bims-medica Biomed News
on Metabolism and diet in cancer
Issue of 2024‒11‒10
sixteen papers selected by
Brett Chrest, Wake Forest University
  1. Cancer Metabolism: Historical Landmarks, New Concepts, and Opportunities.
  2. Cellular ATP demand creates metabolically distinct subpopulations of mitochondria.
  3. Alveolar and Bone Marrow-derived Macrophages Differ in Metabolism and Glutamine Utilization.
  4. Adulthood dietary and lifestyle patterns and risk of breast cancer: Global Cancer Update Programme (CUP Global) systematic literature review.
  5. The multifaceted modulation of mitochondrial metabolism in tumorigenesis.
  6. The association between empirical dietary inflammatory pattern and risk of cancer and cancer-specific mortality: a systematic review and meta-analysis of prospective cohort studies.
  7. Mitochondrial elongation impairs breast cancer metastasis.
  8. Mitochondrial respiratory capacity is not altered in aging rat brains with or without memory impairment.
  9. The combination of venetoclax and quercetin exerts a cytotoxic effect on acute myeloid leukemia.
  10. Does a transmembrane sodium gradient control membrane potential in mammalian mitochondria?
  11. Reduced expressions of TCA cycle genes during aging in humans and mice.
  12. ACAT1 Induces the Differentiation of Glioblastoma Cells by Rewiring Choline Metabolism.
  13. Investigating serum concentration profiles of orally ingested short-chain fatty acid supplements.
  14. Characterising Cancer Cell Responses to Cyclic Hypoxia Using Mathematical Modelling.
  15. Mass spectrometry-based proteomics to study mutants and interactomes of mitochondrial translocation proteins.
  16. Mitochondrial respiratory complex II is altered in renal carcinoma.
  1. Cold Spring Harb Perspect Med. 2024 Nov 05. pii: a041814. [Epub ahead of print]
    Cancer Metabolism: Historical Landmarks, New Concepts, and Opportunities.
    Navdeep S Chandel, Karen H Vousden, Ralph J DeBerardinis.
      Cancer cells undergo changes in metabolism that distinguish them from non-malignant tissue. These may provide a growth advantage by promoting oncogenic signaling and redirecting intermediates to anabolic pathways that provide building blocks for new cellular components. Cancer metabolism is far from uniform, however, and recent work has shed light on its heterogenity within and between tumors. This work is also revealing how cancer metabolism adapts to the tumor microenvironment, as well as ways in which we may capitalize on metabolic changes in cancer cells to create new therapies.
    DOI:  https://doi.org/10.1101/cshperspect.a041814
  2. Nature. 2024 Nov 06.
    Cellular ATP demand creates metabolically distinct subpopulations of mitochondria.
    Keun Woo Ryu, Tak Shun Fung, Daphne C Baker, Michelle Saoi, Jinsung Park, Christopher A Febres-Aldana, Rania G Aly, Ruobing Cui, Anurag Sharma, Yi Fu, Olivia L Jones, Xin Cai, H Amalia Pasolli, Justin R Cross, Charles M Rudin, Craig B Thompson.
      Mitochondria serve a crucial role in cell growth and proliferation by supporting both ATP synthesis and the production of macromolecular precursors. Whereas oxidative phosphorylation (OXPHOS) depends mainly on the oxidation of intermediates from the tricarboxylic acid cycle, the mitochondrial production of proline and ornithine relies on reductive synthesis1. How these competing metabolic pathways take place in the same organelle is not clear. Here we show that when cellular dependence on OXPHOS increases, pyrroline-5-carboxylate synthase (P5CS)-the rate-limiting enzyme in the reductive synthesis of proline and ornithine-becomes sequestered in a subset of mitochondria that lack cristae and ATP synthase. This sequestration is driven by both the intrinsic ability of P5CS to form filaments and the mitochondrial fusion and fission cycle. Disruption of mitochondrial dynamics, by impeding mitofusin-mediated fusion or dynamin-like-protein-1-mediated fission, impairs the separation of P5CS-containing mitochondria from mitochondria that are enriched in cristae and ATP synthase. Failure to segregate these metabolic pathways through mitochondrial fusion and fission results in cells either sacrificing the capacity for OXPHOS while sustaining the reductive synthesis of proline, or foregoing proline synthesis while preserving adaptive OXPHOS. These findings provide evidence of the key role of mitochondrial fission and fusion in maintaining both oxidative and reductive biosyntheses in response to changing nutrient availability and bioenergetic demand.
