bims-mitdyn 2023-02-19 papers

bims-mitdyn

Biomed News

on Mitochondrial dynamics: mechanisms

Issue of 2023‒02‒19
fourteen papers selected by
Edmond Chan
Queen’s University, School of Medicine

Reverse electron transfer is activated during aging and contributes to aging and age-related disease.
The outer mitochondrial membrane protein TMEM11 demarcates spatially restricted BNIP3/BNIP3L-mediated mitophagy.
Quantifying organellar ultrastructure in cryo-electron tomography using a surface morphometrics pipeline.
Cone photoreceptors transfer damaged mitochondria to Müller glia.
Dysregulation of mitochondrial translation caused by CBFB deficiency cooperates with mutant PIK3CA and is a vulnerability in breast cancer.
TRIM21 inhibits irradiation-induced mitochondrial DNA release and impairs antitumour immunity in nasopharyngeal carcinoma tumour models.
DddA homolog search and engineering expand sequence compatibility of mitochondrial base editing.
A trans-kingdom T6SS effector induces the fragmentation of the mitochondrial network and activates innate immune receptor NLRX1 to promote infection.
Mitochondrial control of microglial phagocytosis by the translocator protein and hexokinase 2 in Alzheimer's disease.
Iron status influences mitochondrial disease progression in Complex I-deficient mice.
ERK1/2-dependent TSPO overactivation associates with the loss of mitophagy and mitochondrial respiration in ALS.
Mitochondrial single-cell ATAC-seq for high-throughput multi-omic detection of mitochondrial genotypes and chromatin accessibility.
Disease-associated mutations in Drp1 have fundamentally different effects on the mitochondrial fission machinery.
Mitochondrial respiration reduces exposure of the nucleus to oxygen.

EMBO Rep. 2023 Feb 16. e55548

Reverse electron transfer is activated during aging and contributes to aging and age-related disease.

Suman Rimal, Ishaq Tantray, Yu Li, Tejinder Pal Khaket, Yanping Li, Sunil Bhurtel, Wen Li, Cici Zeng, Bingwei Lu.

  Mechanisms underlying the depletion of NAD+ and accumulation of reactive oxygen species (ROS) in aging and age-related disorders remain poorly defined. We show that reverse electron transfer (RET) at mitochondrial complex I, which causes increased ROS production and NAD+ to NADH conversion and thus lowered NAD+ /NADH ratio, is active during aging. Genetic or pharmacological inhibition of RET decreases ROS production and increases NAD+ /NADH ratio, extending the lifespan of normal flies. The lifespan-extending effect of RET inhibition is dependent on NAD+ -dependent Sirtuin, highlighting the importance of NAD+ /NADH rebalance, and on longevity-associated Foxo and autophagy pathways. RET and RET-induced ROS and NAD+ /NADH ratio changes are prominent in human induced pluripotent stem cell (iPSC) model and fly models of Alzheimer's disease (AD). Genetic or pharmacological inhibition of RET prevents the accumulation of faulty translation products resulting from inadequate ribosome-mediated quality control, rescues relevant disease phenotypes, and extends the lifespan of Drosophila and mouse AD models. Deregulated RET is therefore a conserved feature of aging, and inhibition of RET may open new therapeutic opportunities in the context of aging and age-related diseases including AD.

Keywords:  Alzheimer's disease; NAD+/NADH ratio; lifespan; mitochondrial complex I; reverse electron transport

DOI:  https://doi.org/10.15252/embr.202255548
J Cell Biol. 2023 Apr 03. pii: e202204021. [Epub ahead of print]222(4):

The outer mitochondrial membrane protein TMEM11 demarcates spatially restricted BNIP3/BNIP3L-mediated mitophagy.

Mehmet Oguz Gok, Olivia M Connor, Xun Wang, Cameron J Menezes, Claire B Llamas, Prashant Mishra, Jonathan R Friedman.

