bims-mibica 2022-06-05 papers

bims-mibica

Biomed News

on Mitochondrial bioenergetics in cancer

Issue of 2022‒06‒05
35 papers selected by
Kelsey Fisher-Wellman, East Carolina University

Mitochondrial matrix-localized Src kinase regulates mitochondrial morphology.
Inhibition of nuclear deacetylase Sirtuin-1 induces mitochondrial acetylation and calcium overload leading to cell death.
Diverse reaction behaviors of artificial ubiquinones in mitochondrial respiratory complex I.
The Mitochondrial Pyruvate Carrier at the Crossroads of Intermediary Metabolism.
Inhibition of B-cell lymphoma 2 family proteins alters optical redox ratio, mitochondrial polarization, and cell energetics independent of cell state.
REDD1 interacts with AIF and regulates mitochondrial reactive oxygen species generation in the keratinocyte response to UVB.
Fatty acid oxidation protects cancer cells from apoptosis by increasing mitochondrial membrane lipids.
High-intensity interval training remodels the proteome and acetylome of human skeletal muscle.
MAPK15 protects from oxidative stress-dependent cellular senescence by inducing the mitophagic process.
Mitochondrial phosphatidylethanolamine synthesis affects mitochondrial energy metabolism and quiescence entry through attenuation of Snf1/AMPK signaling in yeast.
Redox regulation by TXNRD3 during epididymal maturation underlies capacitation-associated mitochondrial activity and sperm motility in mice.
A mitochondrial unfolded protein response inhibitor suppresses prostate cancer growth in mice via HSP60.
PFKP alleviates glucose starvation-induced metabolic stress in lung cancer cells via AMPK-ACC2 dependent fatty acid oxidation.
LncRNA OVAAL enhances nucleotide synthesis via pyruvate carboxylase to promote 5-FU resistance in gastric cancer.
Neddylation inhibition induces glutamine uptake and metabolism by targeting CRL3SPOP E3 ligase in cancer cells.
PHYSICAL EXERCISE IMPROVES MITOCHONDRIAL FUNCTION IN OVARIECTOMIZED RATS.
Platinum-induced mitochondrial OXPHOS contributes to cancer stem cell enrichment in ovarian cancer.
Melatonin inhibits glycolysis in hepatocellular carcinoma cells by downregulating mitochondrial respiration and mTORC1 activity.
Ribosome stalling during selenoprotein translation exposes a ferroptosis vulnerability.
Comparison of structural properties of cyclosporin A and its analogue alisporivir and their effects on mitochondrial bioenergetics and membrane behavior.
Leukocyte cytokine responses in adult patients with mitochondrial DNA defects.
Reciprocal Regulation of Mitofusin 2-Mediated Mitophagy and Mitochondrial Fusion by Different PINK1 Phosphorylation Events.
Iron-Regulated Assembly of the Cytosolic Iron-Sulfur Cluster Biogenesis Machinery.
Quantitative analysis of redox proteome reveals oxidation-sensitive protein thiols acting in fundamental processes of developmental hematopoiesis.
TRPML1-induced autophagy inhibition triggers mitochondrial mediated apoptosis.
Mesaconate is synthesized from itaconate and exerts immunomodulatory effects in macrophages.
Intracellular acetyl CoA potentiates the therapeutic efficacy of anti-tumor CD8+ T cells.
Synthesis of a fluorinated pyronin that enables blue light to rapidly depolarize mitochondria.
Targeting mitochondrial DNA polymerase gamma for selective inhibition of MLH1 deficient colon cancer growth.
SREBF2-STARD4 axis confers sorafenib resistance in HCC by regulating mitochondrial cholesterol homeostasis.
The longitudinal dynamics and natural history of clonal haematopoiesis.
Low Adenylate Kinase 5 expression is predictive of poor prognosis and promotes tumor growth by regulating the cell cycle pathway.
A Metabolic Paradigm for H2S Signaling via ETC Plasticity.
A Genome-wide screen identifies PDPK1 as a target to enhance the efficacy of MEK1/2 inhibitors in NRAS mutant melanoma.
Clonal dynamics of haematopoiesis across the human lifespan.

Cell Mol Life Sci. 2022 May 30. 79(6): 327

Mitochondrial matrix-localized Src kinase regulates mitochondrial morphology.

Olivier Lurette, Hala Guedouari, Jordan L Morris, Rebeca Martín-Jiménez, Julie-Pier Robichaud, Geneviève Hamel-Côté, Mehtab Khan, Nicholas Dauphinee, Nicolas Pichaud, Julien Prudent, Etienne Hebert-Chatelain.

  The architecture of mitochondria adapts to physiological contexts: while mitochondrial fragmentation is usually associated to quality control and cell death, mitochondrial elongation often enhances cell survival during stress. Understanding how these events are regulated is important to elucidate how mitochondrial dynamics control cell fate. Here, we show that the tyrosine kinase Src regulates mitochondrial morphology. Deletion of Src increased mitochondrial size and reduced cellular respiration independently of mitochondrial mass, mitochondrial membrane potential or ATP levels. Re-expression of Src targeted to the mitochondrial matrix, but not of Src targeted to the plasma membrane, rescued mitochondrial morphology in a kinase activity-dependent manner. These findings highlight a novel function for Src in the control of mitochondrial dynamics.

Keywords:  Cellular respiration; Mitochondria-shaping protein; Mitochondrial dynamics; Oxidative phosphorylation

DOI:  https://doi.org/10.1007/s00018-022-04325-y
Redox Biol. 2022 May 19. pii: S2213-2317(22)00106-9. [Epub ahead of print]53 102334

Inhibition of nuclear deacetylase Sirtuin-1 induces mitochondrial acetylation and calcium overload leading to cell death.

Yue Sun, Yan-Ming Yang, Yu-Yu Hu, Lan Ouyang, Zheng-Hua Sun, Xing-Feng Yin, Nan Li, Qing-Yu He, Yang Wang.

  Sirtuin-1 (SIRT1) is a critical nuclear deacetylase that participates in a wide range of biological processes. We hereby employed quantitative acetyl-proteomics to globally reveal the landscape of SIRT1-dependent acetylation in colorectal cancer (CRC) cells stimulated by specific SIRT1 inhibitor Inauhzin (INZ). We strikingly observed that SIRT1 inhibition enhances protein acetylation levels, with the multisite-acetylated proteins (acetyl sites >4/protein) mainly enriched in mitochondria. INZ treatment increases mitochondrial fission and depolarization in CRC cells. The acetylation of mitochondrial proteins promoted by SIRT1 inhibition prevents the recruitment of ubiquitin and LC3 for mitophagic degradation. We then found that, SIRT1 inhibition increases the acetylation of mitochondrial calcium uniporter (MCU) at residue K332, resulting in mitochondrial Ca2+ overload and depolarization, and ultimately CRC apoptosis. Arginine substitution of the K332 (K332R) dramatically decreases the mitochondrial Ca2+ influx, mitochondrial membrane potential loss and ROS burst induced by INZ. This finding uncovers a non-canonical role of SIRT1 in regulating mitochondrial function and implicates a possible way for anticancer intervention through SIRT1 inhibition.

Keywords:  Acetylation; Calcium; Inauhzin; MCU; Mitochondria; SIRT1

DOI:  https://doi.org/10.1016/j.redox.2022.102334
J Biol Chem. 2022 May 25. pii: S0021-9258(22)00515-4. [Epub ahead of print] 102075

Diverse reaction behaviors of artificial ubiquinones in mitochondrial respiratory complex I.

Shinpei Uno, Takahiro Masuya, Oleksii Zdorevskyi, Ryou Ikunishi, Kyoko Shinzawa-Itoh, Jonathan Lasham, Vivek Sharma, Masatoshi Murai, Hideto Miyoshi.

  The ubiquinone (UQ) reduction step catalyzed by NADH-UQ oxidoreductase (mitochondrial respiratory complex I) is key to triggering proton translocation across the inner mitochondrial membrane (IMM). Structural studies have identified a long, narrow, UQ-accessing tunnel within the enzyme. We previously demonstrated that synthetic oversized UQs, which are unlikely to transit this narrow tunnel, are catalytically reduced by native complex I embedded in submitochondrial particles, but not by the isolated enzyme. To explain this contradiction, we hypothesized that access of oversized UQs to the reaction site is obstructed in the isolated enzyme because their access route is altered following detergent solubilization from the IMM. In the present study, we investigated this using two pairs of photoreactive UQs (pUQm-1/pUQp-1 and pUQm-2/pUQp-2), with each pair having the same chemical properties except for a ∼1.0 Å difference in side chain-widths. Despite this subtle difference, reduction of the wider pUQs by the isolated complex was significantly slower than of the narrower pUQs, but both were similarly reduced by the native enzyme. In addition, photoaffinity labeling experiments using the four [125I]pUQs demonstrated that their side chains predominantly label the ND1 subunit with both enzymes, but at different regions around the tunnel. Finally, we show the suppressive effects of different types of inhibitors on the labeling significantly changed depending on [125I]pUQs used, indicating that [125I]pUQs and these inhibitors do not necessarily share a common binding cavity. Altogether, we conclude that the reaction behaviors of pUQs cannot be simply explained by the canonical UQ tunnel model.