    DOI:  https://doi.org/10.1038/s41586-024-08146-w
  3. Am J Respir Cell Mol Biol. 2024 Nov 05.
    Alveolar and Bone Marrow-derived Macrophages Differ in Metabolism and Glutamine Utilization.
    Christine L Vigeland, Jordan D Link, Henry S Beggs, Yazan Alwarawrah, Brandie M Ehrmann, Hong Dang, Claire M Doerschuk.
      Changes in metabolic activity are key regulators of macrophage activity. Pro-inflammatory macrophages upregulate glycolysis, which promotes an inflammatory phenotype, whereas pro-repair macrophages rely upon oxidative metabolism and glutaminolysis to support their activity. Work to understand how metabolism regulates macrophage phenotype has been done primarily in macrophage cell lines and bone marrow-derived macrophages (BMDM). Our study sought to understand changes in metabolic activity of murine tissue-resident alveolar macrophages (AM) in response to LPS stimulation and to contrast them to BMDM. These studies also determined the contribution of glutamine metabolism using the glutamine inhibitor, DON. We found that compared to BMDM, AM have higher rates of oxygen consumption and contain a higher concentration of intracellular metabolites involved in fatty acid oxidation. In response to LPS, BMDM but not AM increased rates of glycolysis. Inhibition of glutamine metabolism using DON altered the metabolic activity of AM but not BMDM. Within AM, glutamine inhibition led to increases in intracellular metabolites involved in glycolysis, the TCA cycle, fatty acid oxidation, and amino acid metabolism. Glutamine inhibition also altered the metabolic response to LPS within AM but not BMDM. Our data reveal striking differences in the metabolic activity of AM and BMDM.
    Keywords:  glutamine; glycolysis; lipids; macrophage; metabolomics
    DOI:  https://doi.org/10.1165/rcmb.2023-0249OC
  4. Am J Clin Nutr. 2024 Oct 23. pii: S0002-9165(24)00810-4. [Epub ahead of print]
    Adulthood dietary and lifestyle patterns and risk of breast cancer: Global Cancer Update Programme (CUP Global) systematic literature review.
    Jadwiga Konieczna, Alice Chaplin, Indira Paz-Graniel, Helen Croker, Nerea Becerra-Tomás, Georgios Markozannes, Konstantinos K Tsilidis, Laure Dossus, Esther M Gonzalez-Gil, Yikyung Park, John Krebs, Matty P Weijenberg, Monica L Baskin, Ellen Copson, Sarah J Lewis, Jacob C Seidell, Rajiv Chowdhury, Lynette Hill, Doris Sm Chan, Dora Romaguera.
      BACKGROUND: An increasing number of studies in recent years investigate various dietary and lifestyle patterns and associated breast cancer (BC) risk.OBJECTIVES: This study aimed to comprehensively synthesize and grade the evidence on dietary and lifestyle patterns and BC risk.
    METHODS: Databases were systematically searched up to 31 March, 2022, for evidence from RCTs and prospective cohort studies on adherence to a dietary pattern alone or in combination with lifestyle behaviors and incidence of or mortality from primary BC in adult females. Findings in all, premenopausal, and postmenopausal females were descriptively synthesized instead of meta-analyzed due to patterns heterogeneity. An independent Global Cancer Update Programme Expert Panel graded the strength of the evidence.
    RESULTS: A total of 84 publications were included. Results for patterns reflecting both a healthy diet and lifestyle were more consistent than for patterns that included diet only. There was strong-probable evidence that a priori World Cancer Research Fund/American Institute for Cancer Research (WCRF/AICR) and American Cancer Society (ACS) dietary and lifestyle scores may reduce BC risk in all and postmenopausal females, whereas in premenopausal females, less evidence was found contributing to limited-suggestive grade. There was also a limited-suggestive evidence that adherence to the Healthy Lifestyle Index and other diet and lifestyle scores may reduce BC risk in postmenopausal females; a posteriori Western/meat/alcohol dietary patterns may increase BC risk in postmenopausal females; and prudent/vegetarian/Mediterranean dietary patterns may reduce BC risk in all females. For the remaining patterns, evidence was graded as limited-no conclusions.