Mitochondria play critical roles in cellular metabolism and to maintain their integrity, they are regulated by several quality control pathways, including mitophagy. During BNIP3/BNIP3L-dependent receptor-mediated mitophagy, mitochondria are selectively targeted for degradation by the direct recruitment of the autophagy protein LC3. BNIP3 and/or BNIP3L are upregulated situationally, for example during hypoxia and developmentally during erythrocyte maturation. However, it is not well understood how they are spatially regulated within the mitochondrial network to locally trigger mitophagy. Here, we find that the poorly characterized mitochondrial protein TMEM11 forms a complex with BNIP3 and BNIP3L and co-enriches at sites of mitophagosome formation. We find that mitophagy is hyper-active in the absence of TMEM11 during both normoxia and hypoxia-mimetic conditions due to an increase in BNIP3/BNIP3L mitophagy sites, supporting a model that TMEM11 spatially restricts mitophagosome formation.

DOI: https://doi.org/10.1083/jcb.202204021
J Cell Biol. 2023 Apr 03. pii: e202204093. [Epub ahead of print]222(4):

Quantifying organellar ultrastructure in cryo-electron tomography using a surface morphometrics pipeline.

Benjamin A Barad, Michaela Medina, Daniel Fuentes, R Luke Wiseman, Danielle A Grotjahn.

Cellular cryo-electron tomography (cryo-ET) enables three-dimensional reconstructions of organelles in their native cellular environment at subnanometer resolution. However, quantifying ultrastructural features of pleomorphic organelles in three dimensions is challenging, as is defining the significance of observed changes induced by specific cellular perturbations. To address this challenge, we established a semiautomated workflow to segment organellar membranes and reconstruct their underlying surface geometry in cryo-ET. To complement this workflow, we developed an open-source suite of ultrastructural quantifications, integrated into a single pipeline called the surface morphometrics pipeline. This pipeline enables rapid modeling of complex membrane structures and allows detailed mapping of inter- and intramembrane spacing, curvedness, and orientation onto reconstructed membrane meshes, highlighting subtle organellar features that are challenging to detect in three dimensions and allowing for statistical comparison across many organelles. To demonstrate the advantages of this approach, we combine cryo-ET with cryo-fluorescence microscopy to correlate bulk mitochondrial network morphology (i.e., elongated versus fragmented) with membrane ultrastructure of individual mitochondria in the presence and absence of endoplasmic reticulum (ER) stress. Using our pipeline, we demonstrate ER stress promotes adaptive remodeling of ultrastructural features of mitochondria including spacing between the inner and outer membranes, local curvedness of the inner membrane, and spacing between mitochondrial cristae. We show that differences in membrane ultrastructure correlate to mitochondrial network morphologies, suggesting that these two remodeling events are coupled. Our pipeline offers opportunities for quantifying changes in membrane ultrastructure on a single-cell level using cryo-ET, opening new opportunities to define changes in ultrastructural features induced by diverse types of cellular perturbations.

DOI: https://doi.org/10.1083/jcb.202204093
Cell Rep. 2023 Feb 15. pii: S2211-1247(23)00126-2. [Epub ahead of print]42(2): 112115

Cone photoreceptors transfer damaged mitochondria to Müller glia.

Rachel A Hutto, Kaitlyn M Rutter, Michelle M Giarmarco, Edward D Parker, Zachary S Chambers, Susan E Brockerhoff.

  Mitochondria are vital organelles that require sophisticated homeostatic mechanisms for maintenance. Intercellular transfer of damaged mitochondria is a recently identified strategy broadly used to improve cellular health and viability. Here, we investigate mitochondrial homeostasis in the vertebrate cone photoreceptor, the specialized neuron that initiates our daytime and color vision. We find a generalizable response to mitochondrial stress that leads to loss of cristae, displacement of damaged mitochondria from their normal cellular location, initiation of degradation, and transfer to Müller glia cells, a key non-neuronal support cell in the retina. Our findings show transmitophagy from cones to Müller glia as a response to mitochondrial damage. Intercellular transfer of damaged mitochondria represents an outsourcing mechanism that photoreceptors use to support their specialized function.

