Nucleus-Tethered Mitochondria are essential for Ca²⁺ homeostasis

Nucleus-Tethered Mitochondria are essential for Ca²⁺ homeostasis | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Letter Nucleus-Tethered Mitochondria are essential for Ca² ⁺ homeostasis Michelangelo Campanella, Danilo Faccenda, Tong Guo, Eva Sidlauskaite, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6015469/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted You are reading this latest preprint version Abstract Mitochondria form dynamic membrane contact sites (MCSs) with the surrounding organelles, including the nucleus, which we termed ‘nucleus-associated mitochondria’ (NAM). In this manuscript, we report that the stoichiometry of the NAM-forming molecule TSPO is modified in neurodegeneration diseases, and the nucleus-based component of the chromatin- and lamin-binding protein thymopoietin (TMPO/LAP2) is part of the tethering complex. Furthermore, we show that the NAM-regulated distance between mitochondria and the nucleus is pivotal for the physiological cycling of Ca 2+ in neuronal cells. On this, the transcription of Ca 2+ -regulated genes and those that dictate the spatiotemporal execution of Ca 2+ -such as inositol-1,4,5-trisphosphate (IP3) and ryanodine (RyR) is also dependent. This data set advances our understanding of the NAM biochemistry and function, shedding light on the mechanisms that govern mitochondrial coupling with the nucleus, aiding the homeostatic transduction of cell signalling. Biological sciences/Cell biology/Cell signalling/Calcium signalling Biological sciences/Cell biology/Organelles/Mitochondria Mitochondria form dynamic membrane contact sites (MCSs) TSPO TMPO/LAP2 Ca2+ signalling Figures Figure 1 Figure 2 Figure 3 Figure 4 Full Text Additional Declarations There is NO Competing Interest. Tables are available in the Supplementary Files section. Supplementary Files TablesFinalMC.pdf SFiguresFinal.pdf Cite Share Download PDF Status: Under Review Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6015469","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Letter","associatedPublications":[],"authors":[{"id":419107535,"identity":"f7ae9758-e302-4ad3-8376-a1f8e3060d74","order_by":0,"name":"Michelangelo Campanella","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA9ElEQVRIiWNgGAWjYFADduYGhg9Amg/CTSBCCzNjA+MMIM1GkhZmHmK06DYwP/x0c4ddHj8zY+Nnm7JtiW0MzA8/MLal4dRidoDNWDr3THKxZDNjs3TOudtALWzGEoxtOXi08DBI57YxJ244zNgAZIC0MJgxMLZV4NPC/Du3rT5x/2HG5t+WYC3s3whpYQMafjhxAzNjmzQjWAsPyBY8DjvMZmade+Z44ozDjG2WPeduG7cx8xRLJJzD4/3jzY9v5+6oTuxvbz5840fZbdl+9vaNHz6UJePUwsAMxIwNMB4bVCQBtwYIQNUyCkbBKBgFowANAAAXelA3nVa2zgAAAABJRU5ErkJggg==","orcid":"","institution":"QMUL William Harvey Research Institute","correspondingAuthor":true,"prefix":"","firstName":"Michelangelo","middleName":"","lastName":"Campanella","suffix":""},{"id":419107536,"identity":"f67d4b49-632d-4391-8eee-cd31af0aeb60","order_by":1,"name":"Danilo Faccenda","email":"","orcid":"","institution":"Pharmaceutical and Biological Sciences, University of Hertfordshire","correspondingAuthor":false,"prefix":"","firstName":"Danilo","middleName":"","lastName":"Faccenda","suffix":""},{"id":419107537,"identity":"3c8d9226-b76c-4805-8679-0e7cf419e69d","order_by":2,"name":"Tong Guo","email":"","orcid":"","institution":"William Harvey Research Institute, Queen Mary University of London, London, United Kingdom","correspondingAuthor":false,"prefix":"","firstName":"Tong","middleName":"","lastName":"Guo","suffix":""},{"id":419107538,"identity":"cf0404cb-d7df-4496-9fbe-94c25b514165","order_by":3,"name":"Eva Sidlauskaite","email":"","orcid":"","institution":"William Harvey Research Institute, Queen Mary University of London, London, United Kingdom","correspondingAuthor":false,"prefix":"","firstName":"Eva","middleName":"","lastName":"Sidlauskaite","suffix":""},{"id":419107539,"identity":"09a556e2-0cf1-40ce-8364-c0bc65073ce5","order_by":4,"name":"Diana Cadena-Castaneda","email":"","orcid":"","institution":"UMR9018CNRS, Institute Gustave Roussy, Villejuif, France.","correspondingAuthor":false,"prefix":"","firstName":"Diana","middleName":"","lastName":"Cadena-Castaneda","suffix":""},{"id":419107540,"identity":"1b3df533-d512-4e83-bd31-8c418cdcf56a","order_by":5,"name":"Jill Richardson","email":"","orcid":"","institution":"Discovery Research MRL UK, MSD, LBIC, London, United Kingdom.","correspondingAuthor":false,"prefix":"","firstName":"Jill","middleName":"","lastName":"Richardson","suffix":""},{"id":419107541,"identity":"f98b61d5-d59d-4b04-8e48-56618d6d0443","order_by":6,"name":"Joana Gomes Neto","email":"","orcid":"","institution":"William Harvey Research Institute, Queen Mary University of London, London, United Kingdom","correspondingAuthor":false,"prefix":"","firstName":"Joana","middleName":"Gomes","lastName":"Neto","suffix":""},{"id":419107542,"identity":"6dfeb868-5ad6-490f-8256-54670f56144c","order_by":7,"name":"Radha Desai","email":"","orcid":"","institution":"Discovery Research MRL UK, MSD, LBIC, London, United Kingdom","correspondingAuthor":false,"prefix":"","firstName":"Radha","middleName":"","lastName":"Desai","suffix":""}],"badges":[],"createdAt":"2025-02-12 13:10:51","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6015469/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6015469/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":78750629,"identity":"4b019738-ccaf-4f86-9aad-56cdc8e27d96","added_by":"auto","created_at":"2025-03-18 11:42:56","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":2396695,"visible":true,"origin":"","legend":"\u003cp\u003eTSPO KO SH-SY5Y cells show reduced NAM A) Cartoon depicting the molecules forming the MCSs in mammalian cells during activation of the mitochondrial retrograde response. B, C) Spinning disc confocal microscopy (SDCM) analysis of mito-nuclear contact sites in WT and TSPO KO cells immunostained with anti-LaminB1 (red) and anti-TOMM20 (green) antibodies, as shown in the representative images (B) and the bar chart (C), mito-nuclear membrane interaction is greatly reduced in TSPO KO cells (scale bar: 10 μm; n=3; N=30-45). D, E) Super-resolution analysis of mito-nuclear interaction in WT and TSPO KO SH-SY5Y cells immunostained as in A; images (D) were produced with direct stochastic optical reconstruction microscopy (dSTORM); MCSs occur at a greater frequency in WT cells, as highlighted by the small yellow dots (co-localization events) present on the nuclear envelope (red) and quantified in (E) (scale bar: 5 μm; n = 3; N = 5). F-H) Transmission electron microscopy (TEM) analysis of mito-nuclear membrane interaction in WT and TSPO KO SH-SY5Y cells; as shown in the representative micrographs (F) and quantified in the corresponding bar charts (G, H), loss of TSPO inhibits NAM formation by increasing the average distance (G) and reducing the occurrence of MCSs (H) between mitochondria and the nucleus (E scale bars = 500 nm; M: mitochondrion; NE: nuclear envelope; CS: contact site; n = 3; N = 20-30). I, J) Proximity ligation assay (PLA) in WT and TSPO KO SH-SY5Y incubated with anti-Lamin B1 and anti-TOMM20 antibodies; representative images (H) show a reduced number of PLA dots (red) around the nucleus (stained with DAPI) in TSPO KO cells; the average number of PLA dots per cell is reported in I (scale bar: 5 