Keywords:  bioenergetics; chemical biology; complex I; mitochondria; ubiquinone

DOI:  https://doi.org/10.1016/j.jbc.2022.102075
Am J Physiol Endocrinol Metab. 2022 May 30.

The Mitochondrial Pyruvate Carrier at the Crossroads of Intermediary Metabolism.

Nicole K H Yiew, Brian N Finck.

  Pyruvate metabolism, a central nexus of carbon homeostasis, is an evolutionarily-conserved process and aberrant pyruvate metabolism is associated with and contributes to numerous human metabolic disorders including diabetes, cancer, and heart disease. As a product of glycolysis, pyruvate is primarily generated in the cytosol before being transported into the mitochondrion for further metabolism. Pyruvate entry into the mitochondrial matrix is a critical step for efficient generation of reducing equivalents and ATP and for the biosynthesis of glucose, fatty acids, and amino acids from pyruvate. However, for many years the identity of the carrier protein(s) that transported pyruvate into the mitochondrial matrix remained a mystery. In 2012, the molecular-genetic identification of the mitochondrial pyruvate carrier (MPC), a heterodimeric complex composed of protein subunits MPC1 and MPC2, enabled studies that shed light on the many metabolic and physiologic processes regulated by pyruvate metabolism. A better understanding of the mechanisms regulating pyruvate transport and the processes affected by pyruvate metabolism may enable novel therapeutics to modulate mitochondrial pyruvate flux to treat a variety disorders. Herein, we review our current knowledge of the MPC, discuss recent advances in the understanding of mitochondrial pyruvate metabolism in various tissue and cell types, and address some of the outstanding questions relevant to this field.

Keywords:  adipose tissue; heart; liver; mitochondrion; pyruvate

DOI:  https://doi.org/10.1152/ajpendo.00074.2022
J Biomed Opt. 2022 May;27(5):

Inhibition of B-cell lymphoma 2 family proteins alters optical redox ratio, mitochondrial polarization, and cell energetics independent of cell state.

Amani A Gillette, Rebecca A DeStefanis, Stephanie L Pritzl, Dustin A Deming, Melissa C Skala.

  SIGNIFICANCE: The optical redox ratio (ORR) [autofluorescence intensity of the reduced form of nicotinamide adenine dinucleotide (phosphate) (NAD(P)H)/flavin adenine dinucleotide (FAD)] provides a label-free method to quantify cellular metabolism. However, it is unclear whether changes in the ORR with B-cell lymphoma 2 (Bcl-2) family protein inhibition are due to metabolic stress alone or compromised cell viability.AIM: Determine whether ABT-263 (navitoclax, Bcl-2 family inhibitor) changes the ORR due to changes in mitochondrial function that are independent of changes in cell viability.
APPROACH: SW48 colon cancer cells were used to investigate changes in ORR, mitochondrial membrane potential, oxygen consumption rates, and cell state (cell growth, viability, proliferation, apoptosis, autophagy, and senescence) with ABT-263, TAK-228 [sapanisertib, mammalian target of rapamycin complex 1/2 (mTORC 1/2) inhibitor], and their combination at 24 h.
RESULTS: Changes in the ORR with Bcl-2 inhibition are driven by increases in both NAD(P)H and FAD autofluorescence, corresponding with increased basal metabolic rate and increased mitochondrial polarization. ABT-263 treatment does not change cell viability or induce autophagy but does induce a senescent phenotype. The metabolic changes seen with ABT-263 treatment are mitigated by combination with mTORC1/2 inhibition.
CONCLUSIONS: The ORR is sensitive to increases in mitochondrial polarization, energetic state, and cell senescence, which can change independently from cell viability.

Keywords:  autofluorescence; flavin adenine dinucleotide; mitochondria; nicotinamide adenine dinucleotide hydrogen; optical redox ratio

DOI:  https://doi.org/10.1117/1.JBO.27.5.056505
Biochem Biophys Res Commun. 2022 May 20. pii: S0006-291X(22)00777-X. [Epub ahead of print]616 56-62

REDD1 interacts with AIF and regulates mitochondrial reactive oxygen species generation in the keratinocyte response to UVB.

Robert P Feehan, Catherine S Coleman, Shauna Ebanks, Charles H Lang, Lisa M Shantz.

  Non-melanoma skin cancer (NMSC) incidence is rising, especially in high-risk, immunocompromised groups such as organ transplant patients, who often develop numerous, aggressive cutaneous squamous cell carcinomas. Identifying the pathways that support NMSC development will result in new approaches for prevention and therapy. Our goal is to define the function of REDD1 (Regulated in DNA Damage and Development 1) in the UVB stress response. REDD1 is rapidly induced by a variety of stressors to repress mechanistic target of rapamycin complex I (mTORC1), and it has been reported that REDD1 loss causes dysfunctional mitochondria with increased reactive oxygen species (ROS) and impaired oxidative phosphorylation (OXPHOS). We now show that knockout of REDD1 in human keratinocytes sensitizes them to UVB-induced apoptosis in an mTORC1-independent manner and intensifies mitochondrial ROS generation. Upon REDD1 knockout, we observe reduced levels of apoptosis inducing factor (AIF), a mitochondrial intermembrane space NADH oxidase that is required for electron transport chain Complex I biogenesis. Further, we show that keratinocyte REDD1 interacts with both AIF and the mitochondrial import protein CHCHD4, a direct binding partner of AIF that ensures functional OXPHOS. Our results support the hypothesis that REDD1 is part of a mitochondrial complex that protects cells from UVB-induced ROS toxicity and suggest novel therapeutic targets for prevention and therapy of NMSC.

Keywords:  AIF; Apoptosis; Non-melanoma skin cancer; REDD1; UVB; mTOR

DOI:  https://doi.org/10.1016/j.bbrc.2022.05.066
Cell Rep. 2022 May 31. pii: S2211-1247(22)00645-3. [Epub ahead of print]39(9): 110870

Fatty acid oxidation protects cancer cells from apoptosis by increasing mitochondrial membrane lipids.

Yi-Jia Li, Johannes Francois Fahrmann, Maryam Aftabizadeh, Qianqian Zhao, Satyendra C Tripathi, Chunyan Zhang, Yuan Yuan, David Ann, Samir Hanash, Hua Yu.

  Overcoming resistance to chemotherapies remains a major unmet need for cancers, such as triple-negative breast cancer (TNBC). Therefore, mechanistic studies to provide insight for drug development are urgently needed to overcome TNBC therapy resistance. Recently, an important role of fatty acid β-oxidation (FAO) in chemoresistance has been shown. But how FAO might mitigate tumor cell apoptosis by chemotherapy is unclear. Here, we show that elevated FAO activates STAT3 by acetylation via elevated acetyl-coenzyme A (CoA). Acetylated STAT3 upregulates expression of long-chain acyl-CoA synthetase 4 (ACSL4), resulting in increased phospholipid synthesis. Elevating phospholipids in mitochondrial membranes leads to heightened mitochondrial integrity, which in turn overcomes chemotherapy-induced tumor cell apoptosis. Conversely, in both cultured tumor cells and xenograft tumors, enhanced cancer cell apoptosis by inhibiting ASCL4 or specifically targeting acetylated-STAT3 is associated with a reduction in phospholipids within mitochondrial membranes. This study demonstrates a critical mechanism underlying tumor cell chemoresistance.

Keywords:  ACSL; CP: Cancer; CP: Metabolism; STAT3 acetylation; anti-apoptosis; chemoresistance; fatty acid oxidation; mitochondrial membrane potential; phospholipids

DOI:  https://doi.org/10.1016/j.celrep.2022.110870
Elife. 2022 May 31. pii: e69802. [Epub ahead of print]11

High-intensity interval training remodels the proteome and acetylome of human skeletal muscle.

Morten Hostrup, Anders Krogh Lemminger, Ben Stocks, Alba Gonzalez-Franquesa, Jeppe Kjærgaard Larsen, Julia Prats Quesada, Martin Thomassen, Brian Tate Weinert, Jens Bangsbo, Atul Shahaji Deshmukh.