Keywords:  CP: Neuroscience; Müller glia; mitochondria; mitophagy; photoreceptor; retina; zebrafish

DOI:  https://doi.org/10.1016/j.celrep.2023.112115
Cancer Res. 2023 Feb 14. pii: CAN-22-2525. [Epub ahead of print]

Dysregulation of mitochondrial translation caused by CBFB deficiency cooperates with mutant PIK3CA and is a vulnerability in breast cancer.

Navdeep Malik, Young-Im Kim, Hualong Yan, Yu-Chou Tseng, Wendy du Bois, Gamze Ayaz, Andy D Tran, Laura Vera Ramírez, Howard H Yang, Aleksandra M Michalowski, Michael Kruhlak, Maxwell Lee, Kent W Hunter, Jing Huang.

Understanding functional interactions between cancer mutations is an attractive strategy for discovering unappreciated cancer pathways and developing new combination therapies to improve personalized treatment. However, distinguishing driver gene pairs from passenger pairs remains challenging. Here, we designed an integrated omics approach to identify driver gene pairs by leveraging genetic interaction analyses of top mutated breast cancer genes and the proteomics interactome data of their encoded proteins. This approach identified that PIK3CA oncogenic gain-of-function (GOF) and CBFB loss-of-function (LOF) mutations cooperate to promote breast tumor progression in both mice and humans. The transcription factor CBFB localized to mitochondria and moonlighted in translating the mitochondrial genome. Mechanistically, CBFB enhanced the binding of mitochondrial mRNAs to TUFM, a mitochondrial translation elongation factor. Independent of mutant PI3K, mitochondrial translation defects caused by CBFB LOF led to multiple metabolic reprogramming events, including defective oxidative phosphorylation (OXPHOS), the Warburg effect, and autophagy/mitophagy addiction. Furthermore, autophagy and PI3K inhibitors synergistically killed breast cancer cells and impaired the growth of breast tumors, including patient-derived xenografts (PDXs) carrying CBFB LOF and PIK3CA GOF mutations. Thus, our study offers mechanistic insights into the functional interaction between mutant PI3K and mitochondrial translation dysregulation in breast cancer progression and provides a strong preclinical rationale for combining autophagy and PI3K inhibitors in precision medicine for breast cancer.

DOI: https://doi.org/10.1158/0008-5472.CAN-22-2525
Nat Commun. 2023 Feb 16. 14(1): 865

TRIM21 inhibits irradiation-induced mitochondrial DNA release and impairs antitumour immunity in nasopharyngeal carcinoma tumour models.

Jun-Yan Li, Yin Zhao, Sha Gong, Miao-Miao Wang, Xu Liu, Qing-Mei He, Ying-Qin Li, Sheng-Yan Huang, Han Qiao, Xi-Rong Tan, Ming-Liang Ye, Xun-Hua Zhu, Shi-Wei He, Qian Li, Ye-Lin Liang, Kai-Lin Chen, Sai-Wei Huang, Qing-Jie Li, Jun Ma, Na Liu.

Although radiotherapy can promote antitumour immunity, the mechanisms underlying this phenomenon remain unclear. Here, we demonstrate that the expression of the E3 ubiquitin ligase, tumour cell-intrinsic tripartite motif-containing 21 (TRIM21) in tumours, is inversely associated with the response to radiation and CD8+ T cell-mediated antitumour immunity in nasopharyngeal carcinoma (NPC). Knockout of TRIM21 modulates the cGAS/STING cytosolic DNA sensing pathway, potentiates the antigen-presenting capacity of NPC cells, and activates cytotoxic T cell-mediated antitumour immunity in response to radiation. Mechanistically, TRIM21 promotes the degradation of the mitochondrial voltage-dependent anion-selective channel protein 2 (VDAC2) via K48-linked ubiquitination, which inhibits pore formation by VDAC2 oligomers for mitochondrial DNA (mtDNA) release, thereby inhibiting type-I interferon responses following radiation exposure. In patients with NPC, high TRIM21 expression was associated with poor prognosis and early tumour relapse after radiotherapy. Our findings reveal a critical role of TRIM21 in radiation-induced antitumour immunity, providing potential targets for improving the efficacy of radiotherapy in patients with NPC.

