bims-mitpro Biomed News
on Mitochondrial Proteostasis
Issue of 2024‒03‒17
twelve papers selected by
Andreas Kohler, Umeå University
  1. The mitochondrial UPR induced by ATF5 attenuates intervertebral disc degeneration via cooperating with mitophagy.
  2. Modular Assembly of Mitochondrial β-Barrel Proteins.
  3. Enhancing healthy aging with small molecules: A mitochondrial perspective.
  4. Tracking the Activity and Position of Mitochondrial β-Barrel Proteins.
  5. Loss of FoxO1 activates an alternate mechanism of mitochondrial quality control for healthy adipose browning.
  6. The Ubiquitin Ligase RBX2/SAG Regulates Mitochondrial Ubiquitination and Mitophagy.
  7. Yeast Bxi1/Ybh3 mediates conserved mitophagy and apoptosis in yeast and mammalian cells: convergence in Bcl-2 family.
  8. BAG3 localizes to mitochondria in cardiac fibroblasts and regulates mitophagy.
  9. Mitochondria-Targeted Oligomeric α-Synuclein Induces TOM40 Degradation and Mitochondrial Dysfunction in Parkinson's Disease and Parkinsonism-Dementia of Guam.
  10. MoAti1 mediates mitophagy by facilitating recruitment of MoAtg8 to promote invasive growth in Magnaporthe oryzae.
  11. MitoLuc: A Luminescence-Based Assay to Study Real-Time Protein Import into Mitochondria.
  12. NME3 is a gatekeeper for DRP1-dependent mitophagy in hypoxia.
  1. Cell Biol Toxicol. 2024 Mar 13. 40(1): 16
    The mitochondrial UPR induced by ATF5 attenuates intervertebral disc degeneration via cooperating with mitophagy.
    Wen-Ning Xu, Huo-Liang Zheng, Run-Ze Yang, Yuan-Fang Sun, Bi-Rong Peng, Chun Liu, Jian Song, Sheng-Dan Jiang, Li-Xin Zhu.
      Intervertebral disc degeneration (IVDD) is an aging disease that results in a low quality of life and heavy socioeconomic burden. The mitochondrial unfolded protein response (UPRmt) take part in various aging-related diseases. Our research intents to explore the role and underlying mechanism of UPRmt in IVDD. Nucleus pulposus (NP) cells were exposed to IL-1β and nicotinamide riboside (NR) served as UPRmt inducer to treat NP cells. Detection of ATP, NAD + and NADH were used to determine the function of mitochondria. MRI, Safranin O-fast green and Immunohistochemical examination were used to determine the degree of IVDD in vivo. In this study, we discovered that UPRmt was increased markedly in the NP cells of human IVDD tissues than in healthy controls. In vitro, UPRmt and mitophagy levels were promoted in NP cells treated with IL-1β. Upregulation of UPRmt by NR and Atf5 overexpression inhibited NP cell apoptosis and further improved mitophagy. Silencing of Pink1 reversed the protective effects of NR and inhibited mitophagy induced by the UPRmt. In vivo, NR might attenuate the degree of IDD by activating the UPRmt in rats. In summary, the UPRmt was involved in IVDD by regulating Pink1-induced mitophagy. Mitophagy induced by the UPRmt might be a latent treated target for IVDD.
    Keywords:  Atf5; Intervertebral disc degeneration; Mitochondrial unfolded protein response; Mitophagy; Pink1
    DOI:  https://doi.org/10.1007/s10565-024-09854-9
  2. Methods Mol Biol. 2024 ;2778 201-220
    Modular Assembly of Mitochondrial β-Barrel Proteins.
    Rituparna Bhowmik, Fabian den Brave, Thomas Becker.
      Mitochondrial β-barrel proteins fulfill crucial roles in the biogenesis and function of the cell organelle. They mediate the import and membrane insertion of proteins and transport of small metabolites and ions. All β-barrel proteins are made as precursors on cytosolic ribosomes and are imported into mitochondria. The β-barrel proteins fold and assemble with partner proteins in the outer membrane. The in vitro import of radiolabelled proteins into isolated mitochondria is a powerful tool to investigate the import of β-barrel proteins, the folding of the β-barrel proteins, and their assembly into protein complexes. Altogether, the in vitro import assay is a versatile and crucial assay to analyze the mechanisms of the biogenesis of mitochondrial β-barrel proteins.