  Exercise is an effective strategy in the prevention and treatment of metabolic diseases. Alterations in the skeletal muscle proteome, including post-translational modifications, regulate its metabolic adaptations to exercise. Here, we examined the effect of high-intensity interval training (HIIT) on the proteome and acetylome of human skeletal muscle, revealing the response of 3168 proteins and 1263 lysine acetyl-sites on 464 acetylated proteins. We identified global protein adaptations to exercise training involved in metabolism, excitation-contraction coupling, and myofibrillar calcium sensitivity. Furthermore, HIIT increased the acetylation of mitochondrial proteins, particularly those of complex V. We also highlight the regulation of exercise-responsive histone acetyl-sites. These data demonstrate the plasticity of the skeletal muscle proteome and acetylome, providing insight into the regulation of contractile, metabolic and transcriptional processes within skeletal muscle. Herein, we provide a substantial hypothesis-generating resource to stimulate further mechanistic research investigating how exercise improves metabolic health.

Keywords:  acetylation; biochemistry; calcium sensitivity; chemical biology; computational biology; exercise; human; mitochondria; proteomics; skeletal muscle; systems biology

DOI:  https://doi.org/10.7554/eLife.69802
Aging Cell. 2022 Jun 01. e13620

MAPK15 protects from oxidative stress-dependent cellular senescence by inducing the mitophagic process.

Lorenzo Franci, Alessandro Tubita, Franca Maria Bertolino, Alessandro Palma, Giuseppe Cannino, Carmine Settembre, Andrea Rasola, Elisabetta Rovida, Mario Chiariello.

  Mitochondria are the major source of reactive oxygen species (ROS), whose aberrant production by dysfunctional mitochondria leads to oxidative stress, thus contributing to aging as well as neurodegenerative disorders and cancer. Cells efficiently eliminate damaged mitochondria through a selective type of autophagy, named mitophagy. Here, we demonstrate the involvement of the atypical MAP kinase family member MAPK15 in cellular senescence, by preserving mitochondrial quality, thanks to its ability to control mitophagy and, therefore, prevent oxidative stress. We indeed demonstrate that reduced MAPK15 expression strongly decreases mitochondrial respiration and ATP production, while increasing mitochondrial ROS levels. We show that MAPK15 controls the mitophagic process by stimulating ULK1-dependent PRKN Ser108 phosphorylation and inducing the recruitment of damaged mitochondria to autophagosomal and lysosomal compartments, thus leading to a reduction of their mass, but also by participating in the reorganization of the mitochondrial network that usually anticipates their disposal. Consequently, MAPK15-dependent mitophagy protects cells from accumulating nuclear DNA damage due to mitochondrial ROS and, consequently, from senescence deriving from this chronic DNA insult. Indeed, we ultimately demonstrate that MAPK15 protects primary human airway epithelial cells from senescence, establishing a new specific role for MAPK15 in controlling mitochondrial fitness by efficient disposal of old and damaged organelles and suggesting this kinase as a new potential therapeutic target in diverse age-associated human diseases.

Keywords:  MAP kinases; Oxidative DNA damage; autophagy; cellular senescence; mitophagy; signal transduction

DOI:  https://doi.org/10.1111/acel.13620
FASEB J. 2022 Jul;36(7): e22355

Mitochondrial phosphatidylethanolamine synthesis affects mitochondrial energy metabolism and quiescence entry through attenuation of Snf1/AMPK signaling in yeast.

Non Miyata, Takanori Ito, Miyu Nakashima, Satoru Fujii, Osamu Kuge.

  The Ups2-Mdm35 complex mediates intramitochondrial phosphatidylserine (PS) transport to facilitate mitochondrial phosphatidylethanolamine (PE) synthesis. In the present study, we found that ups2∆ yeast showed increased mitochondrial ATP production and enhanced quiescence (G0) entry in the post-diauxic shift phase. Transcriptomic and biochemical analyses revealed that the depletion of Ups2 leads to overactivation of the yeast AMPK homolog Snf1. Inactivation of Snf1 by depletion of an Snf1-activating kinase, Sak1 canceled the changes in mitochondrial ATP production and quiescence entry observed in ups2∆ cells. Furthermore, among the factors regulated by Snf1, upregulation of pyruvate carboxylase, Pyc1 and downregulation of acetyl-CoA carboxylase, Acc1, respectively, were sufficient to increase mitochondrial ATP production and quiescence entry. These results suggested that a normal PE synthesis mediated by Ups2-Mdm35 complex attenuates Snf1/AMPK activity, and that Snf1-mediated regulation of carbon metabolisms has great impacts on mitochondrial energy metabolism and quiescence entry. We also found that depletion of Ups2 together with the cell-cycle regulators Whi5 and Whi7, functional orthologs of the Rb1 tumor suppressor, caused a synthetic growth defect in yeast. Similarly, knockdown of PRELID3b, the human homolog of Ups2, decreased the viability of Rb1-deficient breast cancer cells, suggesting that PRELID3b is a potential target for cancer therapy.

Keywords:  Snf1/AMPK; Ups2-Mdm35 complex; carbon metabolism; mitochondrial energy metabolism; phosphatidylethanolamine; quiescence entry

DOI:  https://doi.org/10.1096/fj.202101600RR
J Biol Chem. 2022 May 25. pii: S0021-9258(22)00517-8. [Epub ahead of print] 102077

Redox regulation by TXNRD3 during epididymal maturation underlies capacitation-associated mitochondrial activity and sperm motility in mice.

Huafeng Wang, Qianhui Dou, Kyung Jo Jeong, Jungmin Choi, Vadim N Gladyshev, Jean-Ju Chung.

  During epididymal transit, redox remodeling protects mammalian spermatozoa, preparing them for survival in the subsequent journey to fertilization. However, molecular mechanisms of redox regulation in sperm development and maturation remain largely elusive. In this study, we report that TXNRD3, a thioredoxin reductase family member particularly abundant in elongating spermatids at the site of mitochondrial sheath formation, regulates redox homeostasis to support male fertility. Using Txnrd3-/- mice, our biochemical, ultrastructural, and live cell imaging analyses revealed impairments in sperm morphology and motility under conditions of TXNRD3 deficiency. We find that mitochondria develop more defined cristae during capacitation in wild type sperm. Furthermore, we show that absence of TXNRD3 alters thiol redox status in both the head and tail during sperm maturation and capacitation, resulting in defective mitochondrial ultrastructure and activity under capacitating conditions. These findings provide insights into molecular mechanisms of redox homeostasis and bioenergetics during sperm maturation, capacitation, and fertilization.

Keywords:  Redox homeostasis; TXNRD3; Thioredoxin Glutathione Reductase; Ultrastructure; male fertility; mitochondrial function

DOI:  https://doi.org/10.1016/j.jbc.2022.102077
J Clin Invest. 2022 Jun 02. pii: e149906. [Epub ahead of print]

A mitochondrial unfolded protein response inhibitor suppresses prostate cancer growth in mice via HSP60.

Rahul Kumar, Ajay Kumar Chaudhary, Jordan Woytash, Joseph R Inigo, Abhiram A Gokhale, Wiam Bshara, Kristopher Attwood, Jianmin Wang, Joseph A Spernyak, Eva Rath, Neelu Yadav, Dirk Haller, David W Goodrich, Dean G Tang, Dhyan Chandra.

  Mitochondrial proteostasis, regulated by the mitochondrial unfolded protein response (UPRmt), is crucial for maintenance of cellular functions and survival. Elevated oxidative and proteotoxic stress in mitochondria must be attenuated by the activation of ubiquitous UPRmt to promote prostate cancer (PCa) growth. Here we show that the two key components of the UPRmt, heat shock protein 60 (HSP60, a mitochondrial chaperonin) and caseinolytic protease (ClpP, a mitochondrial protease) were required for the development of advanced PCa. HSP60 regulated ClpP expression via c-Myc and physically interacted with ClpP to restore mitochondrial functions promoting cancer cell survival. HSP60 maintained the ATP-producing functions of mitochondria, which activated β-catenin pathway leading to the upregulation of c-Myc. We identified an UPRmt inhibitor that blocked HSP60 interaction with ClpP and abrogated survival signaling without altering HSP60 chaperonin function. Disruption of HSP60-ClpP interaction by UPRmt inhibitor triggered metabolic stress and impeded PCa promoting signaling. Treatment with UPRmt inhibitor, or genetic ablation of Hsp60, inhibited PCa growth and progression. Together, our findings identify that HSP60-ClpP mediated UPRmt is essential for prostate tumorigenesis and HSP60-ClpP interaction represents a therapeutic vulnerability in PCa.

Keywords:  Cell Biology; Cell stress; Mitochondria; Oncology; Prostate cancer

DOI:  https://doi.org/10.1172/JCI149906
Cell Discov. 2022 May 31. 8(1): 52

PFKP alleviates glucose starvation-induced metabolic stress in lung cancer cells via AMPK-ACC2 dependent fatty acid oxidation.

Jiaqing Chen, Li Zou, Guang Lu, Oleg Grinchuk, Lei Fang, Derrick Sek Tong Ong, Reshma Taneja, Choon-Nam Ong, Han-Ming Shen.