DOI: https://doi.org/10.1038/s41467-023-36523-y
Nat Commun. 2023 Feb 16. 14(1): 874

DddA homolog search and engineering expand sequence compatibility of mitochondrial base editing.

Li Mi, Ming Shi, Yu-Xuan Li, Gang Xie, Xichen Rao, Damu Wu, Aimin Cheng, Mengxiao Niu, Fengli Xu, Ying Yu, Ning Gao, Wensheng Wei, Xianhua Wang, Yangming Wang.

Expanding mitochondrial base editing tools with broad sequence compatibility is of high need for both research and therapeutic purposes. In this study, we identify a DddA homolog from Simiaoa sunii (Ddd_Ss) which can efficiently deaminate cytosine in DC context in double-stranded DNA (dsDNA). We successfully develop Ddd_Ss-derived cytosine base editors (DdCBE_Ss) and introduce mutations at multiple mitochondrial DNA (mtDNA) loci including disease-associated mtDNA mutations in previously inaccessible GC context. Finally, by introducing a single amino acid substitution from Ddd_Ss, we successfully improve the activity and sequence compatibility of DdCBE derived from DddA of Burkholderia cenocepacia (DdCBE_Bc). Our study expands mtDNA editing tool boxes and provides resources for further screening and engineering dsDNA base editors for biological and therapeutic applications.

DOI: https://doi.org/10.1038/s41467-023-36600-2
Nat Commun. 2023 Feb 16. 14(1): 871

A trans-kingdom T6SS effector induces the fragmentation of the mitochondrial network and activates innate immune receptor NLRX1 to promote infection.

Joana Sá-Pessoa, Sara López-Montesino, Kornelia Przybyszewska, Isabel Rodríguez-Escudero, Helina Marshall, Adelia Ova, Gunnar N Schroeder, Peter Barabas, María Molina, Tim Curtis, Víctor J Cid, José A Bengoechea.

Bacteria can inhibit the growth of other bacteria by injecting effectors using a type VI secretion system (T6SS). T6SS effectors can also be injected into eukaryotic cells to facilitate bacterial survival, often by targeting the cytoskeleton. Here, we show that the trans-kingdom antimicrobial T6SS effector VgrG4 from Klebsiella pneumoniae triggers the fragmentation of the mitochondrial network. VgrG4 colocalizes with the endoplasmic reticulum (ER) protein mitofusin 2. VgrG4 induces the transfer of Ca2+ from the ER to the mitochondria, activating Drp1 (a regulator of mitochondrial fission) thus leading to mitochondrial network fragmentation. Ca2+ elevation also induces the activation of the innate immunity receptor NLRX1 to produce reactive oxygen species (ROS). NLRX1-induced ROS limits NF-κB activation by modulating the degradation of the NF-κB inhibitor IκBα. The degradation of IκBα is triggered by the ubiquitin ligase SCFβ-TrCP, which requires the modification of the cullin-1 subunit by NEDD8. VgrG4 abrogates the NEDDylation of cullin-1 by inactivation of Ubc12, the NEDD8-conjugating enzyme. Our work provides an example of T6SS manipulation of eukaryotic cells via alteration of the mitochondria.

