bims-mimbat 2023-04-16 papers

bims-mimbat

Biomed News

on Mitochondrial metabolism in brown adipose tissue

Issue of 2023–04–16
nine papers selected by
José Carlos de Lima-Júnior, Washington University

SLC25A22 as a key mitochondrial transporter against ferroptosis by producing GSH and MUFAs.
Synergy of uncoupling proteins (1 and 2) with mitochondrial Ca2+ uptake machinery potentiate mitochondrial uncoupling.
Generation of Brown Fat-Specific Knockout Mice Using a Combined Cre-LoxP, CRISPR-Cas9, and Adeno-Associated Virus Single-Guide RNA System.
Evidence supporting the MICU1 occlusion mechanism and against the potentiation model in the mitochondrial calcium uniporter complex.
The anorectic and thermogenic effects of pharmacological lactate in male mice are confounded by treatment osmolarity and co-administered counterions.
Circulating blood eNAMPT drives the circadian rhythms in locomotor activity and energy expenditure.
Defective import of mitochondrial metabolic enzyme elicits ectopic metabolic stress.
Interaction between poly-A binding protein PABPC4 and nuclear receptor corepressor NCoR1 modulates a metabolic stress response.
Cryo-EM structures of mitochondrial ABC transporter ABCB10 in apo and biliverdin-bound form.

Antioxid Redox Signal. 2023 Apr 13.

SLC25A22 as a key mitochondrial transporter against ferroptosis by producing GSH and MUFAs.

Yang Liu, Yuan Wang, Zhi Lin, Rui Kang, Doalin Tang, Jiao Liu.

AIMS: Ferroptosis, a type of oxidative cell death driven by unlimited lipid peroxidation, is emerging as a target for cancer therapy. Although mitochondrial dysfunction may lead to ferroptosis, the underlying molecular mechanisms and metabolic pathways for ferroptosis are incompletely understood. Here, we identify solute carrier family 25 member 22 (SLC25A22), a mitochondrial glutamate transporter, as a driver of ferroptosis resistance in pancreatic ductal adenocarcinoma (PDAC) cells.
RESULTS: The downregulation of SLC25A22 expression was associated with increased sensitivity to ferroptosis, but not to apoptosis. Mechanistically, on the one hand, SLC25A22-dependent NAPDH synthesis blocks ferroptotic cell death in PDAC cells through mediating the product of glutathione, the most important hydrophilic antioxidant. On the other hand, SLC25A22 promotes the expression of stearoyl-CoA desaturase (SCD) in PDAC cells in an AMP-activated protein kinase (AMPK)-dependent manner, resulting in the production of anti-ferroptotic monounsaturated fatty acids (MUFAs). The animal study further confirms that SLC25A22 inhibits ferroptosis-mediated tumor suppression.
INNOVATION: SLC25A22 is a novel metabolic repressor of ferroptosis by producing GSH and MUFAs.
CONCLUSION: These findings establish a previously unrecognized metabolic defense pathway to limit ferroptotic cell death in vitro and in vivo.

DOI: https://doi.org/10.1089/ars.2022.0203
Cell Calcium. 2023 Apr 05. pii: S0143-4160(23)00048-9. [Epub ahead of print]112 102736

Synergy of uncoupling proteins (1 and 2) with mitochondrial Ca2+ uptake machinery potentiate mitochondrial uncoupling.

Furkan E Oflaz, Zhanat Koshenov, Martin Hirtl, Olaf A Bachkoenig, Wolfgang F Graier, Benjamin Gottschalk.

  Mitochondrial uncoupling proteins UCP1 and UCP2 have a structural homology of app. 60%. They execute their mitochondria uncoupling function through different molecular mechanisms. Non-shivering thermogenesis by UCP1 is mediated through a transmembrane dissipation of the proton motive force to create heat during sympathetic stimulation. UCP2, on the other hand, modulates through the interaction with methylated MICU1 the permeability of the cristae junction, which acts as an isolator for the cristae-located mitochondrial membrane potential. In this mini-review, we discuss and compare the recently described molecular mechanism of UCP1 in brown adipose tissue and UCP2 in aged and cancer non-excitable cells that contribute to mitochondrial uncoupling, and the synergistic effects of both UCPs with the mitochondrial Ca2+ uptake machinery.