Cancer cells adopt metabolic reprogramming to promote cell survival under metabolic stress. A key regulator of cell metabolism is AMP-activated protein kinase (AMPK) which promotes catabolism while suppresses anabolism. However, the underlying mechanism of AMPK in handling metabolic stress in cancer remains to be fully understood. In this study, by performing a proteomics screening of AMPK-interacting proteins in non-small-cell lung cancer (NSCLC) cells, we discovered the platelet isoform of phosphofructokinase 1 (PFKP), a rate-limiting enzyme in glycolysis. Moreover, PFKP was found to be highly expressed in NSCLC patients associated with poor survival. We demonstrated that the interaction of PFKP and AMPK was greatly enhanced upon glucose starvation, a process regulated by PFKP-associated metabolites. Notably, the PFKP-AMPK interaction promoted mitochondrial recruitment of AMPK which subsequently phosphorylated acetyl-CoA carboxylase 2 (ACC2) to enhance long-chain fatty acid oxidation, a process helping maintenance of the energy and redox homeostasis and eventually promoting cancer cell survival under glucose starvation. Collectively, we revealed a critical non-glycolysis-related function of PFKP in regulating long-chain fatty acid oxidation via AMPK to alleviate glucose starvation-induced metabolic stress in NSCLC cells.

DOI: https://doi.org/10.1038/s41421-022-00406-1
Cancer Sci. 2022 Jun 03.

LncRNA OVAAL enhances nucleotide synthesis via pyruvate carboxylase to promote 5-FU resistance in gastric cancer.

Jia-Nan Tan, Sheng-Ning Zhou, Wei Zhang, Bin Yang, Guang-Yu Zhong, Jing Huang, Hai Hu, Fang-Hai Han, Man-Li Luo.

  5-fluorouracil (5-FU) is widely used in gastric cancer treatment, yet 5-FU resistance remains an important clinical challenge. Here we established a 5-lncRNA model to effectively assess the prognosis of gastric cancer patients, among which lncRNA OVAAL was markedly upregulated in gastric cancer and associated with poor prognosis and 5-FU resistance. In vitro and in vivo assays confirmed that OVAAL promoted proliferation and 5-FU resistance of gastric cancer cells. Mechanistically, OVAAL bound with pyruvate carboxylase (PC) and stabilized PC from HSC70/CHIP -mediated ubiquitination and degradation. OVAAL knockdown reduced intracellular levels of oxaloacetate and aspartate, and the subsequent pyrimidine synthesis, which could be rescued by PC overexpression. Moreover, OVAAL knockdown increased sensitivity to 5-FU treatment, which could be revered by PC overexpression or repletion of oxaloacetate, aspartate or uridine. OVAAL overexpression enhanced pyrimidine synthesis to promote proliferation and 5-FU resistance of gastric cancer cells, which could be abolished by PC knockdown. Thus, OVAAL promoted gastric cancer cell proliferation and induced 5-FU resistance by enhancing pyrimidine biosynthesis to antagonize 5-FU induced thymidylate synthase dysfunction. Targeting OVAAL-mediated nucleotide metabolic reprograming would be a promising strategy to overcome chemoresistance in gastric cancer.

Keywords:  5-FU resistance; gastric cancer; lncRNA OVAAL; pyrimidine metabolism; pyruvate carboxylase

DOI:  https://doi.org/10.1111/cas.15453
Nat Commun. 2022 May 31. 13(1): 3034

Neddylation inhibition induces glutamine uptake and metabolism by targeting CRL3SPOP E3 ligase in cancer cells.

Qiyin Zhou, Wenyu Lin, Chaoqun Wang, Fei Sun, Siwei Ju, Qian Chen, Yi Wang, Yongxia Chen, Haomin Li, Linbo Wang, Zeping Hu, Hongchuan Jin, Xian Wang, Yi Sun.

Abnormal neddylation activation is frequently observed in human cancers and neddylation inhibition has been proposed as a therapy for cancer. Here, we report that MLN4924, a small-molecule inhibitor of neddylation activating enzyme, increases glutamine uptake in breast cancer cells by causing accumulation of glutamine transporter ASCT2/SLC1A5, via inactivation of CRL3-SPOP E3 ligase. We show the E3 ligase SPOP promotes ASCT2 ubiquitylation, whereas SPOP itself is auto-ubiquitylated upon glutamine deprivation. Thus, SPOP and ASCT2 inversely regulate glutamine uptake and metabolism. SPOP knockdown increases ASCT2 levels to promote growth which is rescued by ASCT2 knockdown. Adding ASCT2 inhibitor V-9302 enhances MLN4924 suppression of tumor growth. In human breast cancer specimens, SPOP and ASCT2 levels are inversely correlated, whereas lower SPOP with higher ASCT2 predicts a worse patient survival. Collectively, our study links neddylation to glutamine metabolism via the SPOP-ASCT2 axis and provides a rational drug combination for enhanced cancer therapy.

DOI: https://doi.org/10.1038/s41467-022-30559-2
J Endocrinol. 2022 May 01. pii: JOE-22-0057. [Epub ahead of print]

PHYSICAL EXERCISE IMPROVES MITOCHONDRIAL FUNCTION IN OVARIECTOMIZED RATS.

Daniele Leão Ignacio, Rodrigo Soares Fortunato, Diego Silvestre, Leonardo Matta, Andressa Lima de Vansconcelos, Denise Pires Carvalho, Antonio Galina, João Pedro Werneck-de-Castro, João Paulo Cavalcanti-de-Albuquerque.

Estrogen deficiency causes metabolic disorders in humans and rodents, including in part due to changes in energy expenditure. We have shown previously that skeletal muscle mitochondrial function is compromised in ovariectomized rats. Since physical exercise is a powerful strategy to improve skeletal muscle mitochondrial content and function, we hypothesize that exercise training would counteract the deficiency-induced skeletal muscle mitochondrial dysfunction in ovariectomized rats. We report that exercised ovariectomized rats, at 60-65% of maximal exercise capacity for eight weeks, exhibited less fat accumulation and body-weight gain compared with sedentary controls. Treadmill exercise training decreased muscle lactate production, indicating a shift to mitochondrial oxidative metabolism. Furthermore, reduced soleus muscle mitochondrial oxygen consumption confirmed that estrogen deficiency is detrimental to mitochondrial function. However, exercise restored mitochondrial oxygen consumption in ovariectomized rats, achieving similar levels as in exercised control rats. Exercise-induced skeletal muscle peroxisome proliferator-activated receptor-γ coactivator-1α, expression was similar in both groups. Therefore, the mechanisms by which exercise improves mitochondrial oxygen consumption appears to be different in ovariectomized-exercised and sham-exercised rats. While there was an increase in mitochondrial content in sham-exercised rats, demonstrated by a greater citrate synthase activity, no induction was observed in ovariectomized-exercised rats. Normalizing mitochondrial respiratory capacity by citrate synthase activity indicates a better oxidative phosphorylation efficiency in the ovariectomized-exercised group. In conclusion, physical exercise sustains mitochondrial function in ovarian hormone-deficient rats through a non-conventional mitochondrial content-independent manner.

DOI: https://doi.org/10.1530/JOE-22-0057
J Transl Med. 2022 May 31. 20(1): 246

Platinum-induced mitochondrial OXPHOS contributes to cancer stem cell enrichment in ovarian cancer.

Shruthi Sriramkumar, Riddhi Sood, Thomas D Huntington, Ahmed H Ghobashi, Truc T Vuong, Tara X Metcalfe, Weini Wang, Kenneth P Nephew, Heather M O'Hagan.

  BACKGROUND: Platinum based agents-cisplatin and carboplatin in combination with taxanes are used for the treatment of ovarian cancer (OC) patients. However, the majority of OC patients develop recurrent, platinum resistant disease that is uniformly fatal. Platinum treatment enriches for chemoresistant aldehyde dehydrogenase (ALDH) + ovarian cancer stem cells (OCSCs), which contribute to tumor recurrence and disease relapse. Acquired platinum resistance also includes metabolic reprograming and switching to oxidative phosphorylation (OXPHOS). Chemosensitive cells rely on glycolysis while chemoresistant cells have the ability to switch between glycolysis and OXPHOS, depending on which pathway drives a selective advantage for growth and chemoresistance. High expression of genes involved in OXPHOS and high production of mitochondrial ROS are characteristics of OCSCs, suggesting that OCSCs favor OXPHOS over glycolysis. Based on connections between OCSCs, chemoresistance and OXPHOS, we hypothesize that platinum treatment induces changes in metabolism that contribute to platinum-induced enrichment of OCSCs.METHODS: The effect of cisplatin on mitochondrial activity was assessed by JC1 staining and expression of OXPHOS genes by RT-qPCR. Cisplatin-induced changes in Sirtuin 1 (SIRT1) levels and activity were assessed by western blot. Small molecule inhibitors of mitochondrial complex I and SIRT1 were used to determine if their enzymatic activity contributes to the platinum-induced enrichment of OCSCs. The percentage of ALDH + OCSCs in OC cells and tumor tissue from xenograft models across different treatment conditions was analyzed using ALDEFLUOR assay and flow cytometry.
RESULTS: We demonstrate that platinum treatment increases mitochondrial activity. Combined treatment of platinum agents and OXPHOS inhibitors blocks the platinum-induced enrichment of ALDH + OCSCs in vitro and in vivo. Furthermore, platinum treatment increases SIRT1 levels and subsequent deacetylase activity, which likely contributes to the increase in platinum-induced mitochondrial activity.
CONCLUSIONS: These findings on metabolic pathways altered by platinum-based chemotherapy have uncovered key targets that can be exploited therapeutically to block the platinum-induced enrichment of OCSCs, ultimately improving the survival of OC patients.

Keywords:  ALDH + cells; Cancer stem cells; Chemoresistance; OXPHOS; Ovarian cancer; Platinum; SIRT1

DOI:  https://doi.org/10.1186/s12967-022-03447-y
BMB Rep. 2022 Jun 02. pii: 5522. [Epub ahead of print]

Melatonin inhibits glycolysis in hepatocellular carcinoma cells by downregulating mitochondrial respiration and mTORC1 activity.

Seunghyeong Lee, Jun-Kyu Byun, Na-Young Kim, Jonghwa Jin, Hyein Woo, Yeon-Kyung Choi, Keun-Gyu Park.

Various mechanisms have been suggested to explain the chemopreventive and tumor-inhibitory effects of melatonin. Despite the growing evidence supporting melatonin-induced mitochondrial dysfunction, it remains largely unknown how this phenomenon modulates metabolic reprogramming in cancer cells. The aim of our study was to identify the mechanism underlying the anti-proliferative and apoptotic effects of melatonin, which is known to inhibit glycolysis. We analyzed the time-dependent effects of melatonin on mitochondrial respiration and glycolysis in liver cancer cells. The results showed that from a cell bioenergetic point of view, melatonin caused an acute reduction in mitochondrial respiration, however, increased reactive oxygen species production, thereby inhibiting mTORC1 activity from an early stage post-treatment without affecting glycolysis. Nevertheless, administration of melatonin for a longer time reduced expression of c-Myc protein, thereby suppressing glycolysis via downregulation of HK2 and LDHA. The data presented herein suggest that melatonin suppresses mitochondrial respiration and glycolysis simultaneously in HCC cells, leading to anti-cancer effects. Thus, melatonin can be used as an adjuvant agent for therapy of liver cancer.
Nat Chem Biol. 2022 May 30.

Ribosome stalling during selenoprotein translation exposes a ferroptosis vulnerability.

Zhipeng Li, Lucas Ferguson, Kirandeep K Deol, Melissa A Roberts, Leslie Magtanong, Joseph M Hendricks, Gergey Alzaem Mousa, Seda Kilinc, Kaitlin Schaefer, James A Wells, Michael C Bassik, Andrei Goga, Scott J Dixon, Nicholas T Ingolia, James A Olzmann.

The selenoprotein glutathione peroxidase 4 (GPX4) prevents ferroptosis by converting lipid peroxides into nontoxic lipid alcohols. GPX4 has emerged as a promising therapeutic target for cancer treatment, but some cancer cells are resistant to ferroptosis triggered by GPX4 inhibition. Using a chemical-genetic screen, we identify LRP8 (also known as ApoER2) as a ferroptosis resistance factor that is upregulated in cancer. Loss of LRP8 decreases cellular selenium levels and the expression of a subset of selenoproteins. Counter to the canonical hierarchical selenoprotein regulatory program, GPX4 levels are strongly reduced due to impaired translation. Mechanistically, low selenium levels result in ribosome stalling at the inefficiently decoded GPX4 selenocysteine UGA codon, leading to ribosome collisions, early translation termination and proteasomal clearance of the N-terminal GPX4 fragment. These findings reveal rewiring of the selenoprotein hierarchy in cancer cells and identify ribosome stalling and collisions during GPX4 translation as ferroptosis vulnerabilities in cancer.

DOI: https://doi.org/10.1038/s41589-022-01033-3
Biochim Biophys Acta Biomembr. 2022 May 25. pii: S0005-2736(22)00110-9. [Epub ahead of print]1864(9): 183972

Comparison of structural properties of cyclosporin A and its analogue alisporivir and their effects on mitochondrial bioenergetics and membrane behavior.

Mikhail V Dubinin, Vyacheslav A Sharapov, Anna I Ilzorkina, Sergey V Efimov, Vladimir V Klochkov, Sergey V Gudkov, Konstantin N Belosludtsev.

  The paper considers the effect of the MPT pore inhibitor cyclosporin A (CsA) and its non-immunosuppressive analogue alisporivir (Ali) on the functioning of rat skeletal muscle mitochondria. We have shown that both agents at a standard in vitro concentration of 1 μM increase the calcium capacity of organelles and have no effect on the parameters of oxidative phosphorylation. However, an increase in their concentration to 5 μM leads to the suppression of oxygen consumption by mitochondria, which is more pronounced in the case of Ali. This effect is accompanied by a decrease in the membrane potential of organelles and, apparently, is based on the inhibition of electron transport along the mitochondrial respiratory chain due to limited mobility of coenzyme Q. We have noted that both agents do not affect the production of hydrogen peroxide by isolated mitochondria. NMR spectroscopy and molecular dynamics simulation did not reveal significant differences in the structure and backbone flexibility of CsA and Ali. Both agents decrease the overall fluidity of the membrane of DPPC liposomes, inducing an increase in laurdan generalized polarization parameter. A similar effect was also found in the case of mitochondrial membranes. We suggested that these effects of CsA and Ali, associated with their lipophilic nature and the ability to accumulate in the lipid phase of membranes, may cause a decrease in the efficiency of electron transport in the respiratory chain of mitochondria and suppression of the bioenergetics of these organelles.

Keywords:  Alisporivir; Cyclosporin a; Membrane fluidity; Molecular dynamics; NMR spectroscopy; Skeletal muscle mitochondria

DOI:  https://doi.org/10.1016/j.bbamem.2022.183972
J Mol Med (Berl). 2022 Jun;100(6): 963-971

Leukocyte cytokine responses in adult patients with mitochondrial DNA defects.

Kalpita R Karan, Caroline Trumpff, Marissa Cross, Kristin M Engelstad, Anna L Marsland, Peter J McGuire, Michio Hirano, Martin Picard.

  Patients with oxidative phosphorylation (OxPhos) defects causing mitochondrial diseases appear particularly vulnerable to infections. Although OxPhos defects modulate cytokine production in vitro and in animal models, little is known about how circulating leukocytes of patients with inherited mitochondrial DNA (mtDNA) defects respond to acute immune challenges. In a small cohort of healthy controls (n = 21) and patients (n = 12) with either the m.3243A > G mutation or single, large-scale mtDNA deletions, we examined (i) cytokine responses (IL-6, TNF-α, IL-1β) in response to acute lipopolysaccharide (LPS) exposure and (ii) sensitivity to the immunosuppressive effects of glucocorticoid signaling (dexamethasone) on cytokine production. In dose-response experiments to determine the half-maximal effective LPS concentration (EC50), relative to controls, leukocytes from patients with mtDNA deletions showed 74-79% lower responses for IL-6 and IL-1β (pIL-6 = 0.031, pIL-1β = 0.009). Moreover, whole blood from patients with mtDNA deletions (pIL-6 = 0.006), but not patients with the m.3243A > G mutation, showed greater sensitivity to the immunosuppressive effects of dexamethasone. Together, these ex vivo data provide preliminary evidence that some systemic OxPhos defects may compromise immune cytokine responses and increase the sensitivity to immune cytokine suppression by glucocorticoids. Further work in larger cohorts is needed to define the nature of immune dysregulation in patients with mitochondrial disease, and their potential implications for disease phenotypes. KEY MESSAGES: Little is known about leukocyte cytokine responses in patients with mitochondrial diseases. Leukocytes of patients with mtDNA deletions show blunted LPS sensitivity and cytokine responses. Leukocytes of patients with mtDNA deletions are more sensitive to glucocorticoid-mediated IL-6 suppression. Work in larger cohorts is needed to delineate potential immune alterations in mitochondrial diseases.

Keywords:  3243A > G; Cytokine; Glucocorticoid; Inflammation; Inflammation Suppression; Interleukin; Mitochondrial disease; mtDNA deletion

DOI:  https://doi.org/10.1007/s00109-022-02206-2
Front Cell Dev Biol. 2022 ;10 868465

Reciprocal Regulation of Mitofusin 2-Mediated Mitophagy and Mitochondrial Fusion by Different PINK1 Phosphorylation Events.

Jiajia Li, Xiawei Dang, Antonietta Franco, Gerald W Dorn.

