bims-mibica 2023-11-26 papers

bims-mibica

Biomed News

on Mitochondrial bioenergetics in cancer

Issue of 2023‒11‒26
24 papers selected by
Kelsey Fisher-Wellman, East Carolina University

Heme Biosynthesis is Crucial for Cell Survival and Mitochondrial OXPHOS after X Irradiation.
Proline and dihydroorotate dehydrogenase promote a hyper-proliferative state and dampen ferroptosis in cancer cells by rewiring mitochondrial redox metabolism.
High Fat Diet-Induced Obesity Dysregulates Splenic B Cell Mitochondrial Activity.
Cell-specific modulation of mitochondrial respiration and metabolism by the pro-apoptotic Bcl-2 family members Bax and Bak.
KATP channel activity and slow oscillations in pancreatic beta cells are regulated by mitochondrial ATP production.
Mitochondrial nucleoid condensates drive peripheral fission through high membrane curvature.
A century of the Warburg effect.
Inferring mitochondrial and cytosolic metabolism by coupling isotope tracing and deconvolution.
Vitamin B5 Supports Oncogenic Metabolism in MYC-Driven Breast Cancer.
Mitochondrial phosphoproteomes are functionally specialized across tissues.
Comprehensive Identification of Mitochondrial Pseudogenes (NUMTs) in the Human Telomere-to-Telomere Reference Genome.
Adaptive changes in tumor cells in response to reductive stress.
Proteome-wide tagging with an H2O2 biosensor reveals highly localized and dynamic redox microenvironments.
Arvanil induces ferroptosis of hepatocellular carcinoma by binding to MICU1.
An alternative NURF complex sustains acute myeloid leukemia by regulating the accessibility of insulator regions.
Mitochondrial hyperfusion induces metabolic remodeling in lung endothelial cells by modifying the activities of electron transport chain complexes I and III.
Synthesis and Preclinical Evaluation of PSMA-Targeted 111In-Radioconjugates Containing a Mitochondria-Tropic Triphenylphosphonium Carrier.
Phosphorus Chemistry at the Roots of Bioenergetics: Ligand Permutation as the Molecular Basis of the Mechanism of ATP Synthesis/Hydrolysis by FOF1-ATP Synthase.
Allosteric SHP2 inhibition increases apoptotic dependency on BCL2 and synergizes with venetoclax in FLT3- and KIT-mutant AML.
In vivo imaging of mitochondrial DNA mutations using an integrated nano Cas12a sensor.
Exploring the Role of Mitochondrial Hydrogen Sulfide in Maintaining Polarity and mtDNA Integrity with a Multichannel Fluorescent Probe.
SETD2 deficiency accelerates sphingomyelin accumulation and promotes the development of renal cancer.
A comprehensive analysis of SLC25A1 expression and its oncogenic role in pan-cancer.
Hydroquinone-selected chronic myelogenous leukemia cells are sensitive to chloroquine-induced cytotoxicity via MCL1 suppression and glycolysis inhibition.

Radiat Res. 2023 Nov 21.

Heme Biosynthesis is Crucial for Cell Survival and Mitochondrial OXPHOS after X Irradiation.

Tomoki Bo, Koen Van Wijk, Osamu Nakajima.

Heme is an essential component of the hemoproteins involved in the mitochondrial electron transport chain (ETC). Cancer cells have been reported to display high heme levels and increased activity of heme-containing proteins. Consistently, inhibition of heme biosynthesis by the ALAD inhibitor succinylacetone (SA) has been shown to reduce tumor cell survival. These observations indicate that heme biosynthesis is essential for cancer cell proliferation. X irradiation has been shown to increase mitochondrial mass, membrane potential, oxygen consumption, reactive oxygen species (ROS) production, and ATP synthesis. This finding suggests that radiation activates mitochondrial oxidative phosphorylation (OXPHOS). However, although heme is an essential component of the mitochondrial ETC, whether radiation influences heme biosynthesis remains unclear. In this study, we evaluated heme biosynthesis activity after X irradiation and examined the effects of heme biosynthesis inhibition by SA on cellular radiosensitivity and mitochondrial OXPHOS function. We demonstrated that X irradiation significantly increased ALAS1 mRNA levels and cellular heme content. Inhibition of heme biosynthesis by SA significantly decreased cellular heme content and sensitized cancer cells to radiation. We also showed that SA reduced cellular ATP levels, mitochondrial membrane potential, and mitochondrial ROS production, suggesting mitochondrial OXPHOS dysfunction. SA decreased the expression of mitochondrial heme-related proteins COX2 and cytochrome c but did not influence COX1 and VDAC expression. These results indicate that inhibition of heme biosynthesis decreased mitochondrial ETC protein expression and OXPHOS activity, which triggered cellular ATP depletion and radiosensitization after X irradiation. In summary, heme biosynthesis is upregulated by X irradiation and is essential for mitochondrial OXPHOS and cell survival.

DOI: https://doi.org/10.1667/RADE-23-00035.1
Biochim Biophys Acta Mol Cell Res. 2023 Nov 21. pii: S0167-4889(23)00212-4. [Epub ahead of print] 119639

Proline and dihydroorotate dehydrogenase promote a hyper-proliferative state and dampen ferroptosis in cancer cells by rewiring mitochondrial redox metabolism.

Ryan J Mailloux.

  Redox realignment is integral to the initiation, progression, and metastasis of cancer. This requires considerable metabolic rewiring to induce aberrant shifts in redox homeostasis that favor high hydrogen peroxide (H2O2) generation for the induction of a hyper-proliferative state. The ability of tumor cells to thrive under the oxidative burden imposed by this high H2O2 is achieved by increasing antioxidant defenses. This shift in the redox stress signaling threshold (RST) also dampens ferroptosis, an iron (Fe)-dependent form of cell death activated by oxidative distress and lipid peroxidation reactions. Mitochondria are central to the malignant transformation of normal cells to cancerous ones since these organelles supply building blocks for anabolism, govern ferroptosis, and serve as the major source of cell H2O2. This review summarizes advances in understanding the rewiring of redox reactions in mitochondria to promote carcinogenesis, focusing on how cancer cells hijack the electron transport chain (ETC) to promote proliferation and evasion of ferroptosis. I then apply emerging concepts in redox homeodynamics to discuss how the rewiring of the Krebs cycle and ETC promotes shifts in the RST to favor high rates of H2O2 generation for cell signaling. This discussion then focuses on proline dehydrogenase (PRODH) and dihydroorotate dehydrogenase (DHODH), two enzymes over expressed in cancers, and how their link to one another through the coenzyme Q10 (CoQ) pool generates a redox connection that forms a H2O2 signaling platform and pyrimidine synthesome that favors a hyper-proliferative state and disables ferroptosis.

Keywords:  Dihydroorotate dehydrogenase;; Ferroptosis; Hydrogen peroxide; Proline dehydrogense; Redox stress signaling threshold

DOI:  https://doi.org/10.1016/j.bbamcr.2023.119639
Nutrients. 2023 Nov 17. pii: 4807. [Epub ahead of print]15(22):

High Fat Diet-Induced Obesity Dysregulates Splenic B Cell Mitochondrial Activity.

Anandita Pal, Chien-Te Lin, Ilya Boykov, Emily Benson, Grahame Kidd, Kelsey H Fisher-Wellman, P Darrell Neufer, Saame Raza Shaikh.

  Diet-induced obesity impairs mitochondrial respiratory responses in tissues that are highly metabolically active, such as the heart. However, less is known about the impact of obesity on the respiratory activity of specific cell types, such as splenic B cells. B cells are of relevance, as they play functional roles in obesity-induced insulin resistance, inflammation, and responses to infection. Here, we tested the hypothesis that high-fat-diet (HFD)-induced obesity could impair the mitochondrial respiration of intact and permeabilized splenic CD19+ B cells isolated from C57BL/6J mice and activated ex vivo with lipopolysaccharide (LPS). High-resolution respirometry was used with intact and permeabilized cells. To reveal potential mechanistic targets by which HFD-induced obesity dysregulates B cell mitochondria, we conducted proteomic analyses and 3D serial block face scanning electron microscopy (SBFEM). High-resolution respirometry revealed that intact LPS-stimulated B cells of obese mice, relative to controls, displayed lower ATP-linked, as well as maximal uncoupled, respiration. To directly investigate mitochondrial function, we used permeabilized LPS-stimulated B cells, which displayed increased H2O2 emission and production with obesity. We also examined oxidative phosphorylation efficiency simultaneously, which revealed that oxygen consumption and ATP production were decreased in LPS-stimulated B cells with obesity relative to controls. Despite minimal changes in total respiratory complex abundance, in LPS-stimulated B cells of obese mice, three of the top ten most downregulated proteins were all accessory subunits of respiratory complex I. SBFEM showed that B cells of obese mice, compared to controls, underwent no change in mitochondrial cristae integrity but displayed increased mitochondrial volume that was linked to bioenergetic function. Collectively, these results establish a proof of concept that HFD-induced obesity dysregulates the mitochondrial bioenergetic metabolism of activated splenic B cells.

Keywords:  B cells; high fat diet; mitochondria; obesity; proteome

DOI:  https://doi.org/10.3390/nu15224807
Apoptosis. 2023 Nov 24.

Cell-specific modulation of mitochondrial respiration and metabolism by the pro-apoptotic Bcl-2 family members Bax and Bak.

Dana Sovilj, Cristina Daniela Kelemen, Sarka Dvorakova, Renata Zobalova, Helena Raabova, Jan Kriska, Zuzana Hermanova, Tomas Knotek, Miroslava Anderova, Pavel Klener, Vlada Filimonenko, Jiri Neuzil, Ladislav Andera.

  Proteins from the Bcl-2 family play an essential role in the regulation of apoptosis. However, they also possess cell death-unrelated activities that are less well understood. This prompted us to study apoptosis-unrelated activities of the Bax and Bak, pro-apoptotic members of the Bcl-2 family. We prepared Bax/Bak-deficient human cancer cells of different origin and found that while respiration in the glioblastoma U87 Bax/Bak-deficient cells was greatly enhanced, respiration of Bax/Bak-deficient B lymphoma HBL-2 cells was slightly suppressed. Bax/Bak-deficient U87 cells also proliferated faster in culture, formed tumours more rapidly in mice, and showed modulation of metabolism with a considerably increased NAD+/NADH ratio. Follow-up analyses documented increased/decreased expression of mitochondria-encoded subunits of respiratory complexes and stabilization/destabilization of the mitochondrial transcription elongation factor TEFM in Bax/Bak-deficient U87 and HBL-2 cells, respectively. TEFM downregulation using shRNAs attenuated mitochondrial respiration in Bax/Bak-deficient U87 as well as in parental HBL-2 cells. We propose that (post)translational regulation of TEFM levels in Bax/Bak-deficient cells modulates levels of subunits of mitochondrial respiratory complexes that, in turn, contribute to respiration and the accompanying changes in metabolism and proliferation in these cells.

Keywords:  Bak; Bax; Cell proliferation; Metabolism; Mitochondrial respiration; TEFM

DOI:  https://doi.org/10.1007/s10495-023-01917-2
J Physiol. 2023 Nov 20.

KATP channel activity and slow oscillations in pancreatic beta cells are regulated by mitochondrial ATP production.

Jeremías Corradi, Benjamin Thompson, Patrick A Fletcher, Richard Bertram, Arthur S Sherman, Leslie S Satin.

  Pancreatic beta cells secrete insulin in response to plasma glucose. The ATP-sensitive potassium channel (KATP ) links glucose metabolism to islet electrical activity in these cells by responding to increased cytosolic [ATP]/[ADP]. It was recently proposed that pyruvate kinase (PK) in close proximity to beta cell KATP locally produces the ATP that inhibits KATP activity. This proposal was largely based on the observation that applying phosphoenolpyruvate (PEP) and ADP to the cytoplasmic side of excised inside-out patches inhibited KATP . To test the relative contributions of local vs. mitochondrial ATP production, we recorded KATP activity using mouse beta cells and INS-1 832/13 cells. In contrast to prior reports, we could not replicate inhibition of KATP activity by PEP + ADP. However, when the pH of the PEP solutions was not corrected for the addition of PEP, strong channel inhibition was observed as a result of the well-known action of protons to inhibit KATP . In cell-attached recordings, perifusing either a PK activator or an inhibitor had little or no effect on KATP channel closure by glucose, further suggesting that PK is not an important regulator of KATP . In contrast, addition of mitochondrial inhibitors robustly increased KATP activity. Finally, by measuring the [ATP]/[ADP] responses to imposed calcium oscillations in mouse beta cells, we found that oxidative phosphorylation could raise [ATP]/[ADP] even when ADP was at its nadir during the burst silent phase, in agreement with our mathematical model. These results indicate that ATP produced by mitochondrial oxidative phosphorylation is the primary controller of KATP in pancreatic beta cells. KEY POINTS: Phosphoenolpyruvate (PEP) plus adenosine diphosphate does not inhibit KATP activity in excised patches. PEP solutions only inhibit KATP activity if the pH is unbalanced. Modulating pyruvate kinase has minimal effects on KATP activity. Mitochondrial inhibition, in contrast, robustly potentiates KATP activity in cell-attached patches. Although the ADP level falls during the silent phase of calcium oscillations, mitochondria can still produce enough ATP via oxidative phosphorylation to close KATP . Mitochondrial oxidative phosphorylation is therefore the main source of the ATP that inhibits the KATP activity of pancreatic beta cells.

Keywords:  KATP channels; [ATP]/[ADP] ratio; beta cells; glucose response; insulin secretion; oxidative phosphorylation; pH modulation; pyruvate kinase

DOI:  https://doi.org/10.1113/JP284982
Cell Rep. 2023 Nov 23. pii: S2211-1247(23)01484-5. [Epub ahead of print]42(12): 113472

Mitochondrial nucleoid condensates drive peripheral fission through high membrane curvature.

Qixin Chen, Liu-Yi Liu, Zhiqi Tian, Zhou Fang, Kang-Nan Wang, Xintian Shao, Chengying Zhang, Weiwei Zou, Fiona Rowan, Kangqiang Qiu, Baohua Ji, Jun-Lin Guan, Dechang Li, Zong-Wan Mao, Jiajie Diao.

  Mitochondria are dynamic organelles that undergo fusion and fission events, in which the mitochondrial membrane and DNA (mtDNA) play critical roles. The spatiotemporal organization of mtDNA reflects and impacts mitochondrial dynamics. Herein, to study the detailed dynamics of mitochondrial membrane and mtDNA, we rationally develop a dual-color fluorescent probe, mtGLP, that could be used for simultaneously monitoring mitochondrial membrane and mtDNA dynamics via separate color outputs. By combining mtGLP with structured illumination microscopy to monitor mitochondrial dynamics, we discover the formation of nucleoid condensates in damaged mitochondria. We further reveal that nucleoid condensates promoted the peripheral fission of damaged mitochondria via asymmetric segregation. Through simulations, we find that the peripheral fission events occurred when the nucleoid condensates interacted with the highly curved membrane regions at the two ends of the mitochondria. Overall, we show that mitochondrial nucleoid condensates utilize peripheral fission to maintain mitochondrial homeostasis.

Keywords:  CP: Cell biology; chemical biology

DOI:  https://doi.org/10.1016/j.celrep.2023.113472
Nat Metab. 2023 Nov;5(11): 1840-1843

A century of the Warburg effect.

Craig B Thompson, Karen H Vousden, Randall S Johnson, Willem H Koppenol, Helmut Sies, Zhimin Lu, Lydia W S Finley, Christian Frezza, Jiyeon Kim, Zeping Hu, Caroline R Bartman.

DOI: https://doi.org/10.1038/s42255-023-00927-3
Nat Commun. 2023 11 18. 14(1): 7525

Inferring mitochondrial and cytosolic metabolism by coupling isotope tracing and deconvolution.

Alon Stern, Mariam Fokra, Boris Sarvin, Ahmad Abed Alrahem, Won Dong Lee, Elina Aizenshtein, Nikita Sarvin, Tomer Shlomi.

The inability to inspect metabolic activities within distinct subcellular compartments has been a major barrier to our understanding of eukaryotic cell metabolism. Previous work addressed this challenge by analyzing metabolism in isolated organelles, which grossly bias metabolic activity. Here, we describe a method for inferring physiological metabolic fluxes and metabolite concentrations in mitochondria and cytosol based on isotope tracing experiments performed with intact cells. This is made possible by computational deconvolution of metabolite isotopic labeling patterns and concentrations into cytosolic and mitochondrial counterparts, coupled with metabolic and thermodynamic modelling. Our approach lowers the uncertainty regarding compartmentalized fluxes and concentrations by one and three orders of magnitude compared to existing modelling approaches, respectively. We derive a quantitative view of mitochondrial and cytosolic metabolic activities in central carbon metabolism across cultured cell lines without performing cell fractionation, finding major variability in compartmentalized malate-aspartate shuttle fluxes. We expect our approach for inferring metabolism at a subcellular resolution to be instrumental for a variety of studies of metabolic dysfunction in human disease and for bioengineering.

DOI: https://doi.org/10.1038/s41467-023-42824-z
Cancer Discov. 2023 Nov 22. OF1

Vitamin B5 Supports Oncogenic Metabolism in MYC-Driven Breast Cancer.

Pantothetic acid is required for metabolic activity that supports MYC-driven breast tumor growth.

DOI: https://doi.org/10.1158/2159-8290.CD-RW2023-185
Life Sci Alliance. 2024 Feb;pii: e202302147. [Epub ahead of print]7(2):

Mitochondrial phosphoproteomes are functionally specialized across tissues.

Fynn M Hansen, Laura S Kremer, Ozge Karayel, Isabell Bludau, Nils-Göran Larsson, Inge Kühl, Matthias Mann.

Mitochondria are essential organelles whose dysfunction causes human pathologies that often manifest in a tissue-specific manner. Accordingly, mitochondrial fitness depends on versatile proteomes specialized to meet diverse tissue-specific requirements. Increasing evidence suggests that phosphorylation may play an important role in regulating tissue-specific mitochondrial functions and pathophysiology. Building on recent advances in mass spectrometry (MS)-based proteomics, we here quantitatively profile mitochondrial tissue proteomes along with their matching phosphoproteomes. We isolated mitochondria from mouse heart, skeletal muscle, brown adipose tissue, kidney, liver, brain, and spleen by differential centrifugation followed by separation on Percoll gradients and performed high-resolution MS analysis of the proteomes and phosphoproteomes. This in-depth map substantially quantifies known and predicted mitochondrial proteins and provides a resource of core and tissue-specific mitochondrial proteins (mitophos.de). Predicting kinase substrate associations for different mitochondrial compartments indicates tissue-specific regulation at the phosphoproteome level. Illustrating the functional value of our resource, we reproduce mitochondrial phosphorylation events on dynamin-related protein 1 responsible for its mitochondrial recruitment and fission initiation and describe phosphorylation clusters on MIGA2 linked to mitochondrial fusion.

DOI: https://doi.org/10.26508/lsa.202302147
Genes (Basel). 2023 Nov 17. pii: 2092. [Epub ahead of print]14(11):

Comprehensive Identification of Mitochondrial Pseudogenes (NUMTs) in the Human Telomere-to-Telomere Reference Genome.

Yichen Tao, Chengpeng He, Deng Lin, Zhenglong Gu, Weilin Pu.

  Practices related to mitochondrial research have long been hindered by the presence of mitochondrial pseudogenes within the nuclear genome (NUMTs). Even though partially assembled human reference genomes like hg38 have included NUMTs compilation, the exhaustive NUMTs within the only complete reference genome (T2T-CHR13) remain unknown. Here, we comprehensively identified the fixed NUMTs within the reference genome using human pan-mitogenome (HPMT) from GeneBank. The inclusion of HPMT serves the purpose of establishing an authentic mitochondrial DNA (mtDNA) mutational spectrum for the identification of NUMTs, distinguishing it from the polymorphic variations found in NUMTs. Using HPMT, we identified approximately 10% of additional NUMTs in three human reference genomes under stricter thresholds. And we also observed an approximate 6% increase in NUMTs in T2T-CHR13 compared to hg38, including NUMTs on the short arms of chromosomes 13, 14, and 15 that were not assembled previously. Furthermore, alignments based on 20-mer from mtDNA suggested the presence of more mtDNA-like short segments within the nuclear genome, which should be avoided for short amplicon or cell free mtDNA detection. Finally, through the assay of transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) on cell lines before and after mtDNA elimination, we concluded that NUMTs have a minimal impact on bulk ATAC-seq, even though 16% of sequencing data originated from mtDNA.

Keywords:  ATAC-seq; NUMTs; human reference genome; mitochondrial DNA

DOI:  https://doi.org/10.3390/genes14112092
Biochem Pharmacol. 2023 Nov 22. pii: S0006-2952(23)00522-1. [Epub ahead of print] 115929

Adaptive changes in tumor cells in response to reductive stress.

Leilei Zhang, Jie Zhang, Zhi-Wei Ye, Aslam Muhammad, Li Li, John W Culpepper, Danyelle M Townsend, Kenneth D Tew.

  Reductive stress is characterized by an excess of cellular electron donors and can be linked with various human pathologies including cancer. We developed melanoma cell lines resistant to reductive stress agents: rotenone (ROTR), n-acetyl-L-cysteine, (NACR), or dithiothreitol (DTTR). Resistant cells divided more rapidly and had intracellular homeostatic redox-couple ratios that were shifted towards the reduced state. Resistance caused alterations in general cell morphology, but only ROTR cells had significant changes in mitochondrial morphology with higher numbers that were more isolated, fragmented and swollen, with greater membrane depolarization and decreased numbers of networks. These changes were accompanied by lower basal oxygen consumption and maximal respiration rates. Whole cell flux analyses and mitochondrial function assays showed that NACR and DTTR preferentially utilized tricarboxylic acid (TCA) cycle intermediates, while ROTR used ketone body substrates such as D, L-β-hydroxybutyric acid. NACR and DTTR cells had constitutively decreased levels of reactive oxygen species (ROS), although this was accompanied by activation of nuclear factor erythroid 2-related factor 2 (Nrf2), with concomitant increased expression of the downstream gene products such as glutathione S-transferase P (GSTP). Further adaptations included enhanced expression of endoplasmic reticulum proteins controlling the unfolded protein response (UPR). Although expression patterns of these UPR proteins were distinct between the resistant cells, a trend implied that resistance to reductive stress is accompanied by a constitutively increased UPR phenotype in each line. Overall, tumor cells, although tolerant of oxidative stress, can adapt their energy and survival mechanisms in lethal reductive stress conditions.

Keywords:  Glycolysis; Melanoma cell lines; Mitochondrial morphology; Reactive oxygen species; Reductive stress; Unfolded protein response

DOI:  https://doi.org/10.1016/j.bcp.2023.115929
Proc Natl Acad Sci U S A. 2023 Nov 28. 120(48): e2314043120

Proteome-wide tagging with an H2O2 biosensor reveals highly localized and dynamic redox microenvironments.

Paraskevi Kritsiligkou, Katharina Bosch, Tzu Keng Shen, Matthias Meurer, Michael Knop, Tobias P Dick.

  Hydrogen peroxide (H2O2) sensing and signaling involves the reversible oxidation of particular thiols on particular proteins to modulate protein function in a dynamic manner. H2O2 can be generated from various intracellular sources, but their identities and relative contributions are often unknown. To identify endogenous "hotspots" of H2O2 generation on the scale of individual proteins and protein complexes, we generated a yeast library in which the H2O2 sensor HyPer7 was fused to the C-terminus of all protein-coding open reading frames (ORFs). We also generated a control library in which a redox-insensitive mutant of HyPer7 (SypHer7) was fused to all ORFs. Both libraries were screened side-by-side to identify proteins located within H2O2-generating environments. Screening under a variety of different metabolic conditions revealed dynamic changes in H2O2 availability highly specific to individual proteins and protein complexes. These findings suggest that intracellular H2O2 generation is much more localized and functionally differentiated than previously recognized.

Keywords:  genetically encoded probes; hydrogen peroxide; redox regulation; redox signaling

DOI:  https://doi.org/10.1073/pnas.2314043120
Cancer Gene Ther. 2023 Nov 20.

Arvanil induces ferroptosis of hepatocellular carcinoma by binding to MICU1.

Xiangying Deng, Yajun Gui, Lin Zhao, Namei Li, Liling Li.

Hepatocellular carcinoma (HCC) is a primary liver cancer with a high mortality rate that requires research and improved treatment strategies. Chemotherapy is still one of the main methods of HCC treatment, but it may lead to drug resistance and damage to normal organs. Capsaicin, a naturally occurring active ingredient in chili peppers, has demonstrated anticancer properties in a variety of malignant tumor cell lines. However, the anti-cancer mechanism of capsaicin needs to be further explored in HCC. In this study, we utilized Arvanil, a non-stimulating synthetic capsaicin analog, in place of capsaicin. We found that Arvanil induced high mitochondrial calcium flow, which contributed to a decrease in mitochondrial membrane permeability transition pore (mPTP) opening and oxidative phosphorylation levels, ultimately triggering cellular ferroptosis by live cells in real time with a high content screening (HCS) platform and confocal microscopy. It was further confirmed by vina molecular docking and point mutation experiments that Arvanil directly binds to two amino acid sites of mitochondrial calcium uptake protein 1 (MICU1), namely Ser47 and Phe128, to trigger this process, which in turn inhibits the growth of HCC cells. In addition, it was confirmed that Arvanil enhances cisplatin chemosensitivity by inducing HCC cellular ferroptosis in vivo. In conclusion, our study suggests that Arvanil induces ferroptosis in HCC cells and is a candidate drug for the treatment of HCC.

DOI: https://doi.org/10.1038/s41417-023-00690-3
EMBO J. 2023 Nov 21. e114221

An alternative NURF complex sustains acute myeloid leukemia by regulating the accessibility of insulator regions.

Aliaksandra Radzisheuskaya, Isabel Peña-Rømer, Eugenia Lorenzini, Richard Koche, Yingqian Zhan, Pavel V Shliaha, Alexandra J Cooper, Zheng Fan, Daria Shlyueva, Jens V Johansen, Ronald C Hendrickson, Kristian Helin.

  Efficient treatment of acute myeloid leukemia (AML) patients remains a challenge despite recent therapeutic advances. Here, using a CRISPRi screen targeting chromatin factors, we identified the nucleosome-remodeling factor (NURF) subunit BPTF as an essential regulator of AML cell survival. We demonstrate that BPTF forms an alternative NURF chromatin remodeling complex with SMARCA5 and BAP18, which regulates the accessibility of a large set of insulator regions in leukemic cells. This ensures efficient CTCF binding and boundary formation between topologically associated domains that is essential for maintaining the leukemic transcriptional programs. We also demonstrate that the well-studied PHD2-BROMO chromatin reader domains of BPTF, while contributing to complex recruitment to chromatin, are dispensable for leukemic cell growth. Taken together, our results uncover how the alternative NURF complex contributes to leukemia and provide a rationale for its targeting in AML.

Keywords:  BPTF; SMARCA5; acute myeloid leukemia; chromatin remodeling; insulator regions

DOI:  https://doi.org/10.15252/embj.2023114221
Free Radic Biol Med. 2023 Nov 16. pii: S0891-5849(23)01101-2. [Epub ahead of print]

Mitochondrial hyperfusion induces metabolic remodeling in lung endothelial cells by modifying the activities of electron transport chain complexes I and III.

Manivannan Yegambaram, Xutong Sun, Qing Lu, Yan Jin, Wojciech Ornatowski, Jamie Soto, Saurabh Aggarwal, Ting Wang, Kim Tieu, Haiwei Gu, Jeffrey R Fineman, Stephen M Black.

  OBJECTIVE: Pulmonary hypertension (PH) is a progressive disease with vascular remodeling as a critical structural alteration. We have previously shown that metabolic reprogramming is an early initiating mechanism in animal models of PH. This metabolic dysregulation has been linked to remodeling the mitochondrial network to favor fission. However, whether the mitochondrial fission/fusion balance underlies the metabolic reprogramming found early in PH development is unknown.METHODS: Utilizing a rat early model of PH, in conjunction with cultured pulmonary endothelial cells (PECs), we utilized metabolic flux assays, Seahorse Bioassays, measurements of electron transport chain (ETC) complex activity, fluorescent microscopy, and molecular approaches to investigate the link between the disruption of mitochondrial dynamics and the early metabolic changes that occur in PH.
RESULTS: We observed increased fusion mediators, including Mfn1, Mfn2, and Opa1, and unchanged fission mediators, including Drp1 and Fis1, in a two-week monocrotaline-induced PH animal model (early-stage PH). We were able to establish a connection between increases in fusion mediator Mfn1 and metabolic reprogramming. Using an adenoviral expression system to enhance Mfn1 levels in pulmonary endothelial cells and utilizing 13C-glucose labeled substrate, we found increased production of 13C lactate and decreased TCA cycle metabolites, revealing a Warburg phenotype. The use of a 13C5-glutamine substrate showed evidence that hyperfusion also induces oxidative carboxylation. The increase in glycolysis was linked to increased hypoxia-inducible factor 1α (HIF-1α) protein levels secondary to the disruption of cellular bioenergetics and higher levels of mitochondrial reactive oxygen species (mt-ROS). The elevation in mt-ROS correlated with attenuated ETC complexes I and III activities. Utilizing a mitochondrial-targeted antioxidant to suppress mt-ROS, limited HIF-1α protein levels, which reduced cellular glycolysis and reestablished mitochondrial membrane potential.
CONCLUSIONS: Our data connects mitochondrial fusion-mediated mt-ROS to the Warburg phenotype in early-stage PH development.

Keywords:  Glycolysis; Metabolomics; Mitochondrial function; Mitofusin; Pulmonary hypertension

DOI:  https://doi.org/10.1016/j.freeradbiomed.2023.11.008
Mol Pharm. 2023 Nov 22.

Synthesis and Preclinical Evaluation of PSMA-Targeted 111In-Radioconjugates Containing a Mitochondria-Tropic Triphenylphosphonium Carrier.

Joana F Santos, Maria T Braz, Paula Raposinho, Frederik Cleeren, Irwin Cassells, Simon Leekens, Christopher Cawthorne, Filipa Mendes, Célia Fernandes, António Paulo.

  Nuclear DNA is the canonical target for biological damage induced by Auger electrons (AE) in the context of targeted radionuclide therapy (TRT) of cancer, but other subcellular components might also be relevant for this purpose, such as the energized mitochondria of tumor cells. Having this in mind, we have synthesized novel DOTA-based chelators carrying a prostate-specific membrane antigen (PSMA) inhibitor and a triphenyl phosphonium (TPP) group that were used to obtain dual-targeted 111In-radioconjugates ([111In]In-TPP-DOTAGA-PSMA and [111In]In-TPP-DOTAGA-G3-PSMA), aiming to promote a selective uptake of an AE-emitter radiometal (111In) by PSMA+ prostate cancer (PCa) cells and an enhanced accumulation in the mitochondria. These dual-targeted 111In-radiocomplexes are highly stable under physiological conditions and in cell culture media. The complexes showed relatively similar binding affinities toward the PSMA compared to the reference tracer [111In]In-PSMA-617, in line with their high cellular uptake and internalization in PSMA+ PCa cells. The complexes compromised cell survival in a dose-dependent manner and in the case of [111In]In-TPP-DOTAGA-G3-PSMA to a higher extent than observed for the single-targeted congener [111In]In-PSMA-617. μSPECT imaging studies in PSMA+ PCa xenografts showed that the TPP pharmacophore did not interfere with the excellent in vivo tumor uptake of the "golden standard" [111In]In-PSMA-617, although it led to a higher kidney retention. Such kidney retention does not necessarily compromise their usefulness as radiotherapeutics due to the short tissue range of the Auger/conversion electrons emitted by 111In. Overall, our results provide valuable insights into the potential use of mitochondrial targeting by PSMA-based radiocomplexes for efficient use of AE-emitting radionuclides in TRT, giving impetus to extend the studies to other AE-emitting trivalent radiometals (e.g., 161Tb or 165Er) and to further optimize the designed dual-targeting constructs.

Keywords:  Auger electron emitters; PSMA; cancer theranostics; dual-targeting; mitochondria; radiopharmaceuticals

DOI:  https://doi.org/10.1021/acs.molpharmaceut.3c00787
Molecules. 2023 Nov 08. pii: 7486. [Epub ahead of print]28(22):

Phosphorus Chemistry at the Roots of Bioenergetics: Ligand Permutation as the Molecular Basis of the Mechanism of ATP Synthesis/Hydrolysis by FOF1-ATP Synthase.

Sunil Nath.

  The integration of phosphorus chemistry with the mechanism of ATP synthesis/hydrolysis requires dynamical information during ATP turnover and catalysis. Oxygen exchange reactions occurring at β-catalytic sites of the FOF1-ATP synthase/F1-ATPase imprint a unique record of molecular events during the catalytic cycle of ATP synthesis/hydrolysis. They have been shown to provide valuable time-resolved information on enzyme catalysis during ATP synthesis and ATP hydrolysis. The present work conducts new experiments on oxygen exchange catalyzed by submitochondrial particles designed to (i) measure the relative rates of Pi-ATP, Pi-HOH, and ATP-HOH isotope exchanges; (ii) probe the effect of ADP removal on the extent of inhibition of the exchanges, and (iii) test their uncoupler sensitivity/resistance. The objectives have been realized based on new experiments on submitochondrial particles, which show that both the Pi-HOH and ATP-HOH exchanges occur at a considerably higher rate relative to the Pi-ATP exchange, an observation that cannot be explained by previous mechanisms. A unifying explanation of the kinetic data that rationalizes these observations is given. The experimental results in (ii) show that ADP removal does not inhibit the intermediate Pi-HOH exchange when ATP and submitochondrial particles are incubated, and that the nucleotide requirement of the intermediate Pi-HOH exchange is adequately met by ATP, but not by ADP. These results contradicts the central postulate in Boyer's binding change mechanism of reversible catalysis at a F1 catalytic site with Keq~1 that predicts an absolute requirement of ADP for the occurrence of the Pi-HOH exchange. The prominent intermediate Pi-HOH exchange occurring under hydrolytic conditions is shown to be best explained by Nath's torsional mechanism of energy transduction and ATP synthesis/hydrolysis, which postulates an essentially irreversible cleavage of ATP by mitochondria/particles, independent from a reversible formation of ATP from ADP and Pi. The explanation within the torsional mechanism is also shown to rationalize the relative insensitivity of the intermediate Pi-HOH exchange to uncouplers observed in the experiments in (iii) compared to the Pi-ATP and ATP-HOH exchanges. This is shown to lead to new concepts and perspectives based on ligand displacement/substitution and ligand permutation for the elucidation of the oxygen exchange reactions within the framework of fundamental phosphorus chemistry. Fast mechanisms that realize the rotation/twist, tilt, permutation and switch of ligands, as well as inversion at the γ-phosphorus synchronously and simultaneously and in a concerted manner, have been proposed, and their stereochemical consequences have been analyzed. These considerations take us beyond the binding change mechanism of ATP synthesis/hydrolysis in bioenergetics.

Keywords:  ATP synthase; ATP synthesis and ATP hydrolysis; Boyer’s binding change mechanism of ATP synthesis/hydrolysis; FOF1/F1-ATPase; Nath’s torsional mechanism of ATP synthesis/hydrolysis; bisite catalysis vs. trisite catalysis; kinetics and catalysis; ligand permutation and ligand displacement/substitution; molecular mechanism; oxygen exchange; phosphorus chemistry and biochemistry; rotary ATPases; two-site vs. three-site models of ATP synthesis/hydrolysis

DOI:  https://doi.org/10.3390/molecules28227486
Cell Rep Med. 2023 Nov 21. pii: S2666-3791(23)00484-6. [Epub ahead of print]4(11): 101290

Allosteric SHP2 inhibition increases apoptotic dependency on BCL2 and synergizes with venetoclax in FLT3- and KIT-mutant AML.

Bogdan Popescu, Carlos Stahlhut, Theodore C Tarver, Sydney Wishner, Bianca J Lee, Cheryl A C Peretz, Cuyler Luck, Paul Phojanakong, Juan Antonio Camara Serrano, Henry Hongo, Jose M Rivera, Simayijiang Xirenayi, John A Chukinas, Veronica Steri, Sarah K Tasian, Elliot Stieglitz, Catherine C Smith.

  Mutations in the receptor tyrosine kinases (RTKs) FLT3 and KIT are frequent and associated with poor outcomes in acute myeloid leukemia (AML). Although selective FLT3 inhibitors (FLT3i) are clinically effective, remissions are short-lived due to secondary resistance characterized by acquired mutations constitutively activating the RAS/MAPK pathway. Hereby, we report the pre-clinical efficacy of co-targeting SHP2, a critical node in MAPK signaling, and BCL2 in RTK-driven AML. The allosteric SHP2 inhibitor RMC-4550 suppresses proliferation of AML cell lines with FLT3 and KIT mutations, including cell lines with acquired resistance to FLT3i. We demonstrate that pharmacologic SHP2 inhibition unveils an Achilles' heel of RTK-driven AML, increasing apoptotic dependency on BCL2 via MAPK-dependent mechanisms, including upregulation of BMF and downregulation of MCL1. Consequently, RMC-4550 and venetoclax are synergistically lethal in AML cell lines and in clinically relevant xenograft models. Our results provide mechanistic rationale and pre-clinical evidence for co-targeting SHP2 and BCL2 in RTK-driven AML.

Keywords:  FLT3; SHP2; acute myeloid leukemia; apoptosis; cancer; drug synergy; targeted therapies

DOI:  https://doi.org/10.1016/j.xcrm.2023.101290
Nat Commun. 2023 Nov 24. 14(1): 7722

In vivo imaging of mitochondrial DNA mutations using an integrated nano Cas12a sensor.

Yanan Li, Yonghua Wu, Ru Xu, Jialing Guo, Fenglei Quan, Yongyuan Zhang, Di Huang, Yiran Pei, Hua Gao, Wei Liu, Junjie Liu, Zhenzhong Zhang, Ruijie Deng, Jinjin Shi, Kaixiang Zhang.

Mutations in mitochondrial DNA (mtDNA) play critical roles in many human diseases. In vivo visualization of cells bearing mtDNA mutations is important for resolving the complexity of these diseases, which remains challenging. Here we develop an integrated nano Cas12a sensor (InCasor) and show its utility for efficient imaging of mtDNA mutations in live cells and tumor-bearing mouse models. We co-deliver Cas12a/crRNA, fluorophore-quencher reporters and Mg2+ into mitochondria. This process enables the activation of Cas12a's trans-cleavage by targeting mtDNA, which efficiently cleave reporters to generate fluorescent signals for robustly sensing and reporting single-nucleotide variations (SNVs) in cells. Since engineered crRNA significantly increase Cas12a's sensitivity to mismatches in mtDNA, we can identify tumor tissue and metastases by visualizing cells with mutant mtDNAs in vivo using InCasor. This CRISPR imaging nanoprobe holds potential for applications in mtDNA mutation-related basic research, diagnostics and gene therapies.

DOI: https://doi.org/10.1038/s41467-023-43552-0
Anal Chem. 2023 Nov 21.

Exploring the Role of Mitochondrial Hydrogen Sulfide in Maintaining Polarity and mtDNA Integrity with a Multichannel Fluorescent Probe.

Shuai Mu, Taihe Han, Xiaoyun Zhang, Jia Liu, Huipeng Sun, Jinlong Zhang, Xiaoyan Liu, Haixia Zhang.

Abnormal mitochondrial state has been implicated in the pathogenesis of various diseases including neurodegenerative disorders, myopathies, cardiovascular diseases, and cancers. Assessing mitochondrial functionality can be achieved by monitoring alterations in mitochondrial polarity and mitochondrial DNA (mtDNA) integrity, which serve as valuable biomarkers. Hydrogen sulfide (H2S), a gaseous signaling molecule, plays a regulatory role in mitochondrial respiratory chain activity, ATP synthesis, and calcium ion balance, thereby influencing cellular metabolism and signal transduction. Investigating the interplay between mitochondrial H2S, polarity, and mtDNA can enhance our understanding of the underlying regulatory mechanisms involved in H2S-mediated mitochondrial functions. To address this, we designed a mitochondria-targeted multichannel fluorescent probe, HNA, capable of cascaded detection of H2S and polarity, as well as parallel detection of mtDNA. The probe exhibited a significant turn-on response to H2S, emitting at approximately 604 nm, while the product HNAP demonstrated high sensitivity to polarity within the wavelength range of 526-591 nm. Additionally, the probe was able to bind to DNA, resulting in an enhanced long-wave emission at 668 nm. Facilitated by HNA, our study provides novel insights into the role of mitochondrial H2S in maintaining mitochondrial polarity and validates its protective effect on mtDNA through antioxidative mechanisms. Overall, this work proposes a potential therapeutic strategy for modulating the inflammatory process in mitochondrial-related diseases.

DOI: https://doi.org/10.1021/acs.analchem.3c03663
Nat Commun. 2023 Nov 21. 14(1): 7572

SETD2 deficiency accelerates sphingomyelin accumulation and promotes the development of renal cancer.