    CONCLUSIONS: Advice to adopt combined aspects of a healthy diet and lifestyle according to WCRF/AICR and ACS scores, encouraging a healthy weight, physical activity, alcohol and smoking avoidance, and a healthy diet rich in fruits, vegetables, (whole)grains and cereals and discouraging red and processed meat, can be proposed to females to lower BC risk. This review was registered at PROSPERO as ID CRD42021270129 (https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42021270129) on 28 August, 2021, and further updated on 4 May, 2022, in order to extend the search period.
    Keywords:  a posteriori dietary patterns; a priori dietary patterns; breast cancer incidence; dietary and lifestyle patterns; evidence grading; hybrid dietary patterns; systematic review
    DOI:  https://doi.org/10.1016/j.ajcnut.2024.10.003
  5. Mitochondrion. 2024 Nov 04. pii: S1567-7249(24)00135-1. [Epub ahead of print] 101977
    The multifaceted modulation of mitochondrial metabolism in tumorigenesis.
    Rajendiran Keerthiga, Yafang Xie, Desheng Pei, Ailing Fu.
      Changes in mitochondrial metabolism produce a malignant transformation from normal cells to tumor cells. Mitochondrial metabolism, comprising bioenergetic metabolism, biosynthetic process, biomolecular decomposition, and metabolic signal conversion, obviously forms a unique sign in the process of tumorigenesis. Several oncometabolites produced by mitochondrial metabolism maintain tumor phenotype, which are recognized as tumor indicators. The mitochondrial metabolism synchronizes the metabolic and genetic outcome to the potent tumor microenvironmental signals, thereby further promoting tumor initiation. Moreover, the bioenergetic and biosynthetic metabolism within tumor mitochondria orchestrates dynamic contributions toward cancer progression and invasion. In this review, we describe the contribution of mitochondrial metabolism in tumorigenesis through shaping several hallmarks such as microenvironment modulation, plasticity, mitochondrial calcium, mitochondrial dynamics, and epithelial-mesenchymal transition. The review will provide a new insight into the abnormal mitochondrial metabolism in tumorigenesis, which will be conducive to tumor prevention and therapy through targeting tumor mitochondria.
    Keywords:  EMT-MET transition; OXPHOS; Oncometabolites; Plasticity; TCA cycle; Tumorigenesis
    DOI:  https://doi.org/10.1016/j.mito.2024.101977
  6. Front Nutr. 2024 ;11 1462931
    The association between empirical dietary inflammatory pattern and risk of cancer and cancer-specific mortality: a systematic review and meta-analysis of prospective cohort studies.
    Fatemeh S Hosseini, Ali Nikparast, Elahe Etesami, Fatemeh Javaheri-Tafti, Golaleh Asghari.
      Background/aim: Current evidence indicates a correlation between the inflammatory potential of diet and the risk of cancer and cancer-specific mortality. This study aimed to assess the association between empirical dietary inflammatory pattern (EDIP), which has recently been designed based on the inflammatory potential of the diet, and the risk of cancer and cancer-specific mortality.Methods: A systematic literature search was conducted across the PubMed/Medline, Scopus, and Web of Science databases from January 2016 to March 2024. A random effects model was used to calculate the pooled effect size (ES) and 95% confidence intervals (95% CI). Heterogeneity between studies was assessed using the Cochran Q test and the I 2 statistic.
    Results: From the initial 229 records, 24 prospective cohort studies with 2,683,350 participants and 37,091 cancer incidence cases, as well as 20,819 cancer-specific mortality, were included in our study. Pooled results indicated a significant association between higher adherence to the EDIP and an increased risk of total cancer (ES: 1.10; 95% CI: 1.05-1.15; I 2 = 41.1), colorectal cancer (ES: 1.19; 95% CI: 1.11-1.27; I 2 = 41.1), and liver cancer (ES: 1.48; 95% CI: 1.14-1.94; I 2 = 36.9). However, no significant association between increased adherence to the EDIP and an increased risk of ovarian or endometrial cancer was found. Furthermore, greater adherence to the EDIP was significantly associated with an increased risk of cancer-specific mortality (ES: 1.18; 95% CI: 1.05-1.33; I 2 = 45.4).
