Additional ATP Research Articles

Pi-based biochemical mechanism of endurance-training-induced improvement of running performance in humans.

Endurance training improves running performance in distances where oxidative phosphorylation (OXPHOS) is the main ATP source. Here, a dynamic computer model is used to assess possible biochemical mechanisms underlying this improvement. The dynamic computer model is based on the "Pi double-threshold" mechanism of muscle fatigue, according to which the additional ATP usage appears when (1) inorganic phosphate (Pi) exceeds a critical value (Picrit); (2) exercise is terminated because of fatigue, when Pi reaches a peak value (Pipeak); (3) the Pi increase and additional ATP usage increase mutually stimulate each other. The endurance-training-induced increase in oxidative phosphorylation (OXPHOS) activity attenuates the reaching of Pipeak by Pi (and thus of O2max by O2) at increased power output. This in turn allows a greater work intensity, and thus higher speed, to be achieved before exercise is terminated because of fatigue at the end of the 1500m run. Thus, identical total work is performed in a shorter time. Probably, endurance training also lowers Pipeak, which improves the homeostasis of "bioenergetic" muscle metabolites: ADP, PCr, Pi and H+ ions. The present dynamic computer model generates clear predictions of metabolic changes that limit performance during 1500m running. It contributes to our mechanistic understanding of training-induced improvement in running performance and stimulates further physiological experimental studies.

Effect of Free Cysteine Residues to Serine Mutation on Cellodextrin Phosphorylase.

Cellodextrin phosphorylase (CDP) plays a key role in energy-efficient cellulose metabolism of anaerobic bacteria by catalyzing phosphorolysis of cellodextrin to produce cellobiose and glucose 1-phosphate, which can be utilized for glycolysis without consumption of additional ATP. As the enzymatic phosphorolysis reaction is reversible, CDP is also employed to produce cellulosic materials in vitro. However, the enzyme is rapidly inactivated by oxidation, which hinders in vitro utilization in aerobic environments. It has been suggested that the cysteine residues of CDP, which do not form disulfide bonds, are responsible for the loss of activity, and the aim of the present work was to test this idea. For this purpose, we replaced all 11 free cysteine residues of CDP from Acetivibrio thermocellus (formerly known as Clostridium thermocellum) with serine, which structurally resembles cysteine in our previous work. Herein, we show that the resulting CDP variant, named CDP-CS, has comparable activity to the wild-type enzyme, but shows increased stability to oxidation during long-term storage. X-Ray crystallography indicated that the mutations did not markedly alter the overall structure of the enzyme. Ensemble refinement of the crystal structures of CDP and CDP-CS indicated that the C372S and C625S mutations reduce structural fluctuations in the protein main chain, which may contribute to the increased stability of CDP-CS to oxidation.

Alternative electron pathways of photosynthesis power green algal CO2 capture.

Microalgae contribute to about half of global net photosynthesis, which converts sunlight into the chemical energy (ATP and NADPH) used to transform CO2 into biomass. Alternative electron pathways of photosynthesis have been proposed to generate additional ATP that is required to sustain CO2 fixation. However, the relative importance of each alternative pathway remains elusive. Here, we dissect and quantify the contribution of cyclic, pseudo-cyclic, and chloroplast-to-mitochondrion electron flows for their ability to sustain net photosynthesis in the microalga Chlamydomonas reinhardtii. We show that (i) each alternative pathway can provide sufficient additional energy to sustain high CO2 fixation rates, (ii) the alternative pathways exhibit cross-compensation, and (iii) the activity of at least one of the three alternative pathways is necessary to sustain photosynthesis. We further show that all pathways have very different efficiencies at energizing CO2 fixation, with the chloroplast-mitochondrion interaction being the most efficient. Overall, our data lay bioenergetic foundations for biotechnological strategies to improve CO2 capture and fixation.

ALPL regulates pro-angiogenic capacity of mesenchymal stem cells through ATP-P2X7 axis controlled exosomes secretion

BackgroundEarly-onset bone dysplasia is a common manifestation of hypophosphatasia (HPP), an autosomal inherited disease caused by ALPL mutation. ALPL ablation induces prototypical premature bone ageing characteristics, resulting in impaired osteogenic differentiation capacity of human bone marrow mesenchymal stem cells (hBMMSCs). As angiogenesis is tightly coupled with osteogenesis, it also plays a necessary role in sustaining bone homeostasis. We have previously observed a decrease in expression of angiogenesis marker gene CD31 in the metaphysis of long bone in Alpl+/− mice. However, the role of ALPL in regulation of angiogenesis in bone has remained largely unknown.MethodsExosomes derived from Normal and HPP hBMMSCs were isolated and identified by ultracentrifugation, transmission electron microscopy, and nanoparticle size measurement. The effects of ALPL on the angiogenic capacity of hBMMSCs from HPP patients were assessed by immunofluorescence, tube formation, wound healing and migration assay. exo-ELISA and Western Blot were used to evaluate the exosomes secretion of hBMMSCs from HPP, and the protein expression of VEGF, PDGFBB, Angiostatin and Endostatin in exosomes respectively.ResultsWe verified that ALPL ablation resulted in impaired pro-angiogenic capacity of hBMMSCs, accounting for reduced migration and tube formation of human umbilