bims-toxgon 2024-01-21 papers

bims-toxgon

Biomed News

on Toxoplasma gondii metabolism

Issue of 2024–01–21
nineteen papers selected by
Lakesh Kumar, BITS Pilani

An apicoplast-localized deubiquitinase contributes to the cell growth and apicoplast homeostasis of Toxoplasma gondii.
A Combination of Four Nuclear Targeted Effectors Protects Toxoplasma Against Interferon Gamma Driven Human Host Cell Death During Acute Infection.
Transcriptomic profiling of long non-coding RNAs and messenger RNAs in the liver of mice during Toxoplasma gondii infection.
RPA-CRISPR/Cas9-based method for the detection of Toxoplasma gondii: A proof of concept.
Investigation of the effects of Toxoplasma gondii on behavioral and molecular mechanism in bradyzoite stage.
Toxoplasma gondii infection and brain inflammation: A two-sample mendelian randomization analysis.
Catalytic isoforms of AMP-activated protein kinase differentially regulate IMPDH activity and photoreceptor neuron function.
The yeast AMP-activated protein kinase Snf1 phosphorylates the inositol polyphosphate kinase Kcs1.
Whole-genome CRISPR screens to understand Apicomplexan-host interactions.
Phosphorylation of INF2 by AMPK promotes mitochondrial fission and oncogenic function in endometrial cancer.
Endoplasmic reticulum localized TMEM33 domain-containing protein is crucial for all life cycle stages of the malaria parasite.
Decoding the function of Atg13 phosphorylation reveals a role of Atg11 in bulk autophagy initiation.
Post-translational regulation of the mTORC1 pathway: A switch that regulates metabolism-related gene expression.
Context-dependent role of SIRT3 in cancer.
MDM2 Antagonist Idasanutlin Reduces HDAC1/2 Abundance and Corepressor Partners but Not HDAC3.
Development of potent and selective ULK1/2 inhibitors based on 7-azaindole scaffold with favorable in vivo properties.
SIRT1 regulates endoplasmic reticulum stress-related organ damage.
Proteomic applications in identifying protein-protein interactions.
Virtual docking screening and quantitative structure-activity relationship studies to explore AKT and PI3K inhibitors acting on mTOR in cancers by theoretical biology and medical modeling.

Vet Res. 2024 Jan 17. 55(1): 10

An apicoplast-localized deubiquitinase contributes to the cell growth and apicoplast homeostasis of Toxoplasma gondii.

Jie Xia, Yimin Yang, Xueqiu Chen, Kaiyue Song, Guangxu Ma, Yi Yang, Chaoqun Yao, Aifang Du.

  Toxoplasma gondii is among the most important parasites worldwide. The apicoplast is a unique organelle shared by all Apicomplexan protozoa. Increasing lines of evidence suggest that the apicoplast possesses its own ubiquitination system. Deubiquitination is a crucial step executed by deubiquitinase (DUB) during protein ubiquitination. While multiple components of ubiquitination have been identified in T. gondii, the deubiquitinases involved remain unknown. The aim of the current study was to delineate the localization of TgOTU7 and elucidate its functions. TgOTU7 was specifically localized at the apicoplast, and its expression was largely regulated during the cell cycle. Additionally, TgOTU7 efficiently breaks down ubiquitin chains, exhibits linkage-nonspecific deubiquitinating activity and is critical for the lytic cycle and apicoplast biogenesis, similar to the transcription of the apicoplast genome and the nuclear genes encoding apicoplast-targeted proteins. Taken together, the results indicate that the newly described deubiquitinase TgOTU7 specifically localizes to the apicoplast and affects the cell growth and apicoplast homeostasis of T. gondii.

Keywords:  TgOTU7; Toxoplasma gondii; apicoplast; deubiquitinase; ubiquitination

DOI:  https://doi.org/10.1186/s13567-023-01261-y
bioRxiv. 2023 Dec 25. pii: 2023.12.24.573224. [Epub ahead of print]

A Combination of Four Nuclear Targeted Effectors Protects Toxoplasma Against Interferon Gamma Driven Human Host Cell Death During Acute Infection.

Brittany Henry, L David Sibley, Alex Rosenberg.