μm; n = 3; N = 46-52). K) Representative histocytometric scan of fluorescent immunostained brain tissue. L, M) Plots of the mean staining intensity for cell populations positive to Dapi and MAP2 (i), Dapi+/laminB1\u0026amp;2+ (ii) and Dapi+/TSPO (iii) from human brain tissue sections isolated from Healthy donor (Ctrl) or patients diagnosed with Alzheimer’s Disease (AD) or MotorNeuron Diseases (MND). N) Quantification of cells Dapi+/MAP2-/LaminB1\u0026amp;2+/TSPO+ or Dapi+/MAP2+/LaminB1\u0026amp;2+/ TSPO+ in brain tissues per condition normalised (normalised to Dapi signal) (n=2, N=4).\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-6015469/v1/c641c4ed2d824b784bb45397.png"},{"id":78750627,"identity":"7d1b3dde-7281-4dbc-8f94-e9179658038a","added_by":"auto","created_at":"2025-03-18 11:42:56","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":1438073,"visible":true,"origin":"","legend":"\u003cp\u003eTSPO interacts with the nuclear membrane protein TMPO/LAP2 to define NAM A) Schematic of the working hypothesis of a nucleus-based tethering molecule that allows for NAM formation in neurons. B) Cellular component functional annotation analysis was obtained by submitting the list of 139 unique proteins identified in TSPO-TurboID biotinylated samples to the DAVID database (functional annotation tool). The analysis shows the cellular components that 133 proteins out of 139 (coverage: 95.7%) are involved in. Among them are the mitochondrial and mitochondria-associated compartments, such as the ER, peroxisomes and the nucleus. C) Co-immunoprecipitation assay (TSPO immunoprecipitation and TMPO/LAP2 immunoblotting) in WT SH-SY5Y cells, showing the interaction of TSPO with TMPO, mainly isoform β (LAP2b); each TSPO-interacting TMPO isoform is indicated with an asterisk on the left side of the immunoprecipitation (IP) lane, while black dots on the right side of the IP lane indicate non-specific bands (IgG heavy chain, ~ 45-50 kDa), also present in the negative control (NC, no protein sample); all TSPO present in the input sample (IN) was successfully immunoprecipitated, as indicated by the absence of a protein band in the flow through sample (FT). E, F) Spinning disc confocal microscopy (SDCM) analysis of mito-nuclear contact sites in WT cells down-regulated for TMPO-β (siTMPOβ-1) or transfected with a non-targeting siRNA (siScrm) and immunostained with anti-LaminB1 (green) and anti-TOMM20 (red) antibodies; TMPO-β KD induces a significant reduction in mito-nuclear membrane interaction in WT cells, as shown in (A) and quantified in (B) (scale bar: 5 μm; n = 3; N = 40-43). F-H) Proximity ligation assay (PLA) in WT and TSPO KO SH-SY5Y transfected with either a non-targeting siRNA (siScrm) or a siRNA against TMPO-β (siTMPOβ-1) and stained with antiLamin B1 and anti-TOMM20 antibodies; a reduced number of PLA dots (red) around the nucleus (stained with DAPI) is noticeable in WT cells down-regulated for TMPO (C), while no additive effect can be seen in TSPO KO cells, as quantified in J (scale bar: 5 μm; n = 3; N = 58-59). I) Schematic of the proposed NAM complex featuring TMPO and TSPO as the two ends of the tether.