    Conclusion: Our results showed that a diet with higher inflammatory properties is associated with an increased risk of cancer and cancer-specific mortality.
    Systematic review registration: PROSPERO registration no. CRD42024496912.
    Keywords:  EDIP; cancer; cancer-specific mortality; empirical dietary inflammatory pattern; meta-analysis
    DOI:  https://doi.org/10.3389/fnut.2024.1462931
  7. Sci Adv. 2024 Nov 08. 10(45): eadm8212
    Mitochondrial elongation impairs breast cancer metastasis.
    Lucía Minarrieta, Matthew G Annis, Yannick Audet-Delage, Hellen Kuasne, Alain Pacis, Catherine St-Louis, Alexander Nowakowski, Marco Biondini, Mireille Khacho, Morag Park, Peter M Siegel, Julie St-Pierre.
      Mitochondrial dynamics orchestrate many essential cellular functions, including metabolism, which is instrumental in promoting cancer growth and metastatic progression. However, how mitochondrial dynamics influences metastatic progression remains poorly understood. Here, we show that breast cancer cells with low metastatic potential exhibit a more fused mitochondrial network compared to highly metastatic cells. To study the impact of mitochondrial dynamics on metastasis, we promoted mitochondrial elongation in metastatic breast cancer cells by individual genetic deletion of three key regulators of mitochondrial fission (Drp1, Fis1, Mff) or by pharmacological intervention with leflunomide. Omics analyses revealed that mitochondrial elongation causes substantial alterations in metabolic pathways and processes related to cell adhesion. In vivo, enhanced mitochondrial elongation by loss of mitochondrial fission mediators or treatment with leflunomide notably reduced metastasis formation. Furthermore, the transcriptomic signature associated with elongated mitochondria correlated with improved clinical outcome in patients with breast cancer. Overall, our findings highlight mitochondrial dynamics as a potential therapeutic target in breast cancer.
    DOI:  https://doi.org/10.1126/sciadv.adm8212
  8. MicroPubl Biol. 2024 ;2024
    Mitochondrial respiratory capacity is not altered in aging rat brains with or without memory impairment.
    Pamela J Yao, Jeffrey M Long, Peter R Rapp, Dimitrios Kapogiannis.
      Mitochondria are essential for supporting the high metabolic demands that are required for brain function. Impairments in mitochondria have been linked to age-related decline in brain functions. Here, we investigate whether the mitochondrial respiratory capacity of brain cells is changed in cognitive aging. We used a rat model of normal cognitive aging and analyzed mitochondrial oxidative phosphorylation in frozen brain samples. Mitochondrial oxygen consumption rate analysis of the frontal cortex did not show any differences between young rats and aged rats with either intact memory or impaired spatial memory. Mitochondrial ATP synthase activity and quantity also did not differ between young and aged rats. These results suggest that the total level of mitochondrial respiratory capacity is preserved in the frontal cortex of aged rats and may not explain aging-associated cognitive impairment.
    DOI:  https://doi.org/10.17912/micropub.biology.001359
  9. Sci Rep. 2024 11 02. 14(1): 26418
    The combination of venetoclax and quercetin exerts a cytotoxic effect on acute myeloid leukemia.
    Renshi Kawakatsu, Kenjiro Tadagaki, Kenta Yamasaki, Yasumichi Kuwahara, Shinichiro Nakada, Tatsushi Yoshida.
      Venetoclax is a BH3 mimetic that was recently approved for the treatment of acute myeloid leukemia (AML) treatment. However, the effect of venetoclax on AML remains limited, and a novel strategy is required. Here, we demonstrate for the first time that the cytotoxic effect of venetoclax drastically increased when by combined with the naturally occurring flavonoid quercetin. Combined treatment with venetoclax and quercetin caused most of AML KG-1 cells to exhibit a condensed morphology. Cell cycle analysis revealed that the combination strongly induced cell death. Caspase inhibitor blocked this cell death, and the combination induced poly (ADP-ribose) polymerase (PARP) cleavage, indicating that apoptosis was the primary mechanism. These effects were also observed in another AML cell line Kasumi-1 but not in chronic myeloid leukemia (CML) K562 cells. Public data analysis demonstrated that B-cell/CLL lymphoma 2 (Bcl-2) expression is increased in AML cells compared to other malignant tumors, and the survival and the growth of AML cell line depends on Bcl-2. We found that quercetin increased Bcl-2-associated X protein (Bax) expression in KG-1. Our study provides a novel function for quercetin and presents a promising strategy for AML treatment using venetoclax.