DOI: https://doi.org/10.1038/s41467-023-36629-3
Proc Natl Acad Sci U S A. 2023 Feb 21. 120(8): e2209177120

Mitochondrial control of microglial phagocytosis by the translocator protein and hexokinase 2 in Alzheimer's disease.

Lauren H Fairley, Kei Onn Lai, Jia Hui Wong, Wei Jing Chong, Anselm Salvatore Vincent, Giuseppe D'Agostino, Xiaoting Wu, Roshan R Naik, Anusha Jayaraman, Sarah R Langley, Christiane Ruedl, Anna M Barron.

  Microglial phagocytosis is an energetically demanding process that plays a critical role in the removal of toxic protein aggregates in Alzheimer's disease (AD). Recent evidence indicates that a switch in energy production from mitochondrial respiration to glycolysis disrupts this important protective microglial function and may provide therapeutic targets for AD. Here, we demonstrate that the translocator protein (TSPO) and a member of its mitochondrial complex, hexokinase-2 (HK), play critical roles in microglial respiratory-glycolytic metabolism and phagocytosis. Pharmacological and genetic loss-of-function experiments showed that TSPO is critical for microglial respiratory metabolism and energy supply for phagocytosis, and its expression is enriched in phagocytic microglia of AD mice. Meanwhile, HK controlled glycolytic metabolism and phagocytosis via mitochondrial binding or displacement. In cultured microglia, TSPO deletion impaired mitochondrial respiration and increased mitochondrial recruitment of HK, inducing a switch to glycolysis and reducing phagocytosis. To determine the functional significance of mitochondrial HK recruitment, we developed an optogenetic tool for reversible control of HK localization. Displacement of mitochondrial HK inhibited glycolysis and improved phagocytosis in TSPO-knockout microglia. Mitochondrial HK recruitment also coordinated the inflammatory switch to glycolysis that occurs in response to lipopolysaccharide in normal microglia. Interestingly, cytosolic HK increased phagocytosis independent of its metabolic activity, indicating an immune signaling function. Alzheimer's beta amyloid drastically stimulated mitochondrial HK recruitment in cultured microglia, which may contribute to microglial dysfunction in AD. Thus, targeting mitochondrial HK may offer an immunotherapeutic approach to promote phagocytic microglial function in AD.

Keywords:  Alzheimer’s disease; hexokinase; immunometabolism; mitochondria; translocator protein

DOI:  https://doi.org/10.1073/pnas.2209177120
Elife. 2023 Feb 17. pii: e75825. [Epub ahead of print]12

Iron status influences mitochondrial disease progression in Complex I-deficient mice.

C J Kelly, Reid K Couch, Vivian T Ha, Camille M Bodart, Judy Wu, Sydney Huff, Nicole T Herrel, Hyunsung D Kim, Azaad O Zimmermann, Jessica Shattuck, Yu-Chen Pan, Matt Kaeberlein, Anthony S Grillo.

  Mitochondrial dysfunction caused by aberrant Complex I assembly and reduced activity of the electron transport chain is pathogenic in many genetic and age-related diseases. Mice missing the Complex I subunit NADH dehydrogenase [ubiquinone] iron-sulfur protein 4 (NDUFS4) are a leading mammalian model of severe mitochondrial disease that exhibit many characteristic symptoms of Leigh Syndrome including oxidative stress, neuroinflammation, brain lesions, and premature death. NDUFS4 knockout mice have decreased expression of nearly every Complex I subunit. As Complex I normally contains at least 8 iron-sulfur clusters and more than 25 iron atoms, we asked whether a deficiency of Complex I may lead to iron perturbations thereby accelerating disease progression. Consistent with this, iron supplementation accelerates symptoms of brain degeneration in these mice while iron restriction delays the onset of these symptoms, reduces neuroinflammation, and increases survival. NDUFS4 knockout mice display signs of iron overload in the liver including increased expression of hepcidin, and show changes in iron responsive element-regulated proteins consistent with increased cellular iron that were prevented by iron restriction. These results suggest that perturbed iron homeostasis may contribute to pathology in Leigh Syndrome and possibly other mitochondrial disorders.