\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-6015469/v1/61ed35609d366cde9b062bdf.png"},{"id":78750630,"identity":"6f3068df-c9dc-4fb0-bd41-7e5ceafb0556","added_by":"auto","created_at":"2025-03-18 11:42:56","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":1403992,"visible":true,"origin":"","legend":"\u003cp\u003eLoss of NAM in TSPO KO cells disrupts nuclear Ca\u003csup\u003e2+\u003c/sup\u003e signalling A, B) Representative recordings of nuclear Ca\u003csup\u003e2+\u003c/sup\u003e levels in WT and TSPO KO SH-SY5Y cells transfected with GCaMP-Xn and stimulated with 100 μM histamine to induce intracellular Ca\u003csup\u003e2+\u003c/sup\u003e signalling; TSPO KO cells are characterised by a significant reduction in histamine-elicited nuclear Ca\u003csup\u003e2+\u003c/sup\u003e transients (C), as quantified in D (n = 6; N = 15-30). C, D) Nuclear Ca\u003csup\u003e2+\u003c/sup\u003e dynamics in cells co-transfected with GCaMP6m-Xn and either TSPOdsRed or dsRed only (control condition); re-expressing TSPO in TSPO KO cells rescues nuclear Ca\u003csup\u003e2+\u003c/sup\u003e uptake, as shown in C (orange dotted line) and quantified in D (n = 3; N = 15- 30). E, F) Analysis of nuclear Ca\u003csup\u003e2+\u003c/sup\u003e transients in WT and TSPO KO SH-SY5Y cells transfected with GCaMP6m-Xn and exposed to 100 μM Dopamine; nuclear Ca\u003csup\u003e2+\u003c/sup\u003e uptake, as shown in E (orange dotted line) and quantified in F (n = 6; N = 15-30). G, H) Analysis of nuclearCa\u003csup\u003e2+\u003c/sup\u003e transients in WT and TSPO KO SH-SY5Y cells co-transfected with GCaMP-Xn and an anti-TMPO-β siRNA, and stimulated with 100 μM histamine to induce intracellular Ca\u003csup\u003e2+\u003c/sup\u003e mobilisation; TMPO-β-down-regulated WT cells show a significant decrease in nuclear Ca\u003csup\u003e2+\u003c/sup\u003e spikes following histamine administration; no additive effect was observed in TSPO KO cells (G, H; orange dotted line; n = 2; N = 10-20). I, J) Histamine-induced mitochondrial Ca\u003csup\u003e2+\u003c/sup\u003e uptake in WT and TSPO KO SH-SY5Y cells transfected with mitochondria-targeted 2mtGCMP6m. TSPO KO cells showed reduced mitochondrial Ca\u003csup\u003e2+\u003c/sup\u003e entry, as depicted in I and quantified in K (n = 6; N = 10-25). K, L) Histamine-induced mitochondrial Ca\u003csup\u003e2+\u003c/sup\u003e uptake in WT and TSPO KO SH-SY5Y cells transfected with mitochondria-targeted 2mtGCaMP6m and stimulated with 100 μM dopamine to induce intracellular Ca\u003csup\u003e2+\u003c/sup\u003e mobilisation; mitochondrial Ca\u003csup\u003e2+\u003c/sup\u003e transients reported in K and quantified in L respectively (n = 6; N = 15-30). M, N) Analysis of mitochondrial membrane potential (ΔYm) in WT and TSPO KO SH-SY5Y cells stained with TMRM (50 nM). As visible from the representative images shown in K, the ΔYm is greatly reduced in TSPO KO cells; quantification of ΔYm in pan-cytoplasmic and perinuclear mitochondria reveals a lower membrane potential in perinuclear mitochondria of WT cells compared to that of pan-cytoplasmic mitochondria, while no differences are noticeable in TSPO KO cells (n = 3). O) Diagram of NAM definition of cellular Ca\u003csup\u003e2+\u003c/sup\u003e homeostasis\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-6015469/v1/a23b1fdb1d7ab5200181c786.png"},{"id":78750631,"identity":"d0a2301d-d97c-47b2-bfff-d44be18cb465","added_by":"auto","created_at":"2025-03-18 11:42:56","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":1592012,"visible":true,"origin":"","legend":"\u003cp\u003eImpaired NAM formation is linked to the loss of Ca\u003csup\u003e2+\u003c/sup\u003e channels in TSPO KO cells A) KEGG pathway enrichment analysis of downregulated differentially expressed genes in TSPO KO SH-SY5T compared to WT cells. Terms are ranked based on the adjusted p-value. Gene count per term is indicated in a colour gradient, and % associated genes per term are indicated by circle size. B) TRRUST analysis of downregulated genes in TSPO KO SH-SY5Y compared to WT cells reveals CREB1 and NFκB as the top transcriptional regulators. Predicted TFs are ranked based on the adjusted