    Keywords:  Blue native electrophoresis; Mitochondria; Protein import assay; SAM complex; TOM complex; β-barrel proteins
    DOI:  https://doi.org/10.1007/978-1-0716-3734-0_13
  3. Med Res Rev. 2024 Mar 14.
    Enhancing healthy aging with small molecules: A mitochondrial perspective.
    Xiujiao Qin, Hongyuan Li, Huiying Zhao, Le Fang, Xiaohui Wang.
      The pursuit of enhanced health during aging has prompted the exploration of various strategies focused on reducing the decline associated with the aging process. A key area of this exploration is the management of mitochondrial dysfunction, a notable characteristic of aging. This review sheds light on the crucial role that small molecules play in augmenting healthy aging, particularly through influencing mitochondrial functions. Mitochondrial oxidative damage, a significant aspect of aging, can potentially be lessened through interventions such as coenzyme Q10, alpha-lipoic acid, and a variety of antioxidants. Additionally, this review discusses approaches for enhancing mitochondrial proteostasis, emphasizing the importance of mitochondrial unfolded protein response inducers like doxycycline, and agents that affect mitophagy, such as urolithin A, spermidine, trehalose, and taurine, which are vital for sustaining protein quality control. Of equal importance are methods for modulating mitochondrial energy production, which involve nicotinamide adenine dinucleotide boosters, adenosine 5'-monophosphate-activated protein kinase activators, and compounds like metformin and mitochondria-targeted tamoxifen that enhance metabolic function. Furthermore, the review delves into emerging strategies that encourage mitochondrial biogenesis. Together, these interventions present a promising avenue for addressing age-related mitochondrial degradation, thereby setting the stage for the development of innovative treatment approaches to meet this extensive challenge.
    Keywords:  aging; mitochondrial; mitophagy; redox homeostasis; unfolded protein response
    DOI:  https://doi.org/10.1002/med.22034
  4. Methods Mol Biol. 2024 ;2778 221-236
    Tracking the Activity and Position of Mitochondrial β-Barrel Proteins.
    Shuo Wang, Stephan Nussberger.
      Total interference reflection fluorescence (TIRF) microscopy of lipid bilayers is an effective technique for studying the lateral movement and ion channel activity of single integral membrane proteins. Here we describe how to integrate the mitochondrial outer membrane preprotein translocase TOM-CC and its β-barrel protein-conducting channel Tom40 into supported lipid bilayers to identify possible relationships between movement and channel activity. We propose that our approach can be readily applied to membrane protein channels where transient tethering to either membrane-proximal or intramembrane structures is accompanied by a change in channel permeation.
    Keywords:  Channels; Mitochondria; Protein translocation; Single molecule; TIRF microscopy; TOM; β-barrel membrane protein
    DOI:  https://doi.org/10.1007/978-1-0716-3734-0_14
  5. Clin Sci (Lond). 2024 Mar 20. 138(6): 371-385
    Loss of FoxO1 activates an alternate mechanism of mitochondrial quality control for healthy adipose browning.
    Limin Shi, Jinying Yang, Zhipeng Tao, Louise Zheng, Tyler F Bui, Ramon L Alonso, Feng Yue, Zhiyong Cheng.