  Mitochondrial repair is essential to metabolic homeostasis. Outer mitochondrial membrane mitofusin (MFN) proteins orchestrate mitochondrial fusion that opposes mitochondrial degeneration caused by senescence. Depending upon physiological context, MFN2 can either mediate mitochondrial fusion or recruit cytosolic Parkin to initiate mitophagic elimination. Because it is not clear how these events are counter-regulated we engineered and expressed MFN2 mutants that mimic phosphorylated or non-phosphorylatable MFN2 at its PINK1 phosphorylation sites: T111, S378, and S442. By interrogating mitochondrial fusion, polarization status, and Parkin binding/mitophagy as a function of inferred MFN2 phosphorylation, we discovered that individual MFN2 phosphorylation events act as a biological "bar-code", directing mitochondrial fate based on phosphorylation site state. Experiments in Pink1 deficient cells supported a central role for PINK1 kinase as the pivotal regulator of MFN2 functionality. Contrary to popular wisdom that Parkin-mediated ubiquitination regulates MFN-mediated mitochondrial fusion, results in Prkn null cells demonstrated the dispensability of Parkin for MFN2 inactivation. These data demonstrate that PINK1-mediated phosphorylation is necessary and sufficient, and that Parkin is expendable, to switch MFN2 from fusion protein to mitophagy effector.

Keywords:  MFN2; PINK1 kinase; Parkin; fusion; mitochondrial quality control; mitofusin regulation; phosphorylation

DOI:  https://doi.org/10.3389/fcell.2022.868465
J Biol Chem. 2022 May 30. pii: S0021-9258(22)00535-X. [Epub ahead of print] 102094

Iron-Regulated Assembly of the Cytosolic Iron-Sulfur Cluster Biogenesis Machinery.

Xiaorui Fan, William D Barshop, Ajay A Vashisht, Vijaya Pandey, Stephanie Leal, Shima Rayatpisheh, Yasaman Jami-Alahmadi, Jihui Sha, James A Wohlschlegel.

  The cytosolic iron-sulfur (Fe-S) cluster assembly (CIA) pathway delivers Fe-S clusters to nuclear and cytosolic Fe-S proteins involved in essential cellular functions. Although the delivery process is regulated by the availability of iron and oxygen, it remains unclear how CIA components orchestrate the cluster transfer under varying cellular environments. Here, we utilized a targeted proteomics assay for monitoring CIA factors and substrates to characterize the CIA machinery. We find that NUBP1 (NBP35), CIAO3 (NARFL) and CIA substrates associate with NUBP2 (CFD1), a component of the CIA scaffold complex. We also show that NUBP2 weakly associates with the CIA targeting complex (MMS19, CIAO1, CIAO2B) indicating the possible existence of a higher order complex. Interactions between CIAO3 and the CIA scaffold complex are strengthened upon iron supplementation or low oxygen tension, while iron chelation and reactive oxygen species weaken CIAO3 interactions with CIA components. We further demonstrate that CIAO3 mutants defective in Fe-S cluster binding fail to integrate into the higher order complexes. However, these mutants exhibit stronger associations with CIA substrates under conditions in which the association with the CIA targeting complex is reduced suggesting that CIAO3 and CIA substrates may associate in complexes independently of the CIA targeting complex. Together, our data suggest that CIA components potentially form a metabolon whose assembly is regulated by environmental cues and requires Fe-S cluster incorporation in CIAO3. These findings provide additional evidence that the CIA pathway adapts to changes in cellular environment through complex reorganization.

Keywords:  Iron-sulfur protein; cytosolic iron-sulfur cluster assembly (CIA); metalloprotein; protein-protein interaction; proteomics; redox regulation

DOI:  https://doi.org/10.1016/j.jbc.2022.102094
Redox Biol. 2022 May 23. pii: S2213-2317(22)00115-X. [Epub ahead of print]53 102343

Quantitative analysis of redox proteome reveals oxidation-sensitive protein thiols acting in fundamental processes of developmental hematopoiesis.

K Pimkova, M Jassinskaja, R Munita, M Ciesla, N Guzzi, P Cao Thi Ngoc, M Vajrychova, E Johansson, C Bellodi, J Hansson.

  Fetal and adult hematopoietic stem and progenitor cells (HSPCs) are characterized by distinct redox homeostasis that may influence their differential cellular behavior in normal and malignant hematopoiesis. In this work, we have applied a quantitative mass spectrometry-based redox proteomic approach to comprehensively describe reversible cysteine modifications in primary mouse fetal and adult HSPCs. We defined the redox state of 4,438 cysteines in fetal and adult HSPCs and demonstrated a higher susceptibility to oxidation of protein thiols in fetal HSPCs. Our data identified ontogenic changes to oxidation state of thiols in proteins with a pronounced role in metabolism and protein homeostasis. Additional redox proteomic analysis identified oxidation changes to thiols acting in mitochondrial respiration as well as protein homeostasis to be triggered during onset of MLL-ENL leukemogenesis in fetal HSPCs. Our data has demonstrated that redox signaling contributes to the regulation of fundamental processes of developmental hematopoiesis and has pinpointed potential targetable redox-sensitive proteins in in utero-initiated MLL-rearranged leukemia.

Keywords:  Cysteine oxidative modifications; Developmental biology; Hematopoiesis; Leukemia; Protein translation; Redox proteomics

DOI:  https://doi.org/10.1016/j.redox.2022.102343
Cancer Lett. 2022 May 26. pii: S0304-3835(22)00236-1. [Epub ahead of print]541 215752

TRPML1-induced autophagy inhibition triggers mitochondrial mediated apoptosis.

Yucheng Liu, Xinyan Wang, Wucheng Zhu, Zhongheng Sui, Xiangqing Wei, Yang Zhang, Jiansong Qi, Yanhong Xing, Wuyang Wang.

  Previous studies have demonstrated that autophagy tightly regulates apoptosis. However, the underlying mechanism whereby autophagy regulates apoptosis remains unclear. Here, we discover a "autophagy inhibition-mitochondrial turnover disruption-ROS elevation-DNA damage-p53 transactivation-apoptosis" axis that explicates the process of autophagy modulating apoptosis. We found that autophagy inhibition induced by TRPML1, a cationic channel localized in the lysosome, results in accumulation of damaged mitochondria via blocking the mitophagic flux to lysosomes in human melanoma and glioblastoma cells. The disrupted mitochondria turnover leads to ROS elevation, which in turn causes severe damage to DNA in these cancer cells. Damage to DNA resulted from TRPML1-mediated autophagy inhibition subsequently activates p53, which ultimately triggers mitochondrial mediated apoptosis by modulating pro- and anti-apoptosis proteins in these cancer cells. As a result, by triggering apoptosis, TRPML1-induced autophagy inhibition greatly suppresses growth of human melanoma and glioma both in vitro and in vivo. In summary, our findings define the mechanism underling the regulation of autophagy inhibition in apoptosis and represent TRPML1 as a novel target for potentially treating melanoma and glioblastoma in the clinical setting.

Keywords:  DNA damage; Glioblastoma; Lysosomes; Melanoma; p53

DOI:  https://doi.org/10.1016/j.canlet.2022.215752
Nat Metab. 2022 Jun 02.

Mesaconate is synthesized from itaconate and exerts immunomodulatory effects in macrophages.

Wei He, Antonia Henne, Mario Lauterbach, Eike Geißmar, Fabian Nikolka, Celia Kho, Alexander Heinz, Catherine Dostert, Melanie Grusdat, Thekla Cordes, Janika Härm, Oliver Goldmann, Anouk Ewen, Charlène Verschueren, Julia Blay-Cadanet, Robert Geffers, Hendrikus Garritsen, Manfred Kneiling, Christian K Holm, Christian M Metallo, Eva Medina, Zeinab Abdullah, Eicke Latz, Dirk Brenner, Karsten Hiller.

Since its discovery in inflammatory macrophages, itaconate has attracted much attention due to its antimicrobial and immunomodulatory activity1-3. However, instead of investigating itaconate itself, most studies used derivatized forms of itaconate and thus the role of non-derivatized itaconate needs to be scrutinized. Mesaconate, a metabolite structurally very close to itaconate, has never been implicated in mammalian cells. Here we show that mesaconate is synthesized in inflammatory macrophages from itaconate. We find that both, non-derivatized itaconate and mesaconate dampen the glycolytic activity to a similar extent, whereas only itaconate is able to repress tricarboxylic acid cycle activity and cellular respiration. In contrast to itaconate, mesaconate does not inhibit succinate dehydrogenase. Despite their distinct impact on metabolism, both metabolites exert similar immunomodulatory effects in pro-inflammatory macrophages, specifically a reduction of interleukin (IL)-6 and IL-12 secretion and an increase of CXCL10 production in a manner that is independent of NRF2 and ATF3. We show that a treatment with neither mesaconate nor itaconate impairs IL-1β secretion and inflammasome activation. In summary, our results identify mesaconate as an immunomodulatory metabolite in macrophages, which interferes to a lesser extent with cellular metabolism than itaconate.

DOI: https://doi.org/10.1038/s42255-022-00565-1
Cancer Res. 2022 May 31. pii: canres.4052.2021. [Epub ahead of print]

Intracellular acetyl CoA potentiates the therapeutic efficacy of anti-tumor CD8+ T cells.

Snehanshu Chowdhury, Anwesha Kar, Debaleena Bhowmik, Anupam Gautam, Debashree Basak, Ishita Sarkar, Puspendu Ghosh, Deborpita Sarkar, Alvina Deka, Paramita Chakraborty, Asima Mukhopadhyay, Shikhar Mehrotra, Soumen Basak, Sandip Paul, Shilpak Chatterjee.

Effector CD8+ T cells rely primarily on glucose metabolism to meet their biosynthetic and functional needs. However, nutritional limitations in the tumor microenvironment can cause T cell hyporesponsiveness. Therefore, T cells must acquire metabolic traits enabling sustained effector function at the tumor site to elicit a robust anti-tumor immune response. Here, we report that IL-12-stimulated CD8+ T cells have elevated intracellular acetyl CoA levels and can maintain IFNγ levels in nutrient-deprived, tumour-conditioned media (TCM). Pharmacological and metabolic analyses demonstrated an active glucose-citrate-acetyl CoA circuit in IL-12-stimulated CD8+ T cells supporting an intracellular pool of acetyl CoA in an ATP-citrate lyase (ACLY)-dependent manner. Intracellular acetyl CoA levels enhanced histone acetylation, lipid synthesis, and IFNγ production, improving the metabolic and functional fitness of CD8+ T cells in tumors. Pharmacological inhibition or genetic knockdown of ACLY severely impaired IFNγ production and viability of CD8+ T cells in nutrient-restricted conditions. Furthermore, CD8+ T cells cultured in high pyruvate-containing media in vitro acquired critical metabolic features of IL-12-stimulated CD8+ T cells and displayed improved anti-tumor potential upon adoptive transfer in murine lymphoma and melanoma models. Overall, this study delineates the metabolic configuration of CD8+ T cells required for stable effector function in tumors and presents an affordable approach to promote the efficacy of CD8+ T cells for adoptive T cell therapy.

DOI: https://doi.org/10.1158/0008-5472.CAN-21-4052
RSC Med Chem. 2022 Apr 20. 13(4): 456-462

Synthesis of a fluorinated pyronin that enables blue light to rapidly depolarize mitochondria.

Zhe Gao, Krishna K Sharma, Angelo E Andres, Brandon Walls, Fadel Boumelhem, Zachary R Woydziak, Blake R Peterson.

Fluorinated analogues of the fluorophore pyronin B were synthesized as a new class of amine-reactive drug-like small molecules. In water, 2,7-difluoropyronin B was found to reversibly react with primary amines to form covalent adducts. When this fluorinated analogue is added to proteins, these adducts undergo additional oxidation to yield fluorescent 9-aminopyronins. Irradiation with visible blue light enhances this oxidation step, providing a photochemical method to modify the biological properties of reactive amines. In living HeLa cells, 2,7-difluoropyronin B becomes localized in mitochondria, where it is partially transformed into fluorescent aminopyronins, as detected by spectral profiling confocal microscopy. Further excitation of these cells with the blue laser of a confocal microscope can depolarize mitochondria within seconds. This biological activity was only observed with 2,7-difluoropyronin B and was not detected with analogues such as pyronin B or 9-methyl-2,7-difluoropyronin B. This irradiation with blue light enhances the cellular production of reactive oxygen species (ROS), suggesting that increased ROS in mitochondria promotes the formation of aminopyronins that inactivate biomolecules critical for maintenance of mitochondrial membrane potential. The unique reactivity of 2,7-difluoropyronin B offers a novel tool for photochemical control of mitochondrial biology.

DOI: https://doi.org/10.1039/d1md00395j
PLoS One. 2022 ;17(6): e0268391

Targeting mitochondrial DNA polymerase gamma for selective inhibition of MLH1 deficient colon cancer growth.

Berna Somuncu, Aysegul Ekmekcioglu, Fatma Merve Antmen, Tugce Ertuzun, Emre Deniz, Nazli Keskin, Joon Park, Ilgu Ece Yazici, Busra Simsek, Batu Erman, Whitney Yin, Burak Erman, Meltem Muftuoglu.

Synthetic lethality in DNA repair pathways is an important strategy for the selective treatment of cancer cells without harming healthy cells and developing cancer-specific drugs. The synthetic lethal interaction between the mismatch repair (MMR) protein, MutL homolog 1 (MLH1), and the mitochondrial base excision repair protein, DNA polymerase γ (Pol γ) was used in this study for the selective treatment of MLH1 deficient cancers. Germline mutations in the MLH1 gene and aberrant MLH1 promoter methylation result in an increased risk of developing many cancers, including nonpolyposis colorectal and endometrial cancers. Because the inhibition of Pol γ in MLH1 deficient cancer cells provides the synthetic lethal selectivity, we conducted a comprehensive small molecule screening from various databases and chemical drug library molecules for novel Pol γ inhibitors that selectively kill MLH1 deficient cancer cells. We characterized these Pol γ inhibitor molecules in vitro and in vivo, and identified 3,3'-[(1,1'-Biphenyl)-4',4'-diyl)bis(azo)]bis[4-amino-1-naphthalenesulfonic acid] (congo red; CR; Zinc 03830554) as a high-affinity binder to the Pol γ protein and potent inhibitor of the Pol γ strand displacement and one-nucleotide incorporation DNA synthesis activities in vitro and in vivo. CR reduced the cell proliferation of MLH1 deficient HCT116 human colon cancer cells and suppressed HCT116 xenograft tumor growth whereas it did not affect the MLH1 proficient cell proliferation and xenograft tumor growth. CR caused mitochondrial dysfunction and cell death by inhibiting Pol γ activity and oxidative mtDNA damage repair, increasing the production of reactive oxygen species and oxidative mtDNA damage in MLH1 deficient cells. This study suggests that the Pol γ inhibitor, CR may be further evaluated for the MLH1 deficient cancers' therapy.

DOI: https://doi.org/10.1371/journal.pone.0268391
Cancer Sci. 2022 May 31.

SREBF2-STARD4 axis confers sorafenib resistance in HCC by regulating mitochondrial cholesterol homeostasis.

Xuetian Yue, Youzi Kong, Yankun Zhang, Min Sun, Shuyue Liu, Zhuanchang Wu, Lifen Gao, Xiaohong Liang, Chunhong Ma.

  Sorafenib resistance limits its survival benefit for treatment of HCC. Cholesterol metabolism is dysregulated in HCC, while its role in sorafenib resistance of HCC has not been fully elucidated. Aiming to elucidate this, in vitro and in vivo sorafenib resistant models were established. SREBF2, the key regulator of cholesterol metabolism, was activated in sorafenib resistant HepG2 and Huh7 cells. And, knockdown of SREBF2 re-sensitized sorafenib resistant cells and xenografts tumors to sorafenib. Further study showed that SREBF2 positively correlated with STARD4 (StAR related lipid transfer domain containing 4) in our sorafenib resistant models and publically available datasets. STARD4, mediating cholesterol trafficking, not only promoted proliferation and migration of HepG2 and Huh7 cells, but also increased sorafenib resistance in liver cancer. Mechanically, SREBF2 promoted expression of STARD4 by directly binding to its promoter region, leading to increase mitochondrial cholesterol levels and inhibit mitochondrial cytochrome c release. Importantly, knockdown of SREBF2 or STARD4 decreased mitochondrial cholesterol levels and increased mitochondrial cytochrome c release, respectively. Moreover, overexpression of STARD4 reversed effect of SREBF2 knockdown on mitochondrial cytochrome c release and sorafenib resistance. In conclusion, SREBF2 promotes STARD4 transcription, which in turn contributes to mitochondrial cholesterol transport and sorafenib resistance in HCC. Therefore, targeting on SREBF2-STARD4 axis would be beneficial to a subset of HCC patients with sorafenib resistance.

Keywords:  SREBF2; StARs; cholesterol transport; mitochondrial membrane permeability; sorafenib resistance

DOI:  https://doi.org/10.1111/cas.15449
Nature. 2022 Jun 01.

The longitudinal dynamics and natural history of clonal haematopoiesis.

Margarete A Fabre, José Guilherme de Almeida, Edoardo Fiorillo, Emily Mitchell, Aristi Damaskou, Justyna Rak, Valeria Orrù, Michele Marongiu, Michael Spencer Chapman, M S Vijayabaskar, Joanna Baxter, Claire Hardy, Federico Abascal, Nicholas Williams, Jyoti Nangalia, Iñigo Martincorena, Peter J Campbell, Eoin F McKinney, Francesco Cucca, Moritz Gerstung, George S Vassiliou.