    DOI:  https://doi.org/10.1038/s41598-024-78221-9
  10. Cell Calcium. 2024 Oct 23. pii: S0143-4160(24)00120-9. [Epub ahead of print]124 102962
    Does a transmembrane sodium gradient control membrane potential in mammalian mitochondria?
    David G Nicholls.
      In a recent publication, Hernansanz-Agusti̒n et al. propose that a sodium gradient across the inner mitochondrial membrane, generated by a Na+/H+ activity integral to Complex I can account for half of the mitochondrial membrane potential. This conflicts with conventional electrophysiological and chemiosmotic understanding.
    Keywords:  Calcium signaling; Goldman equation; Membrane potential; Mitochondria; Sodium proton exchange
    DOI:  https://doi.org/10.1016/j.ceca.2024.102962
  11. Biochem Biophys Res Commun. 2024 Oct 29. pii: S0006-291X(24)01453-0. [Epub ahead of print]738 150917
    Reduced expressions of TCA cycle genes during aging in humans and mice.
    Chao Zhang, Zhiyao Fu, Ren Zhang.
      Aging is associated with a decline in physiological functions and an increased risk of metabolic disorders. The liver, a key organ in metabolism, undergoes significant changes during aging that can contribute to systemic metabolic dysfunction. This study investigates the expression of genes involved in the tricarboxylic acid (TCA) cycle, a critical pathway for energy production, in the aging liver. We analyzed RNA sequencing data from the Genotype-Tissue Expression (GTEx) project to assess age-related changes in gene expression in the human liver. To validate our findings, we conducted complementary studies in young and old mice, examining the expression of key TCA cycle genes using quantitative real-time PCR. Our analysis of the GTEx dataset revealed a significant reduction in the expression of many genes that are critical for metabolism, including fat mass and obesity associated (FTO) and adiponectin receptor 1 (ADIPOR1). The most overrepresented pathway among the statistically enriched ones was the TCA cycle, with multiple genes exhibiting downregulation in older humans. This reduction was consistent with findings in aging mice, which also showed decreased expression of several TCA cycle genes. These results suggest a conserved pattern of age-related downregulation of TCA cycle, potentially leading to diminished mitochondrial function and energy production in the liver. The reduced expression of TCA cycle genes in the aging liver may contribute to metabolic dysfunction and increased susceptibility to age-related diseases. Understanding the molecular basis of these changes provides new insights into the aging process and highlights potential targets for interventions aimed at promoting healthy aging and preventing metabolic disorders.
    DOI:  https://doi.org/10.1016/j.bbrc.2024.150917
  12. Int J Biol Sci. 2024 ;20(14): 5576-5593
    ACAT1 Induces the Differentiation of Glioblastoma Cells by Rewiring Choline Metabolism.
    Shen You, Ming-Jin Wang, Zhen-Yan Hou, Wei-Da Wang, Zhi-Hui Zhang, Ting-Ting Du, Shu-Ying Li, Yi-Chen Liu, Ni-Na Xue, Xiao-Min Hu, Xiao-Guang Chen, Ming Ji.
      Abnormal differentiation of cells is a hallmark of malignancy. Induction of cancer-cell differentiation is emerging as a novel therapeutic strategy with low toxicity in hematological malignances, but whether such treatment can be used in solid tumors is not known. Here, we uncovered a novel function of acetyl coenzyme A acetyltransferase (ACAT1) in regulating the differentiation of glioblastoma (GBM) cells. Inhibition of ACAT1 promoted the differentiation of GBM cells into astrocytes but also delayed tumor growth. Mechanistically, suppression of ACAT1 restored mitochondrial function and led to metabolic "reprogramming" in GBM cells: reduction of fatty-acid oxidation and acetyl-CoA, but an increase in free fatty acids. Importantly, ACAT1 negatively regulated the choline metabolic pathway, which is crucial for the differentiation of GBM cells. Finally, we demonstrated that a naturally available substance, chlorogenic acid (CHA), could inhibit phosphorylation of ACAT1 and so delay GBM progression, CHA is a promising candidate to treat GBM because it could induce the differentiation of cancer cells.