Keywords:  biochemistry; chemical biology; medicine; mouse

DOI:  https://doi.org/10.7554/eLife.75825
Cell Death Dis. 2023 Feb 15. 14(2): 122

ERK1/2-dependent TSPO overactivation associates with the loss of mitophagy and mitochondrial respiration in ALS.

Andrea Magrì, Cristiana Lucia Rita Lipari, Pierpaolo Risiglione, Stefania Zimbone, Francesca Guarino, Antonella Caccamo, Angela Messina.

Mitochondrial dysfunction and the loss of mitophagy, aimed at recycling irreversibly damaged organelles, contribute to the onset of amyotrophic lateral sclerosis (ALS), a fatal neurodegenerative disease affecting spinal cord motor neurons. In this work, we showed that the reduction of mitochondrial respiration, exactly oxygen flows linked to ATP production and maximal capacity, correlates with the appearance of the most common ALS motor symptoms in a transgenic mouse model expressing SOD1 G93A mutant. This is the result of the equal inhibition in the respiration linked to complex I and II of the electron transport chain, but not their protein levels. Since the overall mitochondrial mass was unvaried, we investigated the expression of the Translocator Protein (TSPO), a small mitochondrial protein whose overexpression was recently linked to the loss of mitophagy in a model of Parkinson's disease. Here we clearly showed that levels of TSPO are significantly increased in ALS mice. Mechanistically, this increase is linked to the overactivation of ERK1/2 pathway and correlates with a decrease in the expression of the mitophagy-related marker Atg12, indicating the occurrence of impairments in the activation of mitophagy. Overall, our work sets out TSPO as a key regulator of mitochondrial homeostasis in ALS.

DOI: https://doi.org/10.1038/s41419-023-05643-0
Nat Protoc. 2023 Feb 15.

Mitochondrial single-cell ATAC-seq for high-throughput multi-omic detection of mitochondrial genotypes and chromatin accessibility.

Caleb A Lareau, Vincent Liu, Christoph Muus, Samantha D Praktiknjo, Lena Nitsch, Pauline Kautz, Katalin Sandor, Yajie Yin, Jacob C Gutierrez, Karin Pelka, Ansuman T Satpathy, Aviv Regev, Vijay G Sankaran, Leif S Ludwig.

Natural sequence variation within mitochondrial DNA (mtDNA) contributes to human phenotypes and may serve as natural genetic markers in human cells for clonal and lineage tracing. We recently developed a single-cell multi-omic approach, called 'mitochondrial single-cell assay for transposase-accessible chromatin with sequencing' (mtscATAC-seq), enabling concomitant high-throughput mtDNA genotyping and accessible chromatin profiling. Specifically, our technique allows the mitochondrial genome-wide inference of mtDNA variant heteroplasmy along with information on cell state and accessible chromatin variation in individual cells. Leveraging somatic mtDNA mutations, our method further enables inference of clonal relationships among native ex vivo-derived human cells not amenable to genetic engineering-based clonal tracing approaches. Here, we provide a step-by-step protocol for the use of mtscATAC-seq, including various cell-processing and flow cytometry workflows, by using primary hematopoietic cells, subsequent single-cell genomic library preparation and sequencing that collectively take ~3-4 days to complete. We discuss experimental and computational data quality control metrics and considerations for the extension to other mammalian tissues. Overall, mtscATAC-seq provides a broadly applicable platform to map clonal relationships between cells in human tissues, investigate fundamental aspects of mitochondrial genetics and enable additional modes of multi-omic discovery.

DOI: https://doi.org/10.1038/s41596-022-00795-3
Hum Mol Genet. 2023 Feb 16. pii: ddad029. [Epub ahead of print]

Disease-associated mutations in Drp1 have fundamentally different effects on the mitochondrial fission machinery.