In both mice and humans, Type II interferon-gamma (IFNγ) is crucial for regulation of Toxoplasma gondii ( T. gondii ) infection, during acute or chronic phases. To thwart this defense, T. gondii secretes protein effectors hindering the host's immune response. For example, T. gondii relies on the MYR translocon complex to deploy soluble dense granule effectors (GRAs) into the host cell cytosol or nucleus. Recent genome-wide loss-of-function screens in IFNγ-primed primary human fibroblasts identified MYR translocon components as crucial for parasite resistance against IFNγ driven vacuole clearance. However, these screens did not pinpoint specific MYR-dependent GRA proteins responsible for IFNγ signaling blockade, suggesting potential functional redundancy. Our study reveals that T. gondii depends on the MYR translocon complex to prevent host cell death and parasite premature egress in human cells stimulated with IFNγ post-infection, a unique phenotype observed in various human cell lines but not in murine cells. Intriguingly, inhibiting parasite egress did not prevent host cell death, indicating this mechanism is distinct from those described previously. Genome-wide loss-of-function screens uncovered TgIST, GRA16, GRA24, and GRA28 as effectors necessary for a complete block of IFNγ response. GRA24 and GRA28 directly influenced IFNγ driven transcription, GRA24's action depended on its interaction with p38 MAPK, while GRA28 disrupted histone acetyltransferase activity of CBP/p300. Given the intricate nature of the immune response to T. gondii , it appears that the parasite has evolved equally elaborate mechanisms to subvert IFNγ signaling, extending beyond direct interference with the JAK/STAT1 pathway, to encompass other signaling pathways as well.

DOI: https://doi.org/10.1101/2023.12.24.573224
Parasit Vectors. 2024 Jan 16. 17(1): 20

Transcriptomic profiling of long non-coding RNAs and messenger RNAs in the liver of mice during Toxoplasma gondii infection.

Yang Zou, Xing Yang, Chao Chen, He Ma, Hong-Wei Cao, Jing Jiang, Xin-Yu Wei, Xiao-Xuan Zhang.

   BACKGROUND: Toxoplasma gondii is an intracellular protozoan parasite that can infect a wide range of warm-blooded animals, including humans. It poses significant health risks, particularly in immunocompromised individuals and during pregnancy, leading to severe disease manifestations. The liver, being a crucial organ involved in immune response and metabolic regulation, plays a critical role in the host's defense against T. gondii infection.
METHODS: In this study, we utilized RNA sequencing to investigate the expression profiles of long non-coding RNAs (lncRNAs) and messenger RNAs (mRNAs) in the liver of mice infected with T. gondii. By employing this method, we obtained a comprehensive overview of the alterations in gene expression occurring in the liver during infection.
RESULTS: By comparing the infected groups to the control groups, we identified numerous differentially expressed lncRNAs DElncRNAs and DEmRNAs at two stages of infection. Specifically, at the acute infection stage, we found 628 DElncRNAs, and 6346 DEmRNAs. At the chronic infection stage, we identified 385 DElncRNAs and 2513 DEmRNAs. Furthermore, we identified 1959 commonly expressed DEmRNAs, including IL27, Nos2, and Cxcr2, across two infection stages. Enrichment and co-location analyses revealed pathways linked to immune and inflammatory responses during T. gondii infection. Notably, through co-location analysis, our analysis revealed several DElncRNAs, including Gm29156, Gm29157, and Gm28644, which are potentially implicated in the progression of liver inflammation induced by T. gondii. Additionally, functional enrichment analysis disclosed stage-specific characteristics of liver inflammation and immune response, alongside changes in metabolic regulation and immunosuppression pathways.
CONCLUSIONS: Our findings provide valuable insights into the expression patterns of lncRNAs and mRNAs in the liver at different stages of T. gondii infection. We identified potential regulatory factors and pathways implicated in liver inflammation, thereby enhancing our understanding of the molecular mechanisms underlying liver inflammation and immune responses during T. gondii infection. These findings could contribute to the development of targeted therapeutic strategies for liver inflammation in the context of T. gondii infection.

Keywords:  Co-location; Liver; Long non-coding RNAs; RNA sequencing; Stage-specific characteristics; Toxoplasma gondii

DOI:  https://doi.org/10.1186/s13071-023-06053-z
Vet Parasitol. 2024 Jan 14. pii: S0304-4017(24)00003-7. [Epub ahead of print]327 110115

RPA-CRISPR/Cas9-based method for the detection of Toxoplasma gondii: A proof of concept.

Mengchen Wu, Haiyan Wu, Xueqiu Chen, Fei Wu, Guangxu Ma, Aifang Du, Yi Yang.