    Keywords:  acetyl coenzyme A acetyltransferase; cell differentiation; choline; glioblastoma
    DOI:  https://doi.org/10.7150/ijbs.96651
  13. Food Funct. 2024 Nov 05.
    Investigating serum concentration profiles of orally ingested short-chain fatty acid supplements.
    Christopher G Green, Marilyn L Y Ong, Samantha N Rowland, Tindaro Bongiovanni, Lewis J James, Tom Clifford, Stephen J Bailey, Liam M Heaney.
      Acetate, propionate, and butyrate are naturally-occurring short-chain fatty acids (SCFAs) derived from bacterial metabolism of dietary fibre and have been associated with numerous positive health outcomes. All three acids have been shown to offer unique physiological and metabolic effects and, therefore, could be targeted for co-ingestion as part of a nutritional/medicinal plan. However, a better understanding of the outcomes of supplementing in combination on circulating concentration profiles is necessary to confirm uptake efficacy. This study sought to investigate the acute circulating concentration profiles of acetate, propionate, and butyrate following oral supplementation. Three experimental trials were conducted including investigations to understand the impact of capsule coating on circulating concentration profiles, the effect of supplementation dose on uptake kinetics, and the outcome of a short, repeated, supplementation routine on circulating levels. Serum samples were analysed for SCFA content using a quantitative GC-MS assay. It was observed that an acid-resistant coated capsule caused a delayed and blunted blood concentration response, with the non-acid resistant trial displaying earlier and more intense peak serum concentrations. For dose comparison investigations, all SCFAs peaked within 60 min and returned to baseline concentrations by 120 min post-supplementation. A graded dose relationship was present for propionate and butyrate when considering the total circulating exposure across a 240 min monitoring period. In addition, a one-week, twice-daily, repeated supplementation protocol resulted in no changes in basal serum SCFA concentrations. Overall, these data indicate that acetate, propionate, and butyrate display relatively similar circulating concentration profiles following oral co-ingestion, adding knowledge to help inform supplementation strategies for future outcomes where acute elevation of circulating SCFAs is desired.
    DOI:  https://doi.org/10.1039/d4fo04028g
  14. Bull Math Biol. 2024 Nov 06. 86(12): 145
    Characterising Cancer Cell Responses to Cyclic Hypoxia Using Mathematical Modelling.
    Giulia L Celora, Ruby Nixson, Joe M Pitt-Francis, Philip K Maini, Helen M Byrne.
      In vivo observations show that oxygen levels in tumours can fluctuate on fast and slow timescales. As a result, cancer cells can be periodically exposed to pathologically low oxygen levels; a phenomenon known as cyclic hypoxia. Yet, little is known about the response and adaptation of cancer cells to cyclic, rather than, constant hypoxia. Further, existing in vitro models of cyclic hypoxia fail to capture the complex and heterogeneous oxygen dynamics of tumours growing in vivo. Mathematical models can help to overcome current experimental limitations and, in so doing, offer new insights into the biology of tumour cyclic hypoxia by predicting cell responses to a wide range of cyclic dynamics. We develop an individual-based model to investigate how cell cycle progression and cell fate determination of cancer cells are altered following exposure to cyclic hypoxia. Our model can simulate standard in vitro experiments, such as clonogenic assays and cell cycle experiments, allowing for efficient screening of cell responses under a wide range of cyclic hypoxia conditions. Simulation results show that the same cell line can exhibit markedly different responses to cyclic hypoxia depending on the dynamics of the oxygen fluctuations. We also use our model to investigate the impact of changes to cell cycle checkpoint activation and damage repair on cell responses to cyclic hypoxia. Our simulations suggest that cyclic hypoxia can promote heterogeneity in cellular damage repair activity within vascular tumours.
    Keywords:  Cell cycle; Damage repair; Fluctuating oxygen levels; Individual-based modelling; Mathematical oncology; Tumour Hypoxia
    DOI:  https://doi.org/10.1007/s11538-024-01359-0
  15. Methods Enzymol. 2024 ;pii: S0076-6879(24)00404-X. [Epub ahead of print]707 101-152
    Mass spectrometry-based proteomics to study mutants and interactomes of mitochondrial translocation proteins.
