased in atg5 -/- mutant. Disruption of rab11a in the HeLa cells compromised the proliferation of T. gondii, and increased the transcription of gra2 in the parasite. Compared to the T. gondii wild-type RH∆ku80 strain, the ∆gra2 mutant induces enhanced host autophagy in HeLa cells, and results in slower replication of the parasite.
Discussion: Collectively, these results indicate that host cell autophagy can limit T. gondii proliferation in an IFN-γ independent manner, possibly by affecting the hijack of host Rab11A-positive vesicles by the parasite which involved TgGRA2. The findings provide novel insights into T. gondii infection in host cells and toxoplasmosis research.

Keywords:  Rab11A; TgGRA2; Toxoplasma gondii; autophagy; proliferation

DOI:  https://doi.org/10.3389/fmicb.2022.1052779
Microbiol Spectr. 2022 Dec 21. e0309122

Regulation of Mitochondrial Energy Metabolism by Glutaredoxin 5 in the Apicomplexan Parasite Neospora caninum.

Xingju Song, Xu Yang, Zhu Ying, Kaijian Wu, Jing Liu, Qun Liu.

  Iron-sulfur [Fe-S] clusters are one of the most ancient and functionally versatile natural biosynthetic prosthetic groups required by various proteins involved in important metabolic processes, including the oxidative phosphorylation of proteins, electron transfer, energy metabolism, DNA/RNA metabolism, and protein translation. Apicomplexan parasites harbor two possible [Fe-S] cluster assembly pathways: the iron-sulfur cluster (ISC) pathway in the mitochondria and the sulfur formation (SUF) pathway in the apicoplast. Glutaredoxin 5 (GRX5) is involved in the ISC pathway in many eukaryotes. However, the cellular roles of GRX5 in apicomplexan parasites remain to be explored. Here, we showed that Neospora caninum mitochondrial GRX5 (NcGRX5) deficiency resulted in aberrant mitochondrial ultrastructure and led to a significant reduction in parasite proliferation and virulence in mice, suggesting that NcGRX5 is important for parasite growth in vitro and in vivo. Comparative proteomics and energy metabolomics were used to investigate the effects of NcGRX5 on parasite growth and mitochondrial metabolism. The data showed that disruption of NcGRX5 downregulated the expression of mitochondrial electron transport chain (ETC) and tricarboxylic acid cycle (TCA) cycle proteins and reduced the corresponding metabolic fluxes. Subsequently, we identified 23 proteins that might be adjacent to or interact with NcGRX5 by proximity-based protein labeling techniques and proteomics. The interactions between NcGRX5 and two iron-sulfur cluster synthesis proteins (ISCS and ISCU1) were further confirmed by coimmunoprecipitation assays. In conclusion, NcGRX5 is important for parasite growth and may regulate mitochondrial energy metabolism by mediating the biosynthesis of iron-sulfur clusters. IMPORTANCE Iron-sulfur [Fe-S] clusters are among the oldest and most ubiquitous prosthetic groups, and they are required for a variety of proteins involved in important metabolic processes. The intracellular parasites in the phylum Apicomplexa, including Plasmodium, Toxoplasma gondii, and Neospora caninum, harbor the ISC pathway involved in the biosynthesis of [Fe-S] clusters in mitochondria. These cofactors are required for a variety of important biological processes. However, little is known about the role of oxidoreductase glutaredoxins in these parasites. Our data indicate that NcGRX5 is an essential protein that plays multiple roles in several biological processes of N. caninum. NcGRX5 interacts with the mitochondrial iron-sulfur cluster synthesis proteins ISCS and ISCU1 and also regulates parasite energy metabolism. These data provide an insider's view of the metabolic regulation and iron-sulfur cluster assembly processes in the apicomplexan parasites.

Keywords:  Neospora caninum; energy metabolism; glutaredoxin 5; iron-sulfur cluster assembly

DOI:  https://doi.org/10.1128/spectrum.03091-22
Acta Parasitol. 2022 Dec 21.