  Toxoplasma gondii is a widespread and specialized intracellular protozoan pathogen that affects one third of the world' s population, posing a great threat to public health. As the definitive host, cats excrete oocysts and play a crucial role in the transmission of toxoplasmosis. The current diagnostic tools usually require bulky equipment and expertize, which hinders the efficient diagnosis and intervention of Toxoplasma infection in cats. In this study, we combined (RPA) with clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technique to establish an easier method for the detection of T. gondii oocysts in cat fecal samples. The sensitivity, specificity, and practicability of the established RPA-CRISPR/Cas9 method were evaluated using a lateral flow strip, with the limitation of detection determined at 10 plasmid copies/μL (corresponding to about one oocyst), cross reactivity to none of Giardia lamblia, Cryptosporidium sp., Microsporidium biberi and Blastocystis hominis that also commonly found in cats, and comparable performance in detecting T. gondii in clinical samples to conventional PCR amplification. This RPA-CRISPR/Cas9 method provides an alternative to conventional molecular tools used in the clinical diagnosis of Toxoplasma infection in cats and other animals.

Keywords:  CRISPR/Cas9; Clinical samples; Diagnosis; Recombinase polymerase amplification (RPA); Toxoplasma gondii

DOI:  https://doi.org/10.1016/j.vetpar.2024.110115
Brain Res. 2024 Jan 14. pii: S0006-8993(24)00016-7. [Epub ahead of print] 148762

Investigation of the effects of Toxoplasma gondii on behavioral and molecular mechanism in bradyzoite stage.

Guven Akcay, Anzel Bahadir, Yakup Tatar, Mehmet Nuri Atalar, Cahit Babur, Aysegul Taylan Ozkan.

  Toxoplasma gondii is a single-celled parasite that causes a disease called toxoplasmosis. It can reach the central nervous system, but the mechanism of T. gondii disrupting the functioning of these brain regions occurs in bradyzoite stage of parasite, causing brain damage by forming tissue cysts in brain. In our study, the effects of T. gondii on locomotor activity, anxiety, learning and memory, and norepinephrine (NE), levodopa (L-DOPA), dopamine (DA) and 3,4-D-dihydroxyphenylacetic acid (DOPAC) catecholamines in amygdala, striatum, prefrontal cortex and hippocampus regions of the brain were investigated in bradyzoite stage. Twenty male Albino mice Mus musculus, 4-5 weeks old, weighing 20-25 g, were used. T. gondii inoculated to mice intraperitonealy with 48-50-hour passages of T. gondii RH Ankara strain. For intraperitoneal inoculation of mice 5x104 tachyzoites per mouse. No inoculation was made in control group (n: 20). Locomotor activity behavior in open field test (OFT), anxious behavior in elevated plus maze (EPM), and learning behavior in novel object recognition (NOR) tests were evaluated. NE, L-DOPA, DA and DOPAC were measured by HPLC in brain tissues of amygdala, striatum, prefrontal cortex and hippocampus. A decrease was observed in the locomotor activity, anxiety and learning values of the T. gondii group compared to the control group (p < 0.05). The heighten in NE and L-DOPA levels in amygdala tissue of T. gondii group compared to control group, an elevation in NE, L-DOPA, DA and DOPAC levels in striatum tissue, and an increase in levels of NE in prefrontal cortex tissue were detected in monoamine results. In hippocampus tissue, an increase was observed in DA levels, while a decrease was observed in NE, L-DOPA and DOPAC levels. In our study, it has been shown that T. gondii in bradyzoite stage reduces locomotor activity, causes learning and memory impairment, and has anxiogenic effects.

Keywords:  Anxiety; Bradyzoite; Brain; Dopamine; Learning and Memory; Norepinephrine; Toxoplasma gondii

DOI:  https://doi.org/10.1016/j.brainres.2024.148762
Heliyon. 2024 Jan 15. 10(1): e24228

Toxoplasma gondii infection and brain inflammation: A two-sample mendelian randomization analysis.

Yong Yao, Taiyu Shi, Panyin Shu, Yixin Zhang, Hao Gu.

