Emerging dimensions of mitochondrial specialization
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Tslil Ast
Abstract
The diverse, and sometimes opposing, roles of mitochondria require sophisticated organizational and regulatory strategies. This review examines emerging evidence that mitochondria can solve this challenge through functional specialization – adopting distinct bioenergetic and metabolic programs based on location, contacts, and cellular conditions. We discuss both established principles and recent technological breakthroughs that reveal this hidden complexity. Ongoing advances promise to move the field from describing mitochondrial diversity to uncovering its regulatory mechanisms and therapeutic potential.
1 Introduction
Mitochondria, once narrowly characterized as the ‘powerhouse of the cell,’ are now recognized as multifunctional organelles orchestrating cellular homeostasis and stress adaptation. Beyond ATP synthesis, these dynamic structures maintain cellular redox balance, synthesize macromolecular precursors including nucleotides and amino acids, and serve as critical hubs for calcium signaling and apoptotic regulation (Monzel et al. 2023). However, a fundamental question remains: how do mitochondria simultaneously fulfill such varied functional demands without compromising efficiency? One answer lies in their capacity for specialization – rather than functioning as a homogeneous pool, mitochondria can exhibit specialized profiles across multiple organizational scales, from distinct subcellular regions and organellar contact sites to tissue-level spatial gradients. Importantly, this specialization also represents an important strategy for maintaining cellular homeostasis under diverse physiological and stress conditions. While excellent reviews have recently addressed mitochondrial heterogeneity at the tissue-level and across development (Granath-Panelo and Kajimura 2024; Monzel et al. 2023), here we focus on the growing body of evidence for additional scales of specialization that enable this remarkable functional versatility.
2 Historical foundations of mitochondrial heterogeneity
Early studies in striated muscle first demonstrated that mitochondria exist in distinct subpopulations with unique properties in a given cell (Palmer et al. 1977). Subsarcolemmal (SS) and intermyofibrillar (IFM) mitochondria differ in their localization, respiratory control, and substrate preferences. IFM mitochondria, which are embedded among myofibrils, may be better adapted to sustain the energetic demands of muscle contractions through enhanced oxidative phosphorylation (Cogswell et al. 1993). In contrast, SS mitochondria, situated beneath the plasma membrane, have a greater capacity to oxidize lipids via beta-oxidation (Koves et al. 2005). These studies provided the conceptual groundwork for today’s broader recognition of mitochondrial heterogeneity.
3 Spatial partitioning as a strategy for specialization
3.1 Positioning to match local ATP demand
Cells may actively position mitochondria where energy is most needed. In migrating cells, mitochondria are enriched at and power the leading edge, possibly directed there by local energy deprivation cues such as AMPK signaling (Cunniff et al. 2016; Garde et al. 2022; Marlar-Pavey et al. 2025; Schuler et al. 2017). In neurons, presynaptic boutons contain mitochondria whose abundance and morphology scale directly with synaptic activity and ATP demand (Ivannikov et al. 2013; Justs et al. 2022). These examples point to a fundamental principle: mitochondria may be strategically allocated to energy-intense microdomains.
3.2 Organelle contacts and substrate routing
Metabolic specialization also emerges from inter-organelle contacts. Mitochondria form tight contact sites with many organelles, anchoring flux and signals in space rather than relying on diffusion alone (Shai et al. 2018; Valm et al. 2017).
Lipid droplet–bound mitochondria in brown adipocytes have been shown to possess distinct proteomes, cristae morphology, and functions compared to their cytosolic counterparts (Benador et al. 2018). These peri-droplet mitochondria have lower oxidative capacity for fatty acids, and enhanced bioenergetic profiles (with roughly double the respiratory and ATP synthesis rates). Taken together, this raises the possibility that peri-droplet mitochondria support triacylglyceride synthesis and lipid droplet expansion, consistent with a local ATP-supply role at the droplet surface.
Peroxisome–mitochondria contacts provide another archetype of proximity-enabled metabolic specialization. Peroxisomes in contact with mitochondria have been shown to be adjacent to sites of mitochondrial acetyl-CoA synthesis (Cohen et al. 2014), which possibly enables more effective transport of β-oxidation intermediates between these organelles (Shai et al. 2018). Recent work has solidified that these contacts also aid in the maintenance of mitochondrial redox homeostasis, shuttling reactive oxygen species to the peroxisomes (DiGiovanni et al. 2025).
Perhaps the mitochondria’s best studied contact site is with the endoplasmic reticulum, observed over 60 years ago (Copeland and Dalton 1959; Palade 1956). These sites are now appreciated as hotspots of phospholipid and calcium exchange (among other functions) between the two organelles, creating spatially confined microdomains of metabolic control (Csordás et al. 2010; Giacomello et al. 2010; Kawano et al. 2018; Wozny et al. 2023). Together, these architectures emphasize that contacts carve function, positioning mitochondria at organelle interfaces equips them for specialized tasks.