p-value. Gene count per TF is indicated in a colour gradient, and % associated genes per term are indicated by circle size. C) RNA-Seq transcriptome analysis showing differentially expressed CREB and NFκB-target genes detected between WT and TSPO KO SH-SY5Y cells; consistently with impaired nuclear Ca\u003csup\u003e2+\u003c/sup\u003e signalling, TSPO ablation results in significantly lower transcription of around 40% target genes for all four Ca\u003csup\u003e2+ \u003c/sup\u003e-regulated mediators of mitochondrial retrograde response (n = 3). D) RNA-Seq transcriptome analysis of genes encoding intracellular Ca\u003csup\u003e2+\u003c/sup\u003e channels regulating mitochondrial, ER and nuclear Ca\u003csup\u003e2+\u003c/sup\u003e fluxes in WT and TSPO KO SH-SY5Y cells (n = 3); the analysis showed a significant reduction in the expression levels of three Ca\u003csup\u003e2+\u003c/sup\u003e channels also residing on the nucleoplasmic reticulum, including RyR3 and ITPR3. E, F) Western blotting analysis of ITPR3 and RyR3 protein levels in the cytosolic and perinuclear fractions of WT and TSPO KO SH-SY5Y cells (E); as quantified in F showing TSPO KO cells to be characterised by reduced levels of both channels, which might contribute to the observed dysfunctional nuclear Ca\u003csup\u003e2+\u003c/sup\u003e signalling (n = 3). G-I) Western blotting analysis of ITPR3 and NF-κB levels in WT and TSPO KO cells transfected with either nucleus-targeted parvalbumin (PV-NLS-GFP) or an empty plasmid (Ctrl) and treated with either 2 mM EGTA or 50 μM BAPTA, AM (4 h); as shown in H and quantified in I, Ca\u003csup\u003e2+\u003c/sup\u003e chelation with either BAPTA, AM or PV-NLS-GFP greatly reduces ITPR3 expression levels; PV-NLS-GFP, but not BAPTA, AM, induces a further decrease in ITPR3 levels in TSPO KO cells; the same conditions have an opposite effect on NF-κB expression, as quantified in I; protein loading control is shown in I (n = 4). J, K) Fluorescence immunocytochemistry (F-ICC) analysis of perinuclear RyR3 levels in SHSY5Y cells following chelation of nuclear Ca\u003csup\u003e2+\u003c/sup\u003e. WT cells were transfected with GFP-tagged, nucleus-targeted parvalbumin (NLS-PV-GFP) for 48 h before immunostaining. As shown in (J), cells expressing NLS-PV-GFP (white arrows) are characterised by a significant reduction in the levels of RyR3 channels (green) coalescing with the nuclear envelope (red), quantified in (K). L) Working model of TSPO-TMPO/LAP2 dependent NAM formation under physiological conditions (i, observed in SH-SY5Y cells) as opposed to the stress-driven, TSPO-AKAP95- mediated pathway (ii).\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-6015469/v1/7944634a573f435ec6908d40.png"},{"id":78752356,"identity":"84060c30-dd89-4ad7-b6d3-42a621a0e163","added_by":"auto","created_at":"2025-03-18 12:07:04","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2311787,"visible":true,"origin":"","legend":"Article File","description":"","filename":"ManuscriptTextFinal.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6015469/v1_covered_e95bdd93-ab23-4f60-a0cd-189e5af90bd5.pdf"},{"id":78750626,"identity":"b2d6b2f2-5060-44da-9004-d793b9329c0b","added_by":"auto","created_at":"2025-03-18 11:42:56","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":69640,"visible":true,"origin":"","legend":"","description":"","filename":"TablesFinalMC.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6015469/v1/cdbe7c95541e47b59888407b.pdf"},{"id":78751033,"identity":"c8213c8a-7152-43a1-95fd-27c9b77faa1b","added_by":"auto","created_at":"2025-03-18 11:50:56","extension":"pdf","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":2927168,"visible":true,"origin":"","legend":"","description":"","filename":"SFiguresFinal.