   Background: Toxoplasma gondii is an opportunistic parasitic protozoan that can cause highly fatal toxoplasmic encephalitis when the host immune system is compromised. However, the transition from chronic to acute infection remains poorly understood. In this study, we conducted a 180-day observation of tissue damage and inflammation in the brains of mice infected with T. gondii. Subsequently, we investigated the inflammatory factors that T. gondii infection may alter using two-sample Mendelian randomization (MR) analysis.
Methods: We first established a mouse model of T. gondii infection. Subsequently, the mice were euthanized, the brain tissue collected, and immunohistochemistry and hematoxylin and eosin staining performed to observe tissue damage and inflammatory conditions at various time points. Our study also included a published large-scale genome-wide association study meta-analysis that encompassed the circulating concentrations of 41 cytokines. This dataset included 8293 individuals from three independent population cohorts in Finland. Genetic association data for T. gondii were sourced from the Integrative Epidemiology Unit and European Bioinformatics Institute datasets, which included 5010 and 559 individuals of European ancestry, respectively. To assess the causal relationship between T. gondii infection and inflammatory biomarkers, we applied a two-sample MR.
Results: Inflammation and damage resulting from T. gondii infection varied among the distinct regions of the mouse brain. Based on the MR analysis results, three inflammatory biomarkers were chemically assigned to Chemokines and Others, including IP10 (interferon gamma inducible protein-10), MCP1 (monocyte chemoattractant protein-1), and TRAIL (TNF-related apoptosis-inducing ligand).
Conclusion: Our study commenced with the assessment of tissue damage and progression of inflammation in distinct regions of the mouse brain after T. gondii infection. Subsequently, using MR analysis, we detected potential alterations in inflammatory factors associated with this infection. These findings offer valuable insights into the mechanisms underlying toxoplasmic encephalitis and suggest directions for the prevention and treatment of T. gondii infections.

Keywords:  Inflammatory biomarkers; Mendelian randomization study; Pathological examination; Toxoplasma gondii; Toxoplasmic encephalitis

DOI:  https://doi.org/10.1016/j.heliyon.2024.e24228
JCI Insight. 2024 Jan 16. pii: e173707. [Epub ahead of print]

Catalytic isoforms of AMP-activated protein kinase differentially regulate IMPDH activity and photoreceptor neuron function.

Tae Jun Lee, Yo Sasaki, Philip A Ruzycki, Norimitsu Ban, Joseph B Lin, Hung-Ting Wu, Andrea Santeford, Rajendra S Apte.

  AMP-activated protein kinase (AMPK) plays a crucial role in maintaining ATP homeostasis in photoreceptor neurons. AMPK is a heterotrimeric protein consisting of alpha, beta, and gamma subunits. The independent functions of the two isoforms of the catalytic alpha subunit, PRKAA1 and PRKAA2, are uncharacterized in specialized neurons such as photoreceptors. Here we demonstrate in mice that rod photoreceptors lacking PRKAA2, but not PRKAA1, show altered levels of cGMP, GTP, and ATP, suggesting isoform-specific regulation of photoreceptor metabolism. Furthermore, PRKAA2 deficient mice display visual functional deficits on electroretinography and photoreceptor outer segment structural abnormalities on transmission electron microscopy consistent with neuronal dysfunction, but not neurodegeneration. Phosphoproteomics identified inosine monophosphate dehydrogenase (IMPDH) as a molecular driver of PRKAA2-specific photoreceptor dysfunction, and inhibition of IMPDH improved visual function in Prkaa2 rod photoreceptor knockout mice. These findings highlight a novel, therapeutically targetable PRKAA2 isoform-specific function of AMPK in regulating photoreceptor metabolism and function through a previously uncharacterized mechanism affecting IMPDH activity.

Keywords:  Bioenergetics; Metabolism; Ophthalmology; Protein kinases; Signal transduction

DOI:  https://doi.org/10.1172/jci.insight.173707
J Biol Chem. 2024 Jan 13. pii: S0021-9258(24)00033-4. [Epub ahead of print] 105657

The yeast AMP-activated protein kinase Snf1 phosphorylates the inositol polyphosphate kinase Kcs1.

Sham Sunder, Joshua S Bauman, Stuart J Decker, Alexandra R Lifton, Anuj Kumar.

The yeast Snf1/AMP-activated kinase (AMPK) maintains energy homeostasis, controlling metabolic processes and glucose derepression in response to nutrient levels and environmental cues. Under conditions of nitrogen or glucose limitation, Snf1 regulates pseudohyphal growth, a morphological transition characterized by the formation of extended multicellular filaments. During pseudohyphal growth, Snf1 is required for wild-type levels of inositol polyphosphate (InsP), soluble phosphorylated species of the six-carbon cyclitol inositol that function as conserved metabolic second messengers. InsP levels are established through the activity of a family of inositol kinases, including the yeast inositol polyphosphate kinase Kcs1, which principally generates pyrophosphorylated InsP7. Here, we report that Snf1 regulates Kcs1, affecting Kcs1 phosphorylation and inositol kinase activity. A snf1 kinase-defective mutant exhibits decreased Kcs1 phosphorylation, and Kcs1 is phosphorylated in vivo at Ser residues 537 and 646 during pseudohyphal growth. By in vitro analysis, Snf1 directly phosphorylates Kcs1, predominantly at amino acids 537 and 646. A yeast strain carrying kcs1 encoding Ser-to-Ala point mutations at these residues (kcs1-S537A,S646A) shows elevated levels of pyrophosphorylated InsP7, comparable to InsP7 levels observed upon deletion of SNF1. The kcs1-S537A,S646A mutant exhibits decreased pseudohyphal growth, invasive growth, and cell elongation. Transcriptional profiling indicates extensive perturbation of metabolic pathways in kcs1-S537A,S646A. Growth of kcs1-S537A,S646A is affected on medium containing sucrose and antimycin A, consistent with decreased Snf1p signaling. This work identifies Snf1 phosphorylation of Kcs1, collectively highlighting the interconnectedness of AMPK activity and InsP signaling in coordinating nutrient availability, energy homoeostasis, and cell growth.

DOI: https://doi.org/10.1016/j.jbc.2024.105657
Mol Microbiol. 2024 Jan 15.

Whole-genome CRISPR screens to understand Apicomplexan-host interactions.

Eva Hesping, Justin A Boddey.

  Apicomplexan parasites are aetiological agents of numerous diseases in humans and livestock. Functional genomics studies in these parasites enable the identification of biological mechanisms and protein functions that can be targeted for therapeutic intervention. Recent improvements in forward genetics and whole-genome screens utilising CRISPR/Cas technology have revolutionised the functional analysis of genes during Apicomplexan infection of host cells. Here, we highlight key discoveries from CRISPR/Cas9 screens in Apicomplexa or their infected host cells and discuss remaining challenges to maximise this technology that may help answer fundamental questions about parasite-host interactions.

Keywords:  Apicomplexa; CRISPR; Cas9; forward genetics; host; parasite

DOI:  https://doi.org/10.1111/mmi.15221
Cell Death Dis. 2024 Jan 17. 15(1): 65

Phosphorylation of INF2 by AMPK promotes mitochondrial fission and oncogenic function in endometrial cancer.

Yan Ding, Zeheng Lv, Wenxin Cao, Wenming Shi, Qizhi He, Kun Gao.

Mitochondria are highly dynamic organelles capable of altering their sizes and shapes to maintain metabolic balance through coordinated fission and fusion processes. In various cancer types, mitochondrial hyperfragmentation has been frequently observed, contributing to the progression of cancer toward metastasis. Inverted formin 2 (INF2), which resides in the endoplasmic reticulum (ER), has been found to accelerate actin polymerization and drive mitochondrial fission. In this study, we demonstrate that INF2 expression is significantly upregulated in endometrial cancer (EC) and is associated with a poor prognosis in EC patients. INF2 promotes anchorage-dependent and independent EC cell growth in part by facilitating mitochondrial fission. Furthermore, in conditions of energy stress, AMP-activated protein kinase (AMPK) phosphorylates INF2 at Ser1077, leading to increased localization of INF2 to the ER and enhanced recruitment of the dynamin-related protein 1 (DRP1) to mitochondria. This AMPK-mediated phosphorylation of INF2 at Ser1077 facilitates mitochondrial division and promotes EC cell growth. Pathological examination using immunohistochemical analyses revealed a positive correlation between AMPK activity and phosphorylated INF2 (Ser1077) in EC specimens. Collectively, our findings uncover novel molecular mechanisms involving the AMPK-INF2 axis, which regulates mitochondrial dynamics and malignant cell growth in EC.

DOI: https://doi.org/10.1038/s41419-024-06431-0
Mol Microbiol. 2024 Jan 18.

Endoplasmic reticulum localized TMEM33 domain-containing protein is crucial for all life cycle stages of the malaria parasite.

Mohd Kamil, Umit Yasar Kina, Habibe Nur Atmaca, Sinem Unal, Gozde Deveci, Pinar Burak, Ahmed S I Aly.