      Browning of white adipose tissue is hallmarked by increased mitochondrial density and metabolic improvements. However, it remains largely unknown how mitochondrial turnover and quality control are regulated during adipose browning. In the present study, we found that mice lacking adipocyte FoxO1, a transcription factor that regulates autophagy, adopted an alternate mechanism of mitophagy to maintain mitochondrial turnover and quality control during adipose browning. Post-developmental deletion of adipocyte FoxO1 (adO1KO) suppressed Bnip3 but activated Fundc1/Drp1/OPA1 cascade, concurrent with up-regulation of Atg7 and CTSL. In addition, mitochondrial biogenesis was stimulated via the Pgc1α/Tfam pathway in adO1KO mice. These changes were associated with enhanced mitochondrial homeostasis and metabolic health (e.g., improved glucose tolerance and insulin sensitivity). By contrast, silencing Fundc1 or Pgc1α reversed the changes induced by silencing FoxO1, which impaired mitochondrial quality control and function. Ablation of Atg7 suppressed mitochondrial turnover and function, causing metabolic disorder (e.g., impaired glucose tolerance and insulin sensitivity), regardless of elevated markers of adipose browning. Consistently, suppression of autophagy via CTSL by high-fat diet was associated with a reversal of adO1KO-induced benefits. Our data reveal a unique role of FoxO1 in coordinating mitophagy receptors (Bnip3 and Fundc1) for a fine-tuned mitochondrial turnover and quality control, underscoring autophagic clearance of mitochondria as a prerequisite for healthy browning of adipose tissue.
    Keywords:  FoxO1; adipose browning; metabolism; mitochondrial quality control; mitophagy
    DOI:  https://doi.org/10.1042/CS20230973
  6. bioRxiv. 2024 Feb 28. pii: 2024.02.24.581168. [Epub ahead of print]
    The Ubiquitin Ligase RBX2/SAG Regulates Mitochondrial Ubiquitination and Mitophagy.
    Wenjuan Wang, Ermin Li, Jianqiu Zou, Qu Chen, Juan Ayala, Yuan Wen, Md Sadikul Islam, Neal L Weintraub, David J Fulton, Qiangrong Liang, Jiliang Zhou, Jinbao Liu, Jie Li, Yi Sun, Huabo Su.
      Clearance of damaged mitochondria via mitophagy is crucial for cellular homeostasis. While the role of ubiquitin (Ub) ligase PARKIN in mitophagy has been extensively studied, increasing evidence suggests the existence of PARKIN-independent mitophagy in highly metabolically active organs such as the heart. Here, we identify a crucial role for Cullin-RING Ub ligase 5 (CRL5) in basal mitochondrial turnover in cardiomyocytes. CRL5 is a multi-subunit Ub ligase comprised by the catalytic RING box protein RBX2 (also known as SAG), scaffold protein Cullin 5 (CUL5), and a substrate-recognizing receptor. Analysis of the mitochondrial outer membrane-interacting proteome uncovered a robust association of CRLs with mitochondria. Subcellular fractionation, immunostaining, and immunogold electron microscopy established that RBX2 and Cul5, two core components of CRL5, localizes to mitochondria. Depletion of RBX2 inhibited mitochondrial ubiquitination and turnover, impaired mitochondrial membrane potential and respiration, and increased cell death in cardiomyocytes. In vivo , deletion of the Rbx2 gene in adult mouse hearts suppressed mitophagic activity, provoked accumulation of damaged mitochondria in the myocardium, and disrupted myocardial metabolism, leading to rapid development of dilated cardiomyopathy and heart failure. Similarly, ablation of RBX2 in the developing heart resulted in dilated cardiomyopathy and heart failure. Notably, the action of RBX2 in mitochondria is not dependent on PARKIN, and PARKIN gene deletion had no impact on the onset and progression of cardiomyopathy in RBX2-deficient hearts. Furthermore, RBX2 controls the stability of PINK1 in mitochondria. Proteomics and biochemical analyses further revealed a global impact of RBX2 deficiency on the mitochondrial proteome and identified several mitochondrial proteins as its putative substrates. These findings identify RBX2-CRL5 as a mitochondrial Ub ligase that controls mitophagy under physiological conditions in a PARKIN-independent, PINK1-dependent manner, thereby regulating cardiac homeostasis.Non-standard abbreviations and acronyms: RBX2, RING-Box Protein 2; SAG, Sensitive to Apoptosis Gene; Ub, Ubiquitin; pS65-Ub, phosphorylated Ub at serine 65; MAVS, mitochondrial antiviral-signaling protein; AAV, adeno-associated