Decreased, mitochondrial oxidative phosphorylation

Mitochondrial oxidative phosphorylation (OXPHOS) is a fundamental cellular process to generate most of ATP to supplies energy for essential cellular functions.  The decrease of OXPHOS can be due to the inhibition or impairment of one or more components of the ETC or ATP synthase, loss of membrane integrity leading to uncoupling of electron transport from proton translocation, limited availability of electron donors (e.g., NADH, FADH₂), reduced metabolic substrate supply due to diminished photosynthetic carbon fixation and associated energy imbalance in photosynthetic cells, or oxygen limitation, or structural damage to mitochondrial membranes. As a consequence, the ability of mitochondria to maintain proton motive force and to synthesize ATP is reduced.

A decrease in mitochondrial oxidative phosphorylation (OXPHOS) can be measured using a combination of functional, biochemical, and structural approaches that collectively assess mitochondrial respiratory capacity, ATP production, and integrity of the electron transport system. Functional impairment of OXPHOS is most commonly evaluated by respirometry, where oxygen consumption rates are measured in intact or permeabilized cells, tissues, or isolated mitochondria under controlled substrate and inhibitor conditions. High-resolution respirometry allows quantification of basal respiration, ADP-stimulated (ATP-linked) respiration, maximal electron transport system capacity, and coupling efficiency. A reduction in ADP-stimulated respiration or maximal respiratory capacity provides direct evidence for decreased mitochondrial oxidative phosphorylation (Djafarzadeh and Jakob, 2017; Coulson, Duffy and Staples, 2024).

Complementary evidence for reduced OXPHOS can be obtained by quantifying ATP synthesis rates. Luciferase-based assays enable sensitive measurement of ATP production in real time or at defined endpoints and are widely used to assess mitochondrial ATP output. Reduced ATP synthesis, particularly when observed alongside diminished oxygen consumption, indicates impaired coupling between electron transport and phosphorylation (Lundin, Rickardsson and Thore, 1976; Coulson, Duffy and Staples, 2024). In addition, in vivo ³¹P nuclear magnetic resonance (NMR) spectroscopy provides a non-invasive assessment of high-energy phosphate metabolites, allowing evaluation of cellular energetic status and supporting identification of reduced mitochondrial ATP generation (Hitchins, Cieslar and Dobson, 2001).

Biochemical characterization of OXPHOS impairment can be further refined through enzyme activity assays of individual respiratory chain complexes (I–IV). These assays measure the catalytic activity of specific complexes using defined substrates and inhibitors and can identify whether reduced oxidative phosphorylation is associated with inhibition or dysfunction of discrete components of the electron transport chain (Coulson, Duffy and Staples, 2024). Changes in mitochondrial membrane potential (ΔΨm), assessed using potentiometric fluorescent probes, provide additional functional insight, as loss or reduction of ΔΨm reflects impaired proton motive force and compromised capacity for ATP synthesis (Xie et al., 2019).

Finally, blue-native polyacrylamide gel electrophoresis (BN-PAGE) is used to examine the structural organization, abundance, and stability of intact OXPHOS complexes and supercomplexes within the inner mitochondrial membrane. Alterations in complex assembly or loss of specific respiratory complexes detected by BN-PAGE support a mechanistic interpretation of reduced oxidative phosphorylation at the protein organization level (Yan and Forster, 2009). Together, these measurement approaches provide robust and complementary lines of evidence for identifying and characterizing decreases in mitochondrial oxidative phosphorylation.

The Key Event Decrease in mitochondrial oxidative phosphorylation is broadly applicable across biological systems that rely on mitochondria for aerobic energy production. This KE is relevant to eukaryotic organisms, including animals, plants, algae, and fungi, as the core molecular machinery of mitochondrial oxidative phosphorylation—comprising the electron transport chain (Complexes I–IV), ATP synthase (Complex V), and the proton motive force across the inner mitochondrial membrane—is highly conserved across taxa. Consequently, the KE can be applied to a wide range of model and non-model species used in ecotoxicology, physiology, and environmental risk assessment.

The KE is applicable across life stages, but sensitivity may vary. Early developmental stages, growth phases, reproduction, and energetically demanding physiological processes (e.g. molting, metamorphosis, active growth, and stress responses) are expected to be particularly vulnerable to perturbations in mitochondrial ATP production. In photosynthetic organisms, this KE is also applicable under both light and dark conditions, reflecting the metabolic integration of mitochondrial respiration with photosynthetic carbon metabolism.

In terms of stressors, the domain of applicability includes chemical, physical, and environmental factors that impair mitochondrial function. These include direct inhibitors of respiratory chain complexes or ATP synthase, uncouplers of oxidative phosphorylation, agents that damage mitochondrial membranes or proteins, stressors that disrupt cellular redox balance, and conditions that limit substrate or oxygen availability. In photosynthetic organisms, stressors that reduce photosynthesis can indirectly contribute to this KE by decreasing the supply of carbon substrates and reducing equivalents that support mitochondrial respiration, thereby altering cellular energy balance.

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