Recombinant Mouse Prolactin Confers Partial Protection Against Toxoplasma gondii Infection in a Pre-treated Experimental Murine Model.

Amany F Atia, Engy V N Beshay, Samar K Fath-Allah, Dina Sweed, Samar A El-Refai.

   INTRODUCTION: Toxoplasmosis is one of the most important health-threatening diseases with worldwide distribution and global impact. It is caused by Toxoplasma gondii (T. gondii), an intracellular apicomplexan parasite that can evade the host immune responses and establish a chronic infection. The available treatments are not efficient against this stage and have many adverse effects. There are no available effective vaccines, apart from Toxovax®, which is used in sheep to prevent abortion. Studies documented that prolactin (PRL) had in vivo and in vitro anti-Toxoplasma effects. Detailed research was recommended about the mechanisms of such inhibitory effects.
AIM: This study was designed to assess the possible protective role of the recombinant prolactin (rPRL) against T. gondii.
MATERIALS AND METHODS: Sixty experimentally infected mice with T. gondii were used. The treated mice received rPRL for five days before infection. Serum prolactin levels were measured; survival rate was monitored; number, size, and DNA of T. gondii cysts in the brain were measured; and histopathological and immunological studies were done.
RESULTS: There was a significant increase in the survival rate of the rPRL-treated mice, a significant decrease in the number, size, and DNA amount of T. gondii cysts in the brain with a noticeable improvement of histopathological lesions in the brain and liver tissues when compared to the infected untreated group. These effects seem to be achieved through stimulating humoral and cell-mediated immune responses that were evident by the significant rise in serum levels of anti-Toxoplasma IgM, IFN-γ, and TNF-α.
CONCLUSION: The rPRL elicited robust immune responses, which provided efficient protection against murine T. gondii infection.

Keywords:  Recombinant mouse prolactin; Toxoplasma gondii

DOI:  https://doi.org/10.1007/s11686-022-00651-0
Anal Chem. 2022 Dec 22.

A High-Throughput Amenable Dual Luciferase System for Measuring Toxoplasma gondii Bradyzoite Viability after Drug Treatment.

David Smith, Matteo Lunghi, Einar B Olafsson, Olivia Hatton, Manlio Di Cristina, Vern B Carruthers.

It is estimated that more than 2 billion people are chronically infected with the intracellular protozoan parasite Toxoplasma gondii (T. gondii). Despite this, there is currently no vaccine to prevent infection in humans, and there is no recognized curative treatment to clear tissue cysts. A major hurdle for identifying effective drug candidates against chronic-stage cysts has been the low throughput of existing in vitro assays for testing the survival of bradyzoites. We have developed a luciferase-based platform for specifically determining bradyzoite survival within in vitro cysts in a 96-well plate format. We engineered a cystogenic type II T. gondii PruΔku80Δhxgpr strain for stage-specific expression of firefly luciferase in the cytosol of bradyzoites and nanoluciferase for secretion into the lumen of the cyst (DuaLuc strain). Using this DuaLuc strain, we found that the ratio of firefly luciferase to nanoluciferase decreased upon treatment with atovaquone or LHVS, two compounds that are known to compromise bradyzoite viability. The 96-well format allowed us to test several additional compounds and generate dose-response curves for calculation of EC50 values indicating relative effectiveness of a compound. Accordingly, this DuaLuc system should be suitable for screening libraries of diverse compounds and defining the potency of hits or other compounds with a putative antibradyzoite activity.

DOI: https://doi.org/10.1021/acs.analchem.2c02174
Curr Opin Microbiol. 2022 Dec 21. pii: S1369-5274(22)00139-4. [Epub ahead of print]71 102255

New insights into apicoplast metabolism in blood-stage malaria parasites.

Rubayet Elahi, Sean T Prigge.