Hanyu Rao, Changwei Liu, Aiting Wang, Chunxiao Ma, Yue Xu, Tianbao Ye, Wenqiong Su, Peijun Zhou, Wei-Qiang Gao, Li Li, Xianting Ding.

Patients with polycystic kidney disease (PKD) encounter a high risk of clear cell renal cell carcinoma (ccRCC), a malignant tumor with dysregulated lipid metabolism. SET domain-containing 2 (SETD2) has been identified as an important tumor suppressor and an immunosuppressor in ccRCC. However, the role of SETD2 in ccRCC generation in PKD remains largely unexplored. Herein, we perform metabolomics, lipidomics, transcriptomics and proteomics within SETD2 loss induced PKD-ccRCC transition mouse model. Our analyses show that SETD2 loss causes extensive metabolic reprogramming events that eventually results in enhanced sphingomyelin biosynthesis and tumorigenesis. Clinical ccRCC patient specimens further confirm the abnormal metabolic reprogramming and sphingomyelin accumulation. Tumor symptom caused by Setd2 knockout is relieved by myriocin, a selective inhibitor of serine-palmitoyl-transferase and sphingomyelin biosynthesis. Our results reveal that SETD2 deficiency promotes large-scale metabolic reprogramming and sphingomyelin biosynthesis during PKD-ccRCC transition. This study introduces high-quality multi-omics resources and uncovers a regulatory mechanism of SETD2 on lipid metabolism during tumorigenesis.

DOI: https://doi.org/10.1038/s41467-023-43378-w
Discov Oncol. 2023 Nov 19. 14(1): 207

A comprehensive analysis of SLC25A1 expression and its oncogenic role in pan-cancer.

Xin You, Lingling Huang, Ouxiang Huang, Yujie Deng, Xi Shi.

  OBJECTIVE: The solute carrier family 25 member 1 (SLC25A1) is currently the only known human transporter for citrate in the mitochondrial membrane. However, its role in cancer development remains to be elucidated. We aim to analyze the expression profile, prognostic value, potential immunological significance, and effect on tumor growth of SLC25A1 at a pan-cancer level.METHODS: Herein, the role of SLC25A1 in tumorigenesis and progression was investigated based on the Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), Genotype-Tissue Expression (GTEx), Clinical Proteomic Tumor Analysis Consortium (CPTAC), GeneMANIA, STRING and Cancer Dependency Map Project (DepMap) database via online websites or the R software. The protein expression levels were validated in tissue microarrays, and the effects on tumor cell lines were accessed through MTS and colony formation assays.
RESULTS: The expression of SLC25A1 increased in most cancers, and the upregulation of SLC25A1 in colon adenocarcinoma and lung adenocarcinoma was further confirmed by immunohistochemistry. Meanwhile, SLC25A1 was linked to clinical outcomes across multiple tumor types, particularly in lung adenocarcinoma, where its high expression predicted poor prognosis. Moreover, SLC25A1 was positively associated with MSI, TMB, and CD276 and tightly correlated with tumor-infiltrating immune cells. Furthermore, the knockout of SLC25A1 demonstrated inhibitory effects in most cancer cell lines in the DepMap project. Cellular experiments showed that SLC25A1 knockdown significantly reduced the proliferation of lung adenocarcinoma cells.
CONCLUSIONS: Our findings suggest the potential of SLC25A1 as a prognostic biomarker for cancers and a therapeutic target for precise antitumor strategy and cancer immunotherapy.

Keywords:  Pan-cancer; Prognosis; SLC25A1; Therapeutic target; Tumor immunity

DOI:  https://doi.org/10.1007/s12672-023-00830-z
Biochem Pharmacol. 2023 Nov 19. pii: S0006-2952(23)00527-0. [Epub ahead of print]218 115934

Hydroquinone-selected chronic myelogenous leukemia cells are sensitive to chloroquine-induced cytotoxicity via MCL1 suppression and glycolysis inhibition.

Jing-Ting Chiou, Yuan-Chin Lee, Long-Sen Chang.

  Previous studies have provided evidence that repeated exposure to the benzene metabolite hydroquinone (HQ) induces malignant transformation and increases basal autophagy in the chronic myeloid leukemia (CML) cell line K562. This study explored the cytotoxicity of the autophagy inhibitor chloroquine (CQ) on parental and HQ-selected K562 (K562/HQ) cells. CQ triggered apoptosis in these cells independently of inhibiting autophagic flux; however, in K562/HQ cells, CQ-induced cytotoxicity was higher than in K562 cells. Mechanistically, CQ-induced NOXA upregulation led to MCL1 downregulation and mitochondrial depolarization in K562/HQ cells. MCL1 overexpression or NOXA silencing attenuated CQ-mediated cytotoxicity in K562/HQ cells. CQ triggered ERK inactivation to increase Sp1, NFκB, and p300 expression, and co-assembly of Sp1, NFκB, and p300 in the miR-29a promoter region coordinately upregulated miR-29a transcription. CQ-induced miR-29a expression destabilized tristetraprolin (TTP) mRNA, which in turn reduced TTP-mediated NOXA mRNA decay, thereby increasing NOXA protein expression. A similar mechanism explained the CQ-induced downregulation of MCL1 in K562 cells. K562/HQ cells relied more on glycolysis for ATP production than K562 cells, whereas inhibition of glycolysis by CQ was greater in K562/HQ cells than in K562 cells. Likewise, CQ-induced MCL1 suppression and glycolysis inhibition resulted in higher cytotoxicity in CML KU812/HQ cells than in KU812 cells. Taken together, our data confirm that CQ inhibits MCL1 expression through the ERK/miR-29a/TTP/NOXA pathway, and that inhibition of glycolysis is positively correlated to higher cytotoxicity of CQ on HQ-selected CML cells.

Keywords:  Chloroquine; Glycolysis inhibition; Hydroquinone exposure; Leukemia; MCL1 downregulation

DOI:  https://doi.org/10.1016/j.bcp.2023.115934