Brianna L Bauer, Kristy Rochon, Jasmine C Liu, Rajesh Ramachandran, Jason A Mears.

Patient mutations have been identified throughout dynamin-related protein 1 (Drp1), the key protein mediator of mitochondrial fission. These changes generally impact young children and often result in severe neurological defects and, in some instances, death. Until now, the underlying functional defect leading to patient phenotypes has been largely speculative. We therefore analyzed six disease-associated mutations throughout the GTPase and middle domains of Drp1. The middle domain (MD) plays a role in Drp1 oligomerization, and three mutations in this region were predictably impaired in self-assembly. However, another mutant in this region (F370C) retained oligomerization capability on pre-curved membranes despite being assembly-limited in solution. Instead, this mutation impaired membrane remodeling of liposomes, which highlights the importance of Drp1 in generating local membrane curvature before fission. Two GTPase domain mutations were also observed in different patients. The G32A mutation was impaired in GTP hydrolysis both in solution and in the presence of lipid but remains capable of self-assembly on these lipid templates. The G223V mutation also exhibited decreased GTPase activity and was able to assemble on pre-curved lipid templates; however, this change impaired membrane remodeling of unilamellar liposomes similar to F370C. This demonstrates that the Drp1 GTPase domain also contributes to self-assembly interactions that drive membrane curvature. Overall, the functional defects caused by mutations in Drp1 are highly variable even for mutations that reside within the same functional domain. This study provides a framework for characterizing additional Drp1 mutations to provide a comprehensive understanding of functional sites within this essential protein.

DOI: https://doi.org/10.1093/hmg/ddad029
J Biol Chem. 2023 Feb 14. pii: S0021-9258(23)00150-3. [Epub ahead of print] 103018

Mitochondrial respiration reduces exposure of the nucleus to oxygen.

Mateus Prates Mori, Rozhin Penjweini, Jin Ma, Greg Alspaugh, Alessio Andreoni, Young-Chae Kim, Ping-Yuan Wang, Jay R Knutson, Paul M Hwang.

  The endosymbiotic theory posits that ancient eukaryotic cells engulfed O2-consuming prokaryotes, which protected them against O2 toxicity. Previous studies have shown that cells lacking cytochrome c oxidase (COX), required for respiration, have increased DNA damage and reduced proliferation, which could be improved by reducing O2 exposure. With recently developed fluorescence lifetime microscopy (FLIM)-based probes demonstrating that the mitochondrial compartment has lower [O2] than the cytosol, we hypothesized that the perinuclear distribution of mitochondria in cells may create a barrier for O2 to access the nuclear core, potentially affecting cellular physiology and maintaining genomic integrity. To test this hypothesis, we utilized myoglobin (MB)-mCherry FLIM O2 sensors without subcellular targeting ("cytosol") or with targeting to the mitochondrion or nucleus for measuring their localized O2 homeostasis. Our results showed that, similar to the mitochondria, the nuclear [O2] was reduced by ∼20-40% compared to the cytosol under imposed O2 levels of ∼0.5-18.6%. Pharmacologic inhibition of respiration increased nuclear O2 levels, and reconstituting O2 consumption by COX reversed this increase. Similarly, genetic disruption of respiration by deleting SCO2, a gene essential for COX assembly, or restoring COX activity in SCO2-/- cells by transducing with SCO2 cDNA also replicated these changes in nuclear O2 levels. The results were further supported by the expression of genes known to be affected by cellular O2 availability. Our study reveals the potential for dynamic regulation of nuclear O2 levels by mitochondrial respiratory activity, which in turn could affect oxidative stress and cellular processes such as neurodegeneration and aging.

Keywords:  Classification: Cell Biology; Metabolism; hypoxia; mitochondria; nucleus; oxygen; respiration

DOI:  https://doi.org/10.1016/j.jbc.2023.103018