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6015469/v1/af7c3a0783bdcf787c5c9fcf.pdf"}],"financialInterests":"\u003cp\u003eThere is \u003cstrong\u003eNO\u003c/strong\u003e Competing Interest.\u003c/p\u003e\n\u003cp\u003eTables are available in the Supplementary Files section.\u003c/p\u003e","formattedTitle":"\u003cp\u003eNucleus-Tethered Mitochondria are essential for Ca²\u003csup\u003e⁺\u003c/sup\u003e homeostasis\u003c/p\u003e","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"nature-portfolio","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"","title":"Nature Portfolio","twitterHandle":"","acdcEnabled":false,"dfaEnabled":false,"editorialSystem":"ejp","reportingPortfolio":"","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Mitochondria form dynamic membrane contact sites (MCSs), TSPO, TMPO/LAP2, Ca2+ signalling","lastPublishedDoi":"10.21203/rs.3.rs-6015469/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6015469/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eMitochondria form dynamic membrane contact sites (MCSs) with the surrounding organelles, including the nucleus, which we termed ‘nucleus-associated mitochondria’ (NAM).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eIn this manuscript, we report that the stoichiometry of the NAM-forming molecule TSPO is modified in neurodegeneration diseases, and the nucleus-based component of the chromatin- and lamin-binding protein thymopoietin (TMPO/LAP2) is part of the tethering complex.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFurthermore, we show that the NAM-regulated distance between mitochondria and the nucleus is pivotal for the physiological cycling of Ca\u003csup\u003e2+\u003c/sup\u003e in neuronal cells. On this, the transcription of Ca\u003csup\u003e2+\u003c/sup\u003e-regulated genes and those that dictate the spatiotemporal execution of Ca\u003csup\u003e2+\u003c/sup\u003e -such as inositol-1,4,5-trisphosphate (IP3) and ryanodine (RyR) is also dependent.\u003c/p\u003e\n\u003cp\u003eThis data set advances our understanding of the NAM biochemistry and function, shedding light on the mechanisms that govern mitochondrial coupling with the nucleus, aiding the homeostatic transduction of cell signalling.\u003c/p\u003e","manuscriptTitle":"Nucleus-Tethered Mitochondria are essential for Ca²⁺ homeostasis","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-03-18 11:42:51","doi":"10.21203/rs.3.rs-6015469/v1","editorialEvents":[],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"nature-communications","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"NCOMMS","sideBox":"Learn more about [Nature Communications](http://www.nature.com/ncomms/)","snPcode":"","submissionUrl":"https://mts-ncomms.nature.com/","title":"Nature Communications","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"ejp","reportingPortfolio":"Nature Communications","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"5a54bf47-58e9-493a-a26e-45383cc2d4b7","owner":[],"postedDate":"March 18th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[{"id":45845417,"name":"Biological sciences/Cell biology/Cell signalling/Calcium signalling"},{"id":45845418,"name":"Biological sciences/Cell biology/Organelles/Mitochondria"}],"tags":[],"updatedAt":"2025-03-18T11:42:51+00:00","versionOfRecord":[],"versionCreatedAt":"2025-03-18 11:42:51","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6015469","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6015469","identity":"rs-6015469","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}