The apicoplast of Plasmodium falciparum is the only source of essential isoprenoid precursors and Coenzyme A (CoA) in the parasite. Isoprenoid precursor synthesis relies on the iron-sulfur cluster (FeS) cofactors produced within the apicoplast, rendering FeS synthesis an essential function of this organelle. Recent reports provide important insights into the roles of FeS cofactors and the use of isoprenoid precursors and CoA both inside and outside the apicoplast. Here, we review the recent insights into the roles of these metabolites in blood-stage malaria parasites and discuss new questions that have been raised in light of these discoveries.

DOI: https://doi.org/10.1016/j.mib.2022.102255
Vaccines (Basel). 2022 Dec 06. pii: 2082. [Epub ahead of print]10(12):

Exploiting the Macrophage Production of IL-12 in Improvement of Vaccine Development against Toxoplasma gondii and Neospora caninum Infections.

Ragab M Fereig, Mosaab A Omar, Abdullah F Alsayeqh.

  Toxoplasmosis and neosporosis are major protozoan diseases of global distribution. Toxoplasma gondii is the cause of toxoplasmosis, which affects almost all warm-blooded animals, including humans, while Neospora caninum induces neosporosis in many animal species, especially cattle. The current defective situation with control measures is hindering all efforts to overcome the health hazards and economic losses of toxoplasmosis and neosporosis. Adequate understanding of host-parasite interactions and host strategies to combat such infections can be exploited in establishing potent control measures, including vaccine development. Macrophages are the first defense line of innate immunity, which is responsible for the successful elimination of T.gondii or N. caninum. This action is exerted via the immunoregulatory interleukin-12 (IL-12), which orchestrates the production of interferon gamma (IFN-γ) from various immune cells. Cellular immune response and IFN-γ production is the hallmark for successful vaccine candidates against both T. gondii and N. caninum. However, the discovery of potential vaccine candidates is a highly laborious, time-consuming and expensive procedure. In this review, we will try to exploit previous knowledge and our research experience to establish an efficient immunological approach for exploring potential vaccine candidates against T. gondii and N. caninum. Our previous studies on vaccine development against both T. gondii and N. caninum revealed a strong association between the successful and potential vaccine antigens and their ability to promote the macrophage secretion of IL-12 using a murine model. This phenomenon was emphasized using different recombinant antigens, parasites, and experimental approaches. Upon these data and research trials, IL-12 production from murine macrophages can be used as an initial predictor for judgment of vaccine efficacy before further evaluation in time-consuming and laborious in vivo experiments. However, more studies and research are required to conceptualize this immunological approach.

Keywords:  N. caninum; T. gondii; antigen; macrophages; neosporosis; vaccine

DOI:  https://doi.org/10.3390/vaccines10122082
Cell Rep. 2022 Dec 20. pii: S2211-1247(22)01758-2. [Epub ahead of print]41(12): 111862

Blocking AMPK β1 myristoylation enhances AMPK activity and protects mice from high-fat diet-induced obesity and hepatic steatosis.

Katyayanee Neopane, Natalie Kozlov, Florentina Negoita, Lisa Murray-Segal, Robert Brink, Ashfaqul Hoque, Ashley J Ovens, Gavin Tjin, Luke M McAloon, Dingyi Yu, Naomi X Y Ling, Matthew J Sanders, Jonathan S Oakhill, John W Scott, Gregory R Steinberg, Kim Loh, Bruce E Kemp, Kei Sakamoto, Sandra Galic.