3.3 Sub-mitochondrial specialization
Beyond inter-organelle variation, specialization may occur within a single mitochondrion, creating local hotspots of activity down to the single cristae level. While evidence for this type of specialization remains preliminary, particularly in mammalian systems, high-resolution imaging has revealed that individual cristae can maintain distinct membrane potentials, effectively functioning as semi-independent bioenergetic units (Wolf et al. 2019). Structural elements, such as OPA1, may act as barriers that contribute to this cristae compartmentalization, shaping localized proton motive force and electron transport (Frezza et al. 2006). In yeast analyzed during logarithmic growth, the pyruvate dehydrogenase complex was not uniformly spread across the mitochondrial matrix, but rather localized to foci (Cohen et al. 2014). These findings project heterogeneity inward: a mitochondrion is not merely one uniform metabolic entity in all regards.
4 Adaptive specialization under stress and scarcity
Stress conditions can cause mitochondria to segregate into functionally distinct subpopulations. Recent work demonstrates that during nutrient scarcity, some mitochondria adopt an oxidative ATP-optimized state while others prioritize reductive biosynthesis pathways (Ryu et al. 2024). Specifically, pyrroline-5-carboxylase synthase (P5CS), a key enzyme in proline and ornithine biosynthesis, can assemble into filaments that localize selectively to a subset of mitochondria. These pools of mitochondria, dedicated to reductive biosynthesis, are depleted of the ATP synthase and lack the prototypical cristae structure. Evidence for P5CS filamentation has also been obtained in Drosophila and Arabidopsis (Guo et al. 2024). The ability to create these stable mitochondrial subpopulations is dependent on fission and fusion.
Complementing these findings, recent work has shown that hepatic mitochondria can shift into distinct states under metabolic stress (Kang et al. 2024). Here, nutrient-responsive kinases such as AMPK and mTOR enable coordinated heterogeneity across the network.
These findings suggest that stress does not merely alter flux through existing pathways. Rather, stress actively drives changes in mitochondrial heterogeneity, increasing or decreasing the proportion of specialized subpopulations to enable division of labor under limiting conditions.
5 Methods to dissect specialized subsets
The recognition of mitochondrial specialization has been accelerated by new technologies. Transgenic MITO-Tag mice enable rapid, cell-type-specific purification of mitochondria, revealing proteomic and metabolomic signatures of this organelle otherwise obscured in bulk analyses (Chen et al. 2016). Proximity labeling methods such as APEX2 or split-TurboID allow selective profiling of mitochondria engaged in particular organelle contacts (Cho et al. 2020). The combination of rapid mitochondrial isolation and proximity labeling has recently been applied to assay endosome- and lipid droplet-associated mitochondria, revealing a proteomic signature distinct from global mitochondria (Cunningham et al. 2025). Looking forward, the development and application of genetically encoded mitochondrial reporters holds great promise for resolving spatial heterogeneity (Choe and Titov 2022). This ever-expanding toolbox can monitor factors such as mitochondrial NAD+/NADH, reactive oxygen burden, pH, calcium levels, and nutrient availability. Such sensors enable real-time monitoring of metabolic dynamics within distinct mitochondrial subpopulations in living cells and tissues. Collectively, these tools provide the resolution necessary to see specialization that bulk approaches once averaged away.
6 Conclusions and future directions
Mitochondrial specialization spans multiple organizational layers – from cristae microdomains to organelle positioning within cells to zonated tissue architectures. This division of labor enables cells and tissues to match metabolic supply with local demand and to flexibly adapt under stress. Key open questions include: How stable are specialized subpopulations over time? What molecular regulators enforce these identities? And can interventions that reprogram or selectively target specialized mitochondria be harnessed in disease therapy? For example, distinct mitochondrial subpopulations may represent untapped vulnerabilities in metabolic disorders, cancer, and neurodegenerative diseases. Identifying these specialized subpopulations and the signaling pathways that maintain them could open new avenues for therapeutic intervention. With spatially resolved omics, advanced imaging, and refined genetic tools, the next decade promises to move the field from recognizing heterogeneity to mechanistically dissecting and manipulating it.
Acknowledgments
T.A. is a recipient of the Alon scholarship and the incumbent of the Leonard and Carol Berall Career Development Chair.
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Research ethics: Not applicable.
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Informed consent: Not applicable.
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Author contributions: The author has accepted responsibility for the entire content of this manuscript and approved its submission.
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Use of Large Language Models, AI and Machine Learning Tools: None declared.
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Conflict of interest: The author states no conflict of interest.
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Research funding: This work was supported by a European Research Council (ERC) Starting Grant (FeSurveil 101163884).
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Data availability: Not applicable.
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