  Endoplasmic reticulum (ER) plays a pivotal role in the regulation of stress responses in multiple eukaryotic cells. However, little is known about the effector mechanisms that regulate stress responses in ER of the malaria parasite. Herein, we aimed to identify the importance of a transmembrane protein 33 (TMEM33)-domain-containing protein in life cycle of the rodent malaria parasite Plasmodium berghei. TMEM33 is an ER membrane-resident protein that is involved in regulating stress responses in various eukaryotic cells. A C-terminal tagged TMEM33 was localized in the ER throughout the blood and mosquito stages of development. Targeted deletion of TMEM33 confirmed its importance for asexual blood stages and ookinete development, in addition to its essential role for sporozoite infectivity in the mammalian host. Pilot scale analysis shows that the loss of TMEM33 results in the initiation of ER stress response and induction of autophagy. Our findings conclude an important role of TMEM33 in the development of all life cycle stages of the malaria parasite, which indicates its potential as an antimalarial target.

Keywords:   Plasmodium berghei ; TMEM33; autophagy; drug resistance; endoplasmic reticulum; malaria

DOI:  https://doi.org/10.1111/mmi.15228
EMBO Rep. 2024 Jan 17.

Decoding the function of Atg13 phosphorylation reveals a role of Atg11 in bulk autophagy initiation.

Anuradha Bhattacharya, Raffaela Torggler, Wolfgang Reiter, Natalie Romanov, Mariya Licheva, Akif Ciftci, Muriel Mari, Lena Kolb, Dominik Kaiser, Fulvio Reggiori, Gustav Ammerer, David M Hollenstein, Claudine Kraft.

  Autophagy is initiated by the assembly of multiple autophagy-related proteins that form the phagophore assembly site where autophagosomes are formed. Atg13 is essential early in this process, and a hub of extensive phosphorylation. How these multiple phosphorylations contribute to autophagy initiation, however, is not well understood. Here we comprehensively analyze the role of phosphorylation events on Atg13 during nutrient-rich conditions and nitrogen starvation. We identify and functionally characterize 48 in vivo phosphorylation sites on Atg13. By generating reciprocal mutants, which mimic the dephosphorylated active and phosphorylated inactive state of Atg13, we observe that disrupting the dynamic regulation of Atg13 leads to insufficient or excessive autophagy, which are both detrimental to cell survival. We furthermore demonstrate an involvement of Atg11 in bulk autophagy even during nitrogen starvation, where it contributes together with Atg1 to the multivalency that drives phase separation of the phagophore assembly site. These findings reveal the importance of post-translational regulation on Atg13 early during autophagy initiation, which provides additional layers of regulation to control bulk autophagy activity and integrate cellular signals.

Keywords:  Atg1 Kinase Complex; Atg11; Atg13; Autophagy; PAS Formation

DOI:  https://doi.org/10.1038/s44319-023-00055-9
Biochim Biophys Acta Gene Regul Mech. 2024 Jan 17. pii: S1874-9399(24)00001-4. [Epub ahead of print] 195005

Post-translational regulation of the mTORC1 pathway: A switch that regulates metabolism-related gene expression.

Yitao Wang, Tobias Engel, Xinchen Teng.

  The mechanistic target of rapamycin complex 1 (mTORC1) is a kinase complex that plays a crucial role in coordinating cell growth in response to various signals, including amino acids, growth factors, oxygen, and ATP. Activation of mTORC1 promotes cell growth and anabolism, while its suppression leads to catabolism and inhibition of cell growth, enabling cells to withstand nutrient scarcity and stress. Dysregulation of mTORC1 activity is associated with numerous diseases, such as cancer, metabolic disorders, and neurodegenerative conditions. This review focuses on how post-translational modifications, particularly phosphorylation and ubiquitination, modulate mTORC1 signaling pathway and their consequential implications for pathogenesis. Understanding the impact of phosphorylation and ubiquitination on the mTORC1 signaling pathway provides valuable insights into the regulation of cellular growth and potential therapeutic targets for related diseases.

Keywords:  Phosphorylation; Transcription factor; Ubiquitination; mTOR; mTORC1 signaling pathway

DOI:  https://doi.org/10.1016/j.bbagrm.2024.195005
Trends Pharmacol Sci. 2024 Jan 18. pii: S0165-6147(23)00262-6. [Epub ahead of print]

Context-dependent role of SIRT3 in cancer.

Jin Zhang, Jing Ye, Shiou Zhu, Bo Han, Bo Liu.