virus; AV, adenovirus; siRNA, Small interfering RNA; GFP, green fluorescent protein; CUL, cullin; RING, Really Interesting New Gene; CRLs, cullin-RING ligases; CSN, COP9 signalosome; APEX2, ascorbate peroxidase 2; mito, mitochondrial; cyto, cytosolic; MOM, mitochondrial outer membrane; CCCP, Carbonyl Cyanide Chlorophenylhydrazone; OMP25, Outer membrane protein 25; PK, proteinase K; HA, hemagglutinin; TMRM, Tetramethylrhodamine methyl ester perchlorate; αMHC,α-myosin heavy chain; CKO, cardiomyocyte-specific knockout; TAM, tamoxifen; TMT, tandem mass tag; KD, knockdown; CTL, control; MCM, MerCreMer; iCKO, inducible cardiomyocyte-specific knockout; BFA, bafilomycin A1; PCA, principle component analysis; MS, Mass spectrometry; DEPs, differentially expressed proteins; FC, fold change; FDR, False Discovery Rate; KEGG, Kyoto encyclopedia of genes and genomes; ER, endoplasmic reticulum; DKO, double knockout; CM, cardiomyocyte; cTnT, cardiac troponin T; NRVCs, neonatal rat ventricular cardiomyocytes; NRVMs, neonatal mouse ventricular cardiomyocytes; NMVFs, neonatal mouse ventricular fibroblasts; HF, heart failure; KO, knockout; MF, Molecular Functions; CC, Cellular Components; BP, Biological Process; TUNEL, Terminal deoxynucleotidyl transferase dUTP nick end labeling; SCF, Skp1-Cullin 1-F-box.
    DOI:  https://doi.org/10.1101/2024.02.24.581168
  7. Biol Chem. 2024 Mar 12.
    Yeast Bxi1/Ybh3 mediates conserved mitophagy and apoptosis in yeast and mammalian cells: convergence in Bcl-2 family.
    Yuying Wang, Zhiyuan Hu, Maojun Jiang, Yanxin Zhang, Linjie Yuan, Ziqian Wang, Ting Song, Zhichao Zhang.
      The process of degrading unwanted or damaged mitochondria by autophagy, called mitophagy, is essential for mitochondrial quality control together with mitochondrial apoptosis. In mammalian cells, pan-Bcl-2 family members including conical Bcl-2 members and non-conical ones are involved in and govern the two processes. We have illustrated recently the BH3 receptor Hsp70 interacts with Bim to mediate both apoptosis and mitophagy. However, whether similar pathways exist in lower eukaryotes where conical Bcl-2 members are absent remained unclear. Here, a specific inhibitor of the Hsp70-Bim PPI, S1g-10 and its analogs were used as chemical tools to explore the role of yeast Bxi1/Ybh3 in regulating mitophagy and apoptosis. Using Om45-GFP processing assay, we illustrated that yeast Ybh3 mediates a ubiquitin-related mitophagy pathway in both yeast and mammalian cells through association with Hsp70, which is in the same manner with Bim. Moreover, by using Bax/Bak double knockout MEF cells, Ybh3 was identified to induce apoptosis through forming oligomerization to trigger mitochondrial outer membrane permeabilization (MOMP) like Bax. We not only illustrated a conserved ubiquitin-related mitophagy pathway in yeast but also revealed the multi-function of Ybh3 which combines the function of BH3-only protein and multi-domain Bax protein as one.
    Keywords:  Bxi1/Ybh3; Hsp70; mitophagy; ubiquitin
    DOI:  https://doi.org/10.1515/hsz-2023-0359
  8. Am J Physiol Heart Circ Physiol. 2024 Mar 15.
    BAG3 localizes to mitochondria in cardiac fibroblasts and regulates mitophagy.
    Thomas G Martin, Laura A Sherer, Jonathan A Kirk.
      The co-chaperone BAG3 is a central node in protein quality control in the heart. In humans and animal models, decreased BAG3 expression is associated with cardiac dysfunction and dilated cardiomyopathy. While previous studies focused on BAG3 in cardiomyocytes, cardiac fibroblasts are also critical drivers of pathologic remodeling. Yet, BAG3's role in cardiac fibroblasts is almost completely unexplored. Here, we show BAG3 is expressed in primary rat neonatal cardiac fibroblasts and preferentially localizes to mitochondria. Knockdown of BAG3 reduces mitophagy and enhances fibroblast activation, which is associated with fibrotic remodeling. Hsp70 is a critical binding partner for BAG3 and inhibiting this interaction in fibroblasts using the drug JG-98 decreased autophagy, decreased mitofusin-2 expression, and disrupted mitochondrial morphology. Together, this data indicates that BAG3 is expressed in cardiac fibroblasts, where it facilitates mitophagy and promotes fibroblast quiescence. This suggests that depressed BAG3 levels in heart failure may exacerbate fibrotic pathology, thus contributing to myocardial dysfunction through sarcomere-independent pathways.