Keywords:  Mitochondrial Ca(2+) uptake machinery; Mitochondrial uncoupling; Uncoupling protein 1; Uncoupling protein 2

DOI:  https://doi.org/10.1016/j.ceca.2023.102736
J Vis Exp. 2023 Mar 24.

Generation of Brown Fat-Specific Knockout Mice Using a Combined Cre-LoxP, CRISPR-Cas9, and Adeno-Associated Virus Single-Guide RNA System.

Tadataka Tsuji, Yang Zhang, Yu-Hua Tseng.

Brown adipose tissue (BAT) is an adipose depot specialized in energy dissipation that can also serve as an endocrine organ via the secretion of bioactive molecules. The creation of BAT-specific knockout mice is one of the most popular approaches for understanding the contribution of a gene of interest to BAT-mediated energy regulation. The conventional gene targeting strategy utilizing the Cre-LoxP system has been the principal approach to generate tissue-specific knockout mice. However, this approach is time-consuming and tedious. Here, we describe a protocol for the rapid and efficient knockout of a gene of interest in BAT using a combined Cre-LoxP, CRISPR-Cas9, and adeno-associated virus (AAV) single-guide RNA (sgRNA) system. The interscapular BAT is located in the deep layer between the muscles. Thus, the BAT must be exposed in order to inject the AAV precisely and directly into the BAT within the visual field. Appropriate surgical handling is crucial to prevent damage to the sympathetic nerves and vessels, such as the Sultzer's vein that connects to the BAT. To minimize tissue damage, there is a critical need to understand the three-dimensional anatomical location of the BAT and the surgical skills required in the technical steps. This protocol highlights the key technical procedures, including the design of sgRNAs targeting the gene of interest, the preparation of AAV-sgRNA particles, and the surgery for the direct microinjection of AAV into both BAT lobes for generating BAT-specific knockout mice, which can be broadly applied to study the biological functions of genes in BAT.

DOI: https://doi.org/10.3791/65083
Proc Natl Acad Sci U S A. 2023 Apr 18. 120(16): e2217665120

Evidence supporting the MICU1 occlusion mechanism and against the potentiation model in the mitochondrial calcium uniporter complex.

Chen-Wei Tsai, Tsung-Yun Liu, Fan-Yi Chao, Yung-Chi Tu, Madison X Rodriguez, Anna M Van Keuren, Zhiwei Ma, John Bankston, Ming-Feng Tsai.

  The mitochondrial calcium uniporter is a Ca2+ channel that imports cytoplasmic Ca2+ into the mitochondrial matrix to regulate cell bioenergetics, intracellular Ca2+ signaling, and apoptosis. The uniporter contains the pore-forming MCU subunit, an auxiliary EMRE protein, and the regulatory MICU1/MICU2 subunits. Structural and biochemical studies have suggested that MICU1 gates MCU by blocking/unblocking the pore. However, mitoplast patch-clamp experiments argue that MICU1 does not block, but instead potentiates MCU via allosteric mechanisms. Here, we address this direct clash of the proposed MICU1 function. Supporting the MICU1-occlusion mechanism, patch-clamp demonstrates that purified MICU1 strongly suppresses MCU Ca2+ currents, and this inhibition is abolished by mutating the MCU-interacting K126 residue. Moreover, a membrane-depolarization assay shows that MICU1 prevents MCU-mediated Na+ flux into intact mitochondria under Ca2+-free conditions. Examining the observations underlying the potentiation model, we found that MICU1 occlusion was not detected in mitoplasts not because MICU1 cannot block, but because MICU1 dissociates from the uniporter complex. Furthermore, MICU1 depletion reduces uniporter transport not because MICU1 can potentiate MCU, but because EMRE is down-regulated. These results firmly establish the molecular mechanisms underlying the physiologically crucial process of uniporter regulation by MICU1.

Keywords:  calcium channels; intracellular calcium signaling; mitochondrial physiology; organellar channels

DOI:  https://doi.org/10.1073/pnas.2217665120
Nat Metab. 2023 Apr 13.

The anorectic and thermogenic effects of pharmacological lactate in male mice are confounded by treatment osmolarity and co-administered counterions.