  Sirtuin 3 (SIRT3), an NAD+-dependent deacetylase, plays a key role in the modulation of metabolic reprogramming and regulation of cell death, as well as in shaping tumor phenotypes. Owing to its critical role in determining tumor-type specificity or the direction of tumor evolution, the development of small-molecule modulators of SIRT3, including inhibitors and activators, is of significant interest. In this review, we discuss recent studies on the oncogenic or tumor-suppressive functions of SIRT3, evaluate advances in SIRT3-targeted drug discovery, and present potential avenues for the design of small-molecule modulators of SIRT3 for cancer therapy.

Keywords:  cancer therapy; drug development; metabolic reprogramming; sirtuin 3 (SIRT3); small-molecule modulator

DOI:  https://doi.org/10.1016/j.tips.2023.12.005
ACS Med Chem Lett. 2024 Jan 11. 15(1): 93-98

MDM2 Antagonist Idasanutlin Reduces HDAC1/2 Abundance and Corepressor Partners but Not HDAC3.

Joshua P Smalley, Shaun M Cowley, James T Hodgkinson.

Histone deacetylases 1-3 (HDAC1, HDAC2, and HDAC3) and their associated corepressor complexes play important roles in regulating chromatin structure and gene transcription. HDAC enzymes are also validated drug targets for oncology and offer promise toward new drugs for neurodegenerative diseases and cardiovascular diseases. We synthesized four novel heterobifunctional molecules designed to recruit the mouse double minute 2 homologue (MDM2) E3 ligase to degrade HDAC1-3 utilizing the MDM2 inhibitor idasanutlin, known as proteolysis targeting chimeras (PROTACs). Idasanutlin inhibits the MDM2-P53 protein-protein interaction and is in clinical trials. Although two MDM2-recruiting heterobifunctional molecules reduced HDAC1 and HDAC2 abundance with complete selectivity over HDAC3 and reduced HDAC1/2 corepressor components LSD1 and SIN3A, we were surprised to observe that idasanutlin alone was also capable of this effect. This finding suggests an association between the MDM2 E3 ligase and HDAC1/2 corepressor complexes, which could be important for designing future dual/bifunctional HDAC- and MDM2-targeting therapeutics, such as PROTACs.

DOI: https://doi.org/10.1021/acsmedchemlett.3c00449
Eur J Med Chem. 2023 Dec 29. pii: S0223-5234(23)01068-1. [Epub ahead of print]266 116101

Development of potent and selective ULK1/2 inhibitors based on 7-azaindole scaffold with favorable in vivo properties.

Alisa Morozova, Sean Chin Chan, Simon Bayle, Luxin Sun, Dylan Grassie, Anna Iermolaieva, Mahalakshmi N Kalaga, Sylvia Frydman, Samer Sansil, Ernst Schönbrunn, Derek Duckett, Andrii Monastyrskyi.

  The UNC-51-like kinase-1 (ULK1) is one of the central upstream regulators of the autophagy pathway, represents a key target for the development of molecular probes to abrogate autophagy and explore potential therapeutic avenues. Here we report the discovery, structure-activity and structure-property relationships of selective, potent, and cell-active ULK1/2 inhibitors based on a 7-azaindole scaffold. Using structure-based drug design, we have developed a series of analogs with excellent binding affinity and biochemical activity against ULK1/2 (IC50 < 25 nM). The validation of cellular target engagement for these compounds was achieved through the employment of the ULK1 NanoBRET intracellular kinase assay. Notably, we have successfully solved the crystal structure of the lead compound, MR-2088, bound to the active site of ULK1. Moreover, the combination treatment of MR-2088 with known KRAS→RAF→MEK→ERK pathway inhibitors, such as trametinib, showed promising synergistic effect in vitro using H2030 (KRASG12C) cell lines. Lastly, our findings underscore MR-2088's potential to inhibit starvation/stimuli-induced autophagic flux, coupled with its suitability for in vivo studies based on its pharmacokinetic properties.

Keywords:  7-Azaindole; Autophagy; KRAS; Non-small cell lung cancer; ULK1/2 kinase

DOI:  https://doi.org/10.1016/j.ejmech.2023.116101
Acta Histochem. 2024 Jan 17. pii: S0065-1281(24)00002-3. [Epub ahead of print]126(1): 152134

SIRT1 regulates endoplasmic reticulum stress-related organ damage.

He Hu, Weichao Xu, Yan Li, Zhicheng Wang, Siyue Wang, Yansheng Liu, Minan Bai, Yingying Lou, Qian Yang.