    Keywords:  BAG3; cardiac fibroblasts; heart; mitophagy
    DOI:  https://doi.org/10.1152/ajpheart.00736.2023
  9. Res Sq. 2024 Feb 21. pii: rs.3.rs-3970470. [Epub ahead of print]
    Mitochondria-Targeted Oligomeric α-Synuclein Induces TOM40 Degradation and Mitochondrial Dysfunction in Parkinson's Disease and Parkinsonism-Dementia of Guam.
    Muralidhar Hegde, Velmarini Vasquez, Manohar Kodavati, Joy Mitra, Indira Vendula, Dale Hamilton, Ralph Garruto, K S Rao.
      Mitochondrial dysfunction is a central aspect of Parkinson's disease (PD) pathology, yet the underlying mechanisms are not fully understood. This study investigates the link between α-Synuclein (α-Syn) pathology and the loss of translocase of the outer mitochondrial membrane 40 (TOM40), unraveling its implications for mitochondrial dysfunctions in neurons. We discovered that TOM40 protein depletion occurs in the brains of patients with Guam Parkinsonism Dementia (Guam PD) and cultured neurons expressing α-Syn proteinopathy, notably, without corresponding changes in TOM40 mRNA levels. Cultured neurons expressing α-Syn mutants, with or without a mitochondria-targeting signal (MTS) underscore the role of α-Syn's mitochondrial localization in inducing TOM40 degradation. Parkinson's Disease related etiological factors, such as 6-hydroxy dopamine or ROS/metal ions stress, which promote α-Syn oligomerization, exacerbate TOM40 depletion in PD patient-derived cells with SNCA gene triplication. Although α-Syn interacts with both TOM40 and TOM20 in the outer mitochondrial membrane, degradation is selective for TOM40, which occurs via the ubiquitin-proteasome system (UPS) pathway. Our comprehensive analyses using Seahorse technology, mitochondrial DNA sequencing, and damage assessments, demonstrate that mutant α-Syn-induced TOM40 loss results in mitochondrial dysfunction, characterized by reduced membrane potential, accumulation of mtDNA damage, deletion/insertion mutations, and altered oxygen consumption rates. Notably, ectopic supplementation of TOM40 or reducing pathological forms of α-Syn using ADP-ribosylation inhibitors ameliorate these mitochondrial defects, suggesting potential therapeutic avenues. In conclusion, our findings provide crucial mechanistic insights into how α-Syn accumulation leads to TOM40 degradation and mitochondrial dysfunction, offering insights for targeted interventions to alleviate mitochondrial defects in PD.
    DOI:  https://doi.org/10.21203/rs.3.rs-3970470/v1
  10. Mol Plant Pathol. 2024 Mar;25(3): e13439
    MoAti1 mediates mitophagy by facilitating recruitment of MoAtg8 to promote invasive growth in Magnaporthe oryzae.
    Huanbin Shi, Shuai Meng, Jiehua Qiu, Shuwei Xie, Nan Jiang, Chaoxi Luo, Naweed I Naqvi, Yanjun Kou.