Jens Lund, Alberte Wollesen Breum, Cláudia Gil, Sarah Falk, Frederike Sass, Marie Sophie Isidor, Oksana Dmytriyeva, Pablo Ranea-Robles, Cecilie Vad Mathiesen, Astrid Linde Basse, Olivia Sveidahl Johansen, Nicole Fadahunsi, Camilla Lund, Trine Sand Nicolaisen, Anders Bue Klein, Tao Ma, Brice Emanuelli, Maximilian Kleinert, Charlotte Mehlin Sørensen, Zachary Gerhart-Hines, Christoffer Clemmensen.

Lactate is a circulating metabolite and a signalling molecule with pleiotropic physiological effects. Studies suggest that lactate modulates energy balance by lowering food intake, inducing adipose browning and increasing whole-body thermogenesis. Yet, like many other metabolites, lactate is often commercially produced as a counterion-bound salt and typically administered in vivo through hypertonic aqueous solutions of sodium L-lactate. Most studies have not controlled for injection osmolarity and the co-injected sodium ions. Here, we show that the anorectic and thermogenic effects of exogenous sodium L-lactate in male mice are confounded by the hypertonicity of the injected solutions. Our data reveal that this is in contrast to the antiobesity effect of orally administered disodium succinate, which is uncoupled from these confounders. Further, our studies with other counterions indicate that counterions can have confounding effects beyond lactate pharmacology. Together, these findings underscore the importance of controlling for osmotic load and counterions in metabolite research.

DOI: https://doi.org/10.1038/s42255-023-00780-4
Nat Commun. 2023 Apr 08. 14(1): 1994

Circulating blood eNAMPT drives the circadian rhythms in locomotor activity and energy expenditure.

Jae Woo Park, Eun Roh, Gil Myoung Kang, So Young Gil, Hyun Kyong Kim, Chan Hee Lee, Won Hee Jang, Se Eun Park, Sang Yun Moon, Seong Jun Kim, So Yeon Jeong, Chae Beom Park, Hyo Sun Lim, Yu Rim Oh, Han Na Jung, Obin Kwon, Byung Soo Youn, Gi Hoon Son, Se Hee Min, Min-Seon Kim.

Nicotinamide adenine dinucleotide (NAD+) is an essential cofactor of critical enzymes including protein deacetylase sirtuins/SIRTs and its levels in mammalian cells rely on the nicotinamide phosphoribosyltransferase (NAMPT)-mediated salvage pathway. Intracellular NAMPT (iNAMPT) is secreted and found in the blood as extracellular NAMPT (eNAMPT). In the liver, the iNAMPT-NAD+ axis oscillates in a circadian manner and regulates the cellular clockwork. Here we show that the hypothalamic NAD+ levels show a distinct circadian fluctuation with a nocturnal rise in lean mice. This rhythm is in phase with that of plasma eNAMPT levels but not with that of hypothalamic iNAMPT levels. Chemical and genetic blockade of eNAMPT profoundly inhibit the nighttime elevations in hypothalamic NAD+ levels as well as those in locomotor activity (LMA) and energy expenditure (EE). Conversely, elevation of plasma eNAMPT by NAMPT administration increases hypothalamic NAD+ levels and stimulates LMA and EE via the hypothalamic NAD+-SIRT-FOXO1-melanocortin pathway. Notably, obese animals display a markedly blunted circadian oscillation in blood eNAMPT-hypothalamic NAD+-FOXO1 axis as well as LMA and EE. Our findings indicate that the eNAMPT regulation of hypothalamic NAD+ biosynthesis underlies circadian physiology and that this system can be significantly disrupted by obesity.

DOI: https://doi.org/10.1038/s41467-023-37517-6
Sci Adv. 2023 Apr 14. 9(15): eadf1956

Defective import of mitochondrial metabolic enzyme elicits ectopic metabolic stress.

Kazuya Nishio, Tomoyuki Kawarasaki, Yuki Sugiura, Shunsuke Matsumoto, Ayano Konoshima, Yuki Takano, Mayuko Hayashi, Fumihiko Okumura, Takumi Kamura, Tsunehiro Mizushima, Kunio Nakatsukasa.