    Silke Oeljeklaus, Lakshita Sharma, Julian Bender, Bettina Warscheid.
      The multiple functions of mitochondria are governed by their proteome comprising 1000-1500 proteins depending on the organism. However, only few proteins are synthesized inside mitochondria, whereas most are "born" outside mitochondria. To reach their destined location, these mitochondrial proteins follow specific import routes established by a mitochondrial translocase network. A detailed understanding of the role and interplay of the different translocases is imperative to understand mitochondrial biology and how mitochondria are integrated into the cellular network. Mass spectrometry (MS) proved to be effective to study the translocase network regarding composition, functions, interplay, and cellular responses evoked by dysfunction. In this chapter, we provide protocols tailored to MS-enabled functional analysis of mutants and interactomes of mitochondrial translocation proteins. In the first part, we exemplify the MS-based proteomics analysis of translocation mutants for delineating the human mitochondrial importome following depletion of the central translocation protein TOMM40. The protocol comprises metabolic stable isotope labeling, TOMM40 knockdown, preparation of mitochondrial fractions, and sample preparation for liquid chromatography (LC)-MS. For deep MS analysis, prefractionation of peptide mixtures by high pH reversed-phase LC is described. In the second part, we outline an affinity purification MS approach to reveal the association of an orphaned protein with the translocase TIM23. The protocol covers FLAG-tag affinity purification of protein complexes from mitochondrial fractions and downstream sample preparation for interactome analysis. In the last unifying part, we describe methods for LC-MS, data processing, statistical analysis and visualization of quantitative MS data, and provide a Python code for effective, customizable analysis.
    Keywords:  Affinity-purification mass spectrometry; Data analysis; Importome; Interactome; Mass spectrometry; Mitochondria; Protein quantification; Proteomics; SILAC; Translocation mutants
    DOI:  https://doi.org/10.1016/bs.mie.2024.07.059
  16. Biochim Biophys Acta Mol Basis Dis. 2024 Oct 31. pii: S0925-4439(24)00550-7. [Epub ahead of print]1871(1): 167556
    Mitochondrial respiratory complex II is altered in renal carcinoma.
    Sona Miklovicova, Luca Volpini, Ondrej Sanovec, Federica Monaco, Katerina Hadrava Vanova, Jaromir Novak, Stepana Boukalova, Renata Zobalova, Petr Klezl, Marco Tomasetti, Vladimir Bobek, Vojtech Fiala, Josef Vcelak, Lory Santarelli, Zuzana Bielcikova, Katerina Komrskova, Katarina Kolostova, Karel Pacak, Sarka Dvorakova, Jiri Neuzil.
      BACKGROUND: Renal cell carcinoma (RCC) is a disease typified by anomalies in cell metabolism. The function of mitochondria, including subunits of mitochondrial respiratory complex II (CII), in particular SDHB, are often affected. Here we investigated the state and function of CII in RCC patients.METHODS: We evaluated tumour tissue as well as the adjacent healthy kidney tissue of 78 patients with RCC of different histotypes, focusing on their mitochondrial function. As clear cell RCC (ccRCC) is by far the most frequent histotype of RCC, we focused on these patients, which were grouped based on the pathological WHO/ISUP grading system to low- and high-grade patients, indicative of prognosis. We also evaluated mitochondrial function in organoids derived from tumour tissue of 7 patients.
    RESULTS: ccRCC tumours were characterized by mutated von Hippel-Lindau gene and high expression of carbonic anhydrase IX. We found low levels of mitochondrial DNA, protein and function, together with CII function in ccRCC tumour tissue, but not in other RCC types and non-tumour tissues. Mitochondrial content increased in high-grade tumours, while the function of CII remained low. Tumour organoids from ccRCC patients recapitulated molecular characteristics of RCC tissue.
    CONCLUSIONS: Our findings suggest that the state of CII, epitomized by its assembly and SDHB levels, deteriorates with the progressive severity of ccRCC. These observations hold the potential for stratification of patients with worse prognosis and may guide the exploration of targeted therapeutic interventions.
    Keywords:  Complex II; Metabolism; Mitochondria; Organoids; Renal cell carcinoma; Succinate dehydrogenase
    DOI:  https://doi.org/10.1016/j.bbadis.2024.167556
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