      Mitophagy is a selective autophagy for the degradation of damaged or excessive mitochondria to maintain intracellular homeostasis. In Magnaporthe oryzae, a filamentous ascomycetous fungus that causes rice blast, the most devastating disease of rice, mitophagy occurs in the invasive hyphae to promote infection. To date, only a few proteins are known to participate in mitophagy and the mechanisms of mitophagy are largely unknown in pathogenic fungi. Here, by a yeast two-hybrid screen with the core autophagy-related protein MoAtg8 as a bait, we obtained a MoAtg8 interactor MoAti1 (MoAtg8-interacting protein 1). Fluorescent observations and protease digestion analyses revealed that MoAti1 is primarily localized to the peripheral mitochondrial outer membrane and is responsible for recruiting MoAtg8 to mitochondria under mitophagy induction conditions. MoAti1 is specifically required for mitophagy, but not for macroautophagy and pexophagy. Infection assays suggested that MoAti1 is required for mitophagy in invasive hyphae during pathogenesis. Notably, no homologues of MoAti1 were found in rice and human protein databases, indicating that MoAti1 may be used as a potential target to control rice blast. By the host-induced gene silencing (HIGS) strategy, transgenic rice plants targeted to silencing MoATI1 showed enhanced resistance against M. oryzae with unchanged agronomic traits. Our results suggest that MoATI1 is required for mitophagy and pathogenicity in M. oryzae and can be used as a target for reducing rice blast.
    Keywords:  Atg8; fungal pathogen; pathogenicity; rice blast; selective autophagy
    DOI:  https://doi.org/10.1111/mpp.13439
  11. Methods Mol Biol. 2024 ;2778 185-200
    MitoLuc: A Luminescence-Based Assay to Study Real-Time Protein Import into Mitochondria.
    Holly C Ford, Ian Collinson.
      All but a few mitochondrial proteins are translated into the cytosol and imported in via complicated and varied pathways. These processes occur over short time frames and, as such, are difficult to monitor with classical approaches such as Western blotting or autoradiography that require sample collection at discrete time points. The development of an assay based on a split version of the small luciferase-Mitoluc-has allowed us to monitor the import of proteins into mitochondria in high resolution and real time (Pereira et al., J Mol Biol 431:1689-1699, 2019). Luminescence measurements are acquired using a plate reader in the order of seconds. This allows scores of experiments to be conducted in parallel in a single multi-well plate and permits kinetic analysis yielding information about import mechanisms (Ford et al., Elife 11:e75426, 2022).
    Keywords:  Luminescence; MitoLuc; Mitochondria; NanoLuc; Protein import; Split-luciferase
    DOI:  https://doi.org/10.1007/978-1-0716-3734-0_12
  12. Nat Commun. 2024 Mar 13. 15(1): 2264
    NME3 is a gatekeeper for DRP1-dependent mitophagy in hypoxia.
    Chih-Wei Chen, Chi Su, Chang-Yu Huang, Xuan-Rong Huang, Xiaojing Cuili, Tung Chao, Chun-Hsiang Fan, Cheng-Wei Ting, Yi-Wei Tsai, Kai-Chien Yang, Ti-Yen Yeh, Sung-Tsang Hsieh, Yi-Ju Chen, Yuxi Feng, Tony Hunter, Zee-Fen Chang.
      NME3 is a member of the nucleoside diphosphate kinase (NDPK) family localized on the mitochondrial outer membrane (MOM). Here, we report a role of NME3 in hypoxia-induced mitophagy dependent on its active site phosphohistidine but not the NDPK function. Mice carrying a knock-in mutation in the Nme3 gene disrupting NME3 active site histidine phosphorylation are vulnerable to ischemia/reperfusion-induced infarction and develop abnormalities in cerebellar function. Our mechanistic analysis reveals that hypoxia-induced phosphatidic acid (PA) on mitochondria is essential for mitophagy and the interaction of DRP1 with NME3. The PA binding function of MOM-localized NME3 is required for hypoxia-induced mitophagy. Further investigation demonstrates that the interaction with active NME3 prevents DRP1 susceptibility to MUL1-mediated ubiquitination, thereby allowing a sufficient amount of active DRP1 to mediate mitophagy. Furthermore, MUL1 overexpression suppresses hypoxia-induced mitophagy, which is reversed by co-expression of ubiquitin-resistant DRP1 mutant or histidine phosphorylatable NME3. Thus, the site-specific interaction with active NME3 provides DRP1 a microenvironment for stabilization to proceed the segregation process in mitophagy.
    DOI:  https://doi.org/10.1038/s41467-024-46385-7
This page is being maintained by Thomas Krichel. It was last updated on 2024‒03‒18 at 8:01.