  AMP-activated protein kinase (AMPK) is a master regulator of cellular energy homeostasis and a therapeutic target for metabolic diseases. Co/post-translational N-myristoylation of glycine-2 (Gly2) of the AMPK β subunit has been suggested to regulate the distribution of the kinase between the cytosol and membranes through a "myristoyl switch" mechanism. However, the relevance of AMPK myristoylation for metabolic signaling in cells and in vivo is unclear. Here, we generated knockin mice with a Gly2-to-alanine point mutation of AMPKβ1 (β1-G2A). We demonstrate that non-myristoylated AMPKβ1 has reduced stability but is associated with increased kinase activity and phosphorylation of the Thr172 activation site in the AMPK α subunit. Using proximity ligation assays, we show that loss of β1 myristoylation impedes colocalization of the phosphatase PPM1A/B with AMPK in cells. Mice carrying the β1-G2A mutation have improved metabolic health with reduced adiposity, hepatic lipid accumulation, and insulin resistance under conditions of high-fat diet-induced obesity.

Keywords:  AMPK; CP: Metabolism; adiposity; myristoylation; phosphatase; signal transduction; steatosis

DOI:  https://doi.org/10.1016/j.celrep.2022.111862
Comp Immunol Microbiol Infect Dis. 2022 Dec 05. pii: S0147-9571(22)00184-9. [Epub ahead of print]92 101927

Immunogenic evaluation of multi-epitope peptide-loaded PCPP microparticles as a vaccine candidate against Toxoplasma Gondii.

Rabia Yilmaz-Ozturk, Hilal Calik, Serkan Yaman, Eslin Ustun-Karatop, Rabia Cakir-Koc.

  Toxoplasmosis is a major health problem and socioeconomic burden, affecting around 30-50% of the global population. Poly(dicarboxylatophenoxy)phosphazene (PCPP) polymer was chosen as adjuvant for the immunogenic peptide antigen. Peptide-loaded PCPP microparticles were synthesized via the coacervation method and the characterization studies of microparticles were conducted to determine their size, charge, morphology, encapsulation efficacy, and loading capacity. To evaluate in vivo efficacy of the vaccine candidate, Balb/c mice were immunized with the formulations. Brain and spleen tissues were isolated from animals to investigate cytokine levels, lymphocyte proliferation, and brain cyst formation. As a result, antibody and cytokine responses in groups immunized with peptide-loaded PCPP microparticles were found to be significantly higher when compared to the control group. In conclusion, our novel multi-epitope peptide-loaded PCPP microparticle-based vaccine formulation demonstrated considerable humoral and cellular immune responses against T. gondii and protected mice against T. gondii infection during Toxoplasmosis.

Keywords:  Immunity; Microparticle; PCPP; Peptide-based vaccine; Toxoplasma gondii

DOI:  https://doi.org/10.1016/j.cimid.2022.101927
Mol Genet Metab. 2022 Nov 30. pii: S1096-7192(22)00442-5. [Epub ahead of print]138(1): 106966

The multiple facets of acetyl-CoA metabolism: Energetics, biosynthesis, regulation, acylation and inborn errors.

Youlin Wang, Hao Yang, Chloé Geerts, Alexandra Furtos, Paula Waters, Denis Cyr, Shupei Wang, Grant A Mitchell.

  Acetyl-coenzyme A (Ac-CoA) is a core metabolite with essential roles throughout cell physiology. These functions can be classified into energetics, biosynthesis, regulation and acetylation of large and small molecules. Ac-CoA is essential for oxidative metabolism of glucose, fatty acids, most amino acids, ethanol, and of free acetate generated by endogenous metabolism or by gut bacteria. Ac-CoA cannot cross lipid bilayers, but acetyl groups from Ac-CoA can shuttle across membranes as part of carrier molecules like citrate or acetylcarnitine, or as free acetate or ketone bodies. Ac-CoA is the basic unit of lipid biosynthesis, providing essentially all of the carbon for the synthesis of fatty acids and of isoprenoid-derived compounds including cholesterol, coenzyme Q and dolichols. High levels of Ac-CoA in hepatocytes stimulate lipid biosynthesis, ketone body production and the diversion of pyruvate metabolism towards gluconeogenesis and away from oxidation; low levels exert opposite effects. Acetylation changes the properties of molecules. Acetylation is necessary for the synthesis of acetylcholine, acetylglutamate, acetylaspartate and N-acetyl amino sugars, and to metabolize/eliminate some xenobiotics. Acetylation is a major post-translational modification of proteins. Different types of protein acetylation occur. The most-studied form occurs at the epsilon nitrogen of lysine residues. In histones, lysine acetylation can alter gene transcription. Acetylation of other proteins has diverse, often incompletely-documented effects. Inborn errors related to Ac-CoA feature a broad spectrum of metabolic, neurological and other features. To date, a small number of studies of animals with inborn errors of CoA thioesters has included direct measurement of acyl-CoAs. These studies have shown that low levels of tissue Ac-CoA correlate with the development of clinical signs, hinting that shortage of Ac-CoA may be a recurrent theme in these conditions. Low levels of Ac-CoA could potentially disrupt any of its roles.

Keywords:  Acetyl-CoA; Acetylation; Acylation; Energy metabolism; Inborn errors

DOI:  https://doi.org/10.1016/j.ymgme.2022.106966
Int J Mol Sci. 2022 Dec 18. pii: 16142. [Epub ahead of print]23(24):

mTOR Signaling Network in Cell Biology and Human Disease.

Jane J Yu, Elena A Goncharova.

The mechanistic target of rapamycin (mTOR) is a serine/threonine protein kinase that regulates multiple processes, including gene transcription, protein synthesis, ribosome biogenesis, autophagy, cell metabolism, and cell growth [...].

DOI: https://doi.org/10.3390/ijms232416142
Exp Gerontol. 2022 Dec 21. pii: S0531-5565(22)00380-1. [Epub ahead of print]172 112071

Changes in AMPK activity induces cellular senescence in human dental follicle cells.

Christian Morsczeck, Anja Reck, Torsten E Reichert.

  Dental Follicle Cells (DFCs) are somatic stem cells with a limited lifespan, but little is known about a possible mechanism of cellular senescence. Previous studies have shown that cellular senescence is associated with increased demand of glycolsis or the "glycolytic metabotype", which can be induced by activation of 5' adenosine monophosphate-activated protein kinase (AMPK), and decreased autophagy. This study examined the role of AMPK in inducing senescence in DFCs. During the induction of cellular senescence, AMPK activity was impaired, suggesting a negative impact on senescence induction. In line with this assumption, cellular senescence was induced upon inhibition of AMPK with a specific siRNA. In addition, after this inhibition, autophagy was also inhibited. Moreover, specific inhibition of autophagy promoted cellular senescence. However, inducers of AMPK such as metformin or AICAR surprisingly increased senescence in DFCs. Interestingly, autophagy was impaired after long-term induction of AMPK with AICAR and metformin. Moreover, activation of AMPK induces the consumption of glucose but decreases NAD/NADH ratio in DFCs that suggest not only "glycolytic metabotype" of DFCs but also Mitochondrial Dysfunction Associated Senescence (MiDAS). Both changes are highly associated with the induction of cellular senescence. Hence, both AMPK activation and inhibition promote the induction of cellular senecence of DFCs.

Keywords:  AMP-kinase; Autophagy; Cellular senescence; Dental follicle cells; Metformin

DOI:  https://doi.org/10.1016/j.exger.2022.112071
Front Cell Infect Microbiol. 2022 ;12 1060202

PfATP4 inhibitors in the Medicines for Malaria Venture Malaria Box and Pathogen Box block the schizont-to-ring transition by inhibiting egress rather than invasion.

Claudia B G Barnes, Madeline G Dans, Thorey K Jonsdottir, Brendan S Crabb, Paul R Gilson.

  The cation efflux pump Plasmodium falciparum ATPase 4 (PfATP4) maintains Na+ homeostasis in malaria parasites and has been implicated in the mechanism of action of many structurally diverse antimalarial agents, including >7% of the antimalarial compounds in the Medicines for Malaria Venture's 'Malaria Box' and 'Pathogen Box'. Recent screens of the 'Malaria Box' and 'Pathogen Box' revealed that many PfATP4 inhibitors prevent parasites from exiting their host red blood cell (egress) or entering new host cells (invasion), suggesting that these compounds may have additional molecular targets involved in egress or invasion. Here, we demonstrate that five PfATP4 inhibitors reduce egress but not invasion. These compounds appear to inhibit egress by blocking the activation of protein kinase G, an enzyme that, once stimulated, rapidly activates parasite egress. We establish a direct link between egress and PfATP4 function by showing that the inhibition of egress is attenuated in a Na+-depleted environment and in parasites with a mutation in pfatp4. Finally, we show that PfATP4 inhibitors induce host cell lysis when administered prior to the completion of parasite replication. Since host cell lysis mimics egress but is not followed by invasion, this phenomenon likely explains why several PfATP4 inhibitors were previously classified as invasion inhibitors. Collectively, our results confirm that PfATP4-mediated Na+ efflux is critical to the regulation of parasite egress.

Keywords:  Malaria Box; Pathogen Box; PfATP4; Plasmodium falciparum; egress; invasion; malaria; sodium ions

DOI:  https://doi.org/10.3389/fcimb.2022.1060202
Genes (Basel). 2022 Dec 16. pii: 2384. [Epub ahead of print]13(12):

AMPK Promotes Larval Metamorphosis of Mytilus coruscus.

Wen Zhang, Yuyi Wang, Xiaomeng Hu, Zhongsheng Zhou, Youting Zhu, Xiao Liang, Jin-Long Yang.

  Metamorphosis is a critical process in the transition from planktonic life to benthic life for marine invertebrates, which is accompanied by a large amount of energy consumption. Previous studies have proved that AMP-activated protein kinase (AMPK), as a vital energy regulator, plays a prominent role in mediating the growth and development of terrestrial animals. However, its function in the growth and development of marine invertebrates, especially in metamorphosis, remains elusive. This study explored the function of AMPK in the larval metamorphosis of Mytilus coruscus. The full-length cDNA of AMPK genes in M. coruscus was cloned and characterized, which is composed of three subunits, McAMPKα, McAMPKβ, and McAMPKγ. Pharmacological tests demonstrated that through the application of an AMPK activator, AMP substantially enhanced the larval metamorphosis rate (p < 0.05). By contrast, the larval metamorphosis rate decreased significantly after being treated with the AMPK inhibitor Compound C (p < 0.05). McAMPK gene knock-down resulted in a reduction in McAMPK gene expression (p < 0.05), and the larval metamorphosis of M. coruscus was significantly restrained (p < 0.05). These results indicated that AMPK signaling is vital in the larval metamorphosis of M. coruscus, which advances further understanding in exploring the molecular mechanisms in the metamorphosis of marine invertebrate larvae.

Keywords:  AMPK genes; Mytilus coruscus; RNA interference; activator; inhibitor; metamorphosis

DOI:  https://doi.org/10.3390/genes13122384
Front Cell Infect Microbiol. 2022 ;12 1063407

Targeting malaria parasites with novel derivatives of azithromycin.

Amy L Burns, Brad E Sleebs, Maria Gancheva, Kimberley T McLean, Ghizal Siddiqui, Henrietta Venter, James G Beeson, Ryan O'Handley, Darren J Creek, Shutao Ma, Sonja Frölich, Christopher D Goodman, Geoffrey I McFadden, Danny W Wilson.