  Endoplasmic reticulum (ER) stress plays a key role in the pathogenesis of several organ damages. Studies show that excessive ER stress (ERS) can destroy cellular homeostasis, causing cell damage and physiological dysfunction in various organs. In recent years, Sirtuin1 (SIRT1) has become a research hotspot on ERS. Increasing evidence suggests that SIRT1 plays a positive role in various ERS-induced organ damage via multiple mechanisms, including inhibiting cellular apoptosis and promoting autophagy. SIRT1 can also alleviate liver, heart, lung, kidney, and intestinal damage by inhibiting ERS. We discuss the possible mechanism of SIRT1, explore potential therapeutic targets of diseases, and provide a theoretical basis for treating ERS-related diseases.

Keywords:  Endoplasmic reticulum stress; Organ damage; Silencing of the regulatory protein 1

DOI:  https://doi.org/10.1016/j.acthis.2024.152134
Adv Protein Chem Struct Biol. 2024 ;pii: S1876-1623(23)00059-7. [Epub ahead of print]138 1-48

Proteomic applications in identifying protein-protein interactions.

Benjamin T Veenstra, Timothy D Veenstra.

  There are many things that can be used to characterize a protein. Size, isoelectric point, hydrophobicity, structure (primary to quaternary), and subcellular location are just a few parameters that are used. The most important feature of a protein, however, is its function. While there are many experiments that can indicate a protein's role, identifying the molecules it interacts with is probably the most definitive way of determining its function. Owing to technology limitations, protein interactions have historically been identified on a one molecule per experiment basis. The advent of high throughput multiplexed proteomic technologies in the 1990s, however, made identifying hundreds and thousands of proteins interactions within single experiments feasible. These proteomic technologies have dramatically increased the rate at which protein-protein interactions (PPIs) are discovered. While the improvement in mass spectrometry technology was an early driving force in the rapid pace of identifying PPIs, advances in sample preparation and chromatography have recently been propelling the field. In this chapter, we will discuss the importance of identifying PPIs and describe current state-of-the-art technologies that demonstrate what is currently possible in this important area of biological research.

Keywords:  Chemical-crosslinking; Co-fractionation; Immunoaffinity; Mass spectrometry; Protein complexes; Protein identification; Proteomics; Proximity labeling; Yeast-two-hybrid screening

DOI:  https://doi.org/10.1016/bs.apcsb.2023.04.001
Contemp Oncol (Pozn). 2023 ;27(3): 155-162

Virtual docking screening and quantitative structure-activity relationship studies to explore AKT and PI3K inhibitors acting on mTOR in cancers by theoretical biology and medical modeling.

Ilham Kandoussi, Hanane Abbou, Ghyzlane El Haddoumi, Mariam Mansouri, Lahcen Belyamani, Azeddine Ibrahimi.

   Introduction: The mechanistic target of rapamycin (mTOR) coordinates the growth and metabolism of eukaryotic cells with a central role in the regulation of many fundamental cellular processes. It is strongly connected to phosphatidylinositol 3-kinase (PI3K) and AKT signaling. Activation of the PI3K/AKT/mTOR pathway leads to a profound disruption in the control of cell growth and survival, which ultimately leads to competitive growth advantage, metastatic competence, angiogenesis and therapeutic resistance.
Material and methods: To explore the common competitive adenosine triphosphate (ATP) inhibitors PI3K/AKT and PI3K/mTOR, we built a 2D mTOR-SAR model that predicted the bioactivity of AKT and PI3K inhibitors towards mTOR. The interaction of the best inhibitors was evaluated by docking analysis and compared to that of the standard AZ8055 and XL388 inhibitors.
Results: A mechanistic target of rapamycin-quantitative structure-activity relationship (mTOR-QSAR) model with a correlation coefficient (R2) of 0.80813 and a root mean square error of 0.17756 was obtained, validated and evaluated by a cross-validation leave-one-out method. The best predicted AKT and PI3K inhibitor pIC50 activities were 9.36-9.95 and 9.23-9.87 respectively.
Conclusions: After docking and several comparisons, the inhibitors with better predictions showed better affinity and interaction with mTOR compared to AZ8055 and XL388, so we have found that 2 AKT inhibitors and 9 mTOR inhibitors met the Lipinski and Veber criteria and could be future drugs.

Keywords:  PI3K/AKT/mTOR; QSAR; docking; dual ATP inhibitors; virtual screening

DOI:  https://doi.org/10.5114/wo.2023.133709