Deficiencies in mitochondrial protein import are associated with a number of diseases. However, although nonimported mitochondrial proteins are at great risk of aggregation, it remains largely unclear how their accumulation causes cell dysfunction. Here, we show that nonimported citrate synthase is targeted for proteasomal degradation by the ubiquitin ligase SCFUcc1. Unexpectedly, our structural and genetic analyses revealed that nonimported citrate synthase appears to form an enzymatically active conformation in the cytosol. Its excess accumulation caused ectopic citrate synthesis, which, in turn, led to an imbalance in carbon flux of sugar, a reduction of the pool of amino acids and nucleotides, and a growth defect. Under these conditions, translation repression is induced and acts as a protective mechanism that mitigates the growth defect. We propose that the consequence of mitochondrial import failure is not limited to proteotoxic insults, but that the accumulation of a nonimported metabolic enzyme elicits ectopic metabolic stress.

DOI: https://doi.org/10.1126/sciadv.adf1956
J Biol Chem. 2023 Apr 12. pii: S0021-9258(23)00344-7. [Epub ahead of print] 104702

Interaction between poly-A binding protein PABPC4 and nuclear receptor corepressor NCoR1 modulates a metabolic stress response.

Oliveira Ag, Oliveira Ld, Cruz Mv, Guimarães Dspsf, Lima Ti, B C Santos-Fávero, Luchessi Ad, B A Pauletti, Leme Ap, Bajgelman Mc, Afonso J, Regitano Lca, Carvalho Hf, Carneiro Em, Kobarg J, Perissi V, Auwerx J, Silveira Lr.

  Mitochondria are organelles known primarily for generating ATP via the oxidative phosphorylation process. Environmental signals are sensed by whole organisms or cells and markedly affect this process, leading to alterations in gene transcription and, consequently, changes in mitochondrial function and biogenesis. The expression of mitochondrial genes is finely regulated by nuclear transcription factors, including nuclear receptors and their coregulators. Among the best-known coregulators is the nuclear receptor corepressor 1 (NCoR1). Muscle-specific knockout of NCoR1 in mice induces an oxidative phenotype, improving glucose and fatty acid metabolism. However, the mechanism by which NCoR1 is regulated remains elusive. In this work, we identified the Poly A Binding Protein (PABPC4) as a new NCoR1 interactor. Unexpectedly, we found that silencing of PABPC4 induced an oxidative phenotype in both C2C12 and MEF cells as indicated by increased oxygen consumption, mitochondria content, and reduced lactate production. Mechanistically, we demonstrated that PABPC4 silencing increased the ubiquitination and consequent degradation of NCoR1, leading to the derepression of PPAR-regulated genes. As a consequence, cells with PABPC4 silencing had a greater capacity to metabolize lipids, reduced intracellular lipid droplets and reduced cell death. Interestingly, in conditions known to induce mitochondrial function and biogenesis, both mRNA expression and PABPC4 protein content were markedly reduced. Our study, therefore, suggests that the lowering of PABPC4 expression may represent an adaptive event required to induce mitochondrial activity in response to metabolic stress in skeletal muscle cells. As such, the NCoR1-PABPC4 interface might be a new road to the treatment of metabolic diseases.

Keywords:  NCoR1; PABPC4; metabolism; mitochondria; nuclear receptors; transcription corepressor

DOI:  https://doi.org/10.1016/j.jbc.2023.104702
Nat Commun. 2023 Apr 11. 14(1): 2030

Cryo-EM structures of mitochondrial ABC transporter ABCB10 in apo and biliverdin-bound form.

Sheng Cao, Yihu Yang, Lili He, Yumo Hang, Xiaodong Yan, Hui Shi, Jiaquan Wu, Zhuqing Ouyang.

ABCB10, a member of ABC transporter superfamily that locates in the inner membrane of mitochondria, plays crucial roles in hemoglobin synthesis, antioxidative stress and stabilization of the iron transporter mitoferrin-1. Recently, it was found that ABCB10 is a mitochondrial biliverdin exporter. However, the molecular mechanism of biliverdin export by ABCB10 remains elusive. Here we report the cryo-EM structures of ABCB10 in apo (ABCB10-apo) and biliverdin-bound form (ABCB10-BV) at 3.67 Å and 2.85 Å resolution, respectively. ABCB10-apo adopts a wide-open conformation and may thus represent the apo form structure. ABCB10-BV forms a closed conformation and biliverdin situates in a hydrophobic pocket in one protomer and bridges the interaction through hydrogen bonds with the opposing one. We also identify cholesterols sandwiched by BVs and discuss the export dynamics based on these structural and biochemical observations.

DOI: https://doi.org/10.1038/s41467-023-37851-9