   Introduction: The spread of artemisinin resistant Plasmodium falciparum parasites is of global concern and highlights the need to identify new antimalarials for future treatments. Azithromycin, a macrolide antibiotic used clinically against malaria, kills parasites via two mechanisms: 'delayed death' by inhibiting the bacterium-like ribosomes of the apicoplast, and 'quick-killing' that kills rapidly across the entire blood stage development.
Methods: Here, 22 azithromycin analogues were explored for delayed death and quick-killing activities against P. falciparum (the most virulent human malaria) and P. knowlesi (a monkey parasite that frequently infects humans).
Results: Seventeen analogues showed improved quick-killing against both Plasmodium species, with up to 38 to 20-fold higher potency over azithromycin after less than 48 or 28 hours of treatment for P. falciparum and P. knowlesi, respectively. Quick-killing analogues maintained activity throughout the blood stage lifecycle, including ring stages of P. falciparum parasites (<12 hrs treatment) and were >5-fold more selective against P. falciparum than human cells. Isopentenyl pyrophosphate supplemented parasites that lacked an apicoplast were equally sensitive to quick-killing analogues, confirming that the quick killing activity of these drugs was not directed at the apicoplast. Further, activity against the related apicoplast containing parasite Toxoplasma gondii and the gram-positive bacterium Streptococcus pneumoniae did not show improvement over azithromycin, highlighting the specific improvement in antimalarial quick-killing activity. Metabolomic profiling of parasites subjected to the most potent compound showed a build-up of non-haemoglobin derived peptides that was similar to chloroquine, while also exhibiting accumulation of haemoglobin-derived peptides that was absent for chloroquine treatment.
Discussion: The azithromycin analogues characterised in this study expand the structural diversity over previously reported quick-killing compounds and provide new starting points to develop azithromycin analogues with quick-killing antimalarial activity.

Keywords:  Plasmodium; antimalarial; azithromycin; malaria; quick-killing

DOI:  https://doi.org/10.3389/fcimb.2022.1063407
Nat Commun. 2022 Dec 19. 13(1): 7822

Bioenergetics of pollen tube growth in Arabidopsis thaliana revealed by ratiometric genetically encoded biosensors.

Jinhong Liu, Shey-Li Lim, Jia Yi Zhong, Boon Leong Lim.

Pollen tube is the fastest-growing plant cell. Its polarized growth process consumes a tremendous amount of energy, which involves coordinated energy fluxes between plastids, the cytosol, and mitochondria. However, how the pollen tube obtains energy and what the biological roles of pollen plastids are in this process remain obscure. To investigate this energy-demanding process, we developed second-generation ratiometric biosensors for pyridine nucleotides which are pH insensitive between pH 7.0 to pH 8.5. By monitoring dynamic changes in ATP and NADPH concentrations and the NADH/NAD+ ratio at the subcellular level in Arabidopsis (Arabidopsis thaliana) pollen tubes, we delineate the energy metabolism that underpins pollen tube growth and illustrate how pollen plastids obtain ATP, NADPH, NADH, and acetyl-CoA for fatty acid biosynthesis. We also show that fermentation and pyruvate dehydrogenase bypass are not essential for pollen tube growth in Arabidopsis, in contrast to other plant species like tobacco and lily.

DOI: https://doi.org/10.1038/s41467-022-35486-w
Nat Metab. 2022 Dec;4(12): 1792-1811

mTORC1 regulates a lysosome-dependent adaptive shift in intracellular lipid species.

Aaron M Hosios, Meghan E Wilkinson, Molly C McNamara, Krystle C Kalafut, Margaret E Torrence, John M Asara, Brendan D Manning.

The mechanistic target of rapamycin complex 1 (mTORC1) senses and relays environmental signals from growth factors and nutrients to metabolic networks and adaptive cellular systems to control the synthesis and breakdown of macromolecules; however, beyond inducing de novo lipid synthesis, the role of mTORC1 in controlling cellular lipid content remains poorly understood. Here we show that inhibition of mTORC1 via small molecule inhibitors or nutrient deprivation leads to the accumulation of intracellular triglycerides in both cultured cells and a mouse tumor model. The elevated triglyceride pool following mTORC1 inhibition stems from the lysosome-dependent, but autophagy-independent, hydrolysis of phospholipid fatty acids. The liberated fatty acids are available for either triglyceride synthesis or β-oxidation. Distinct from the established role of mTORC1 activation in promoting de novo lipid synthesis, our data indicate that mTORC1 inhibition triggers membrane phospholipid trafficking to the lysosome for catabolism and an adaptive shift in the use of constituent fatty acids for storage or energy production.