Clonal expansions driven by somatic mutations become pervasive across human tissues with age, including in the haematopoietic system, where the phenomenon is termed clonal haematopoiesis1-4. The understanding of how and when clonal haematopoiesis develops, the factors that govern its behaviour, how it interacts with ageing and how these variables relate to malignant progression remains limited5,6. Here we track 697 clonal haematopoiesis clones from 385 individuals 55 years of age or older over a median of 13 years. We find that 92.4% of clones expanded at a stable exponential rate over the study period, with different mutations driving substantially different growth rates, ranging from 5% (DNMT3A and TP53) to more than 50% per year (SRSF2P95H). Growth rates of clones with the same mutation differed by approximately ±5% per year, proportionately affecting slow drivers more substantially. By combining our time-series data with phylogenetic analysis of 1,731 whole-genome sequences of haematopoietic colonies from 7 individuals from an older age group, we reveal distinct patterns of lifelong clonal behaviour. DNMT3A-mutant clones preferentially expanded early in life and displayed slower growth in old age, in the context of an increasingly competitive oligoclonal landscape. By contrast, splicing gene mutations drove expansion only later in life, whereas TET2-mutant clones emerged across all ages. Finally, we show that mutations driving faster clonal growth carry a higher risk of malignant progression. Our findings characterize the lifelong natural history of clonal haematopoiesis and give fundamental insights into the interactions between somatic mutation, ageing and clonal selection.

DOI: https://doi.org/10.1038/s41586-022-04785-z
Clin Exp Pharmacol Physiol. 2022 May 31.

Low Adenylate Kinase 5 expression is predictive of poor prognosis and promotes tumor growth by regulating the cell cycle pathway.

Pengfei Zhang, Yan Li, Yankui Liu, Lihua Zhang, Dong Hua.

  Colon adenocarcinoma (COAD) is one of the most common malignant tumors of the digestive system. Specific molecular markers play important role in COAD diagnosis and therapy. Adenylate Kinase 5 (AK5) is an enzyme that is related to energy metabolism and cancer. However, the exact role of AK5 in the progression of COAD is still unclear. In this study, the expression of AK5 in tissue samples and non-cancerous tissues of COAD patients was assessed by the bioinformatics method and western blot. Kaplan-Meier survival analysis and Cox regression analysis evaluated the prognostic significance of AK5. The biological function of AK5 in tumor progression was assessed by MTT assay, colony formation assay, transwell assay, wound healing assay, western blot, and mice xenograft models. The results showed that AK5 expression in tumor tissues was lower than in non-cancerous tissues. Notably, the patients with high AK5 expression possessed a longer overall survival (OS) than the low expression patients. And low AK5 expression promoted proliferation and metastasis in COAD cells by regulating the cell cycle pathway. Importantly, in vivo results showed that reduced AK5 expression is required for tumor growth. This study confirmed the significant role of AK5 in the development and progression of COAD. Therefore, low AK5 expression levels can be an independent prognostic biomarker, which provides new sight for the clinical diagnosis and target therapy of COAD. This article is protected by copyright. All rights reserved.

Keywords:  adenylate Kinase 5; colon adenocarcinoma; prognosis

DOI:  https://doi.org/10.1111/1440-1681.13680
Antioxid Redox Signal. 2022 Jun 01.

A Metabolic Paradigm for H2S Signaling via ETC Plasticity.

David Hanna, Roshan Kumar, Ruma Banerjee.

SIGNIFICANCE: A burgeoning literature has attributed varied physiological effects to H2S, which is a product of eukaryotic sulfur amino acid metabolism. Protein persulfidation represents a major focus of studies elucidating the mechanism underlying H2S signaling. On the other hand, the capacity of H2S to induce reductive stress by targeting the electron transport chain (ETC), and signal by reprogramming redox metabolism have only recently begun to be elucidated.RECENT ADVANCES: In contrast to the nonspecific reaction of H2S with oxidized cysteines to form protein persulfides, its inhibition of complex IV represents a specific mechanism of action. Studies on the dual impact of H2S as an ETC substrate and an inhibitor have led to the exciting discovery of ETC plasticity and the use of fumarate as a terminal electron acceptor. H2S oxidation combined with complex IV targeting generate mitochondrial reductive stress, which is signaled through the metabolic network, leading to increased aerobic glycolysis, glutamine-dependent reductive carboxylation and lipogenesis.
CRITICAL ISSUES: Insights into H2S-induced metabolic reprogramming are ushering in a paradigm shift for understanding the mechanism of its cellular action. It will be critical to reevaluate the physiological effects of H2S e.g., cytoprotection against ischemia-reperfusion injury, through the framework of metabolic reprogramming and ETC remodeling by H2S.
FUTURE DIRECTIONS: The metabolic ramifications of H2S in other cellular compartments, e.g., the endoplasmic reticulum and the nucleus, as well as the intersections between hypoxia and H2S signaling are important future directions that merit elucidation.

DOI: https://doi.org/10.1089/ars.2022.0067
Cancer Res. 2022 Jun 03. pii: canres.3217.2021-9-22 14:53:05.437. [Epub ahead of print]

A Genome-wide screen identifies PDPK1 as a target to enhance the efficacy of MEK1/2 inhibitors in NRAS mutant melanoma.

Weijia Cai, Mai Q Nguyen, Nicole A Wilski, Timothy J Purwin, Megane Vernon, Manoela Tiago, Andrew E Aplin.

Melanomas frequently harbor activating NRAS mutations. However, limited advance has been made in developing targeted therapy options for NRAS mutant melanoma patients. MEK inhibitors (MEKi) show modest efficacy in the clinic and their actions need to be optimized. In this study, we performed a genome-wide CRISPR-Cas9-based screen and demonstrated that loss of Phosphoinositide-dependent kinase-1 (PDPK1) enhances the efficacy of MEKi. The synergistic effects of PDPK1 loss and MEKi was validated in NRAS mutant melanoma cell lines using pharmacological and molecular approaches. Combined PDPK1 inhibitors (PDPK1i) with MEKi suppressed NRAS mutant xenograft growth and induced gasdermin E-associated pyroptosis. In an immune-competent allograft model, PDPK1i+MEKi increased the ratio of intratumoral CD8+ T cells, delayed tumor growth and prolonged survival; the combination treatment was less effective against tumors in immune-deficient mice. These data suggest PDPK1i+MEKi as an efficient immunostimulatory strategy against NRAS mutant melanoma.

DOI: https://doi.org/10.1158/0008-5472.CAN-21-3217
Nature. 2022 Jun 01.

Clonal dynamics of haematopoiesis across the human lifespan.

Emily Mitchell, Michael Spencer Chapman, Nicholas Williams, Kevin J Dawson, Nicole Mende, Emily F Calderbank, Hyunchul Jung, Thomas Mitchell, Tim H H Coorens, David H Spencer, Heather Machado, Henry Lee-Six, Megan Davies, Daniel Hayler, Margarete A Fabre, Krishnaa Mahbubani, Federico Abascal, Alex Cagan, George S Vassiliou, Joanna Baxter, Inigo Martincorena, Michael R Stratton, David G Kent, Krishna Chatterjee, Kourosh Saeb Parsy, Anthony R Green, Jyoti Nangalia, Elisa Laurenti, Peter J Campbell.

Age-related change in human haematopoiesis causes reduced regenerative capacity1, cytopenias2, immune dysfunction3 and increased risk of blood cancer4-6, but the reason for such abrupt functional decline after 70 years of age remains unclear. Here we sequenced 3,579 genomes from single cell-derived colonies of haematopoietic cells across 10 human subjects from 0 to 81 years of age. Haematopoietic stem cells or multipotent progenitors (HSC/MPPs) accumulated a mean of 17 mutations per year after birth and lost 30 base pairs per year of telomere length. Haematopoiesis in adults less than 65 years of age was massively polyclonal, with high clonal diversity and a stable population of 20,000-200,000 HSC/MPPs contributing evenly to blood production. By contrast, haematopoiesis in individuals aged over 75 showed profoundly decreased clonal diversity. In each of the older subjects, 30-60% of haematopoiesis was accounted for by 12-18 independent clones, each contributing 1-34% of blood production. Most clones had begun their expansion before the subject was 40 years old, but only 22% had known driver mutations. Genome-wide selection analysis estimated that between 1 in 34 and 1 in 12 non-synonymous mutations were drivers, accruing at constant rates throughout life, affecting more genes than identified in blood cancers. Loss of the Y chromosome conferred selective benefits in males. Simulations of haematopoiesis, with constant stem cell population size and constant acquisition of driver mutations conferring moderate fitness benefits, entirely explained the abrupt change in clonal structure in the elderly. Rapidly decreasing clonal diversity is a universal feature of haematopoiesis in aged humans, underpinned by pervasive positive selection acting on many more genes than currently identified.

DOI: https://doi.org/10.1038/s41586-022-04786-y