DOI: https://doi.org/10.1038/s42255-022-00706-6
Curr Protoc. 2022 Dec;2(12): e660

Corrections: Principles of Resonance Energy Transfer.

Ágnes Szabó, János Szöllősi, Peter Nagy.

DOI: https://doi.org/10.1002/cpz1.660
Mol Med Rep. 2023 Feb;pii: 35. [Epub ahead of print]27(2):

Role of the AMPK/SIRT1 pathway in non‑alcoholic fatty liver disease (Review).

Putri Anggreini, Hadi Kuncoro, Sri Adi Sumiwi, Jutti Levita.

  Non‑alcoholic fatty liver disease (NAFLD) is an increasingly prevalent ailment worldwide. Moreover, de novo lipogenesis (DNL) is considered a critical factor in the development of NAFLD; hence, its inhibition is a promising target for the prevention of fatty liver disease. There is evidence to indicate that AMP‑activated protein kinase (AMPK) and sirtuin 1 (SIRT1) may play a crucial role in DNL and are the regulatory proteins in type 2 diabetes mellitus, obesity and cardiovascular disease. Therefore, AMPK and SIRT1 may be promising targets for the treatment of NAFLD. The present review article thus aimed to summarize the findings of clinical studies published during the past decade that suggested the beneficial effects of AMPK and SIRT1, using their specific activators and their combined effects on fatty liver disease.

Keywords:  AMP‑activated protein kinase; mechanism; non‑alcoholic fatty liver disease; randomized control trial; sirtuin 1

DOI:  https://doi.org/10.3892/mmr.2022.12922
J Mol Endocrinol. 2022 Dec 01. pii: JME-22-0149. [Epub ahead of print]

Cryo-electron microscopy structures of human thyroid peroxidase (TPO) in complex with TPO antibodies.

Stuart Baker, Ricardo Núñez Miguel, Daniel Thomas, Michael Powell, Jadwiga Furmaniak, Bernard Rees Smith.

Determination of the structure of the extracellular domain of human thyroid peroxidase (hTPO) by cryo-electron microscopy (cryo-EM) is described. TPO, purified to homogeneity was complexed with the human TPO monoclonal autoantibody 2G4 Fab and also with a mouse monoclonal TPO antibody 4F5 Fab (which competes with autoantibody binding to TPO). Both complexes were analysed by cryo-EM. The two structures (global resolution 3.92Å and 3.4Å for the 2G4 complex and 4F5 complex respectively) show TPO as a monomer with 4 domains; the N-terminal domain, the peroxidase domain (POD), the complement control protein like (CCP) domain and the epidermal growth factor like (EGF) domain which are all visible in the structures. The relative positions of the domains are fixed with a disulphide bond between cysteine residues Cys146 in the POD and Cys756 in the CCP domain preventing significant flexibility of the molecule. The entrance to the enzyme active site, the haem group and the calcium binding site are clearly visible on the opposite side of the TPO molecule from the 2G4 and 4F5 binding sites. Extensive interactions are seen between TPO and the two antibodies which both bind to distinct epitopes on the POD domain, including some residues in the immunodominant region B (IDR-B) mainly via different residues. However the epitopes of the two antibodies contain three shared TPO residues. This is the first high resolution structure of TPO to be reported and it should help guide development of new inhibitors of TPO enzyme activity for therapeutic applications.

DOI: https://doi.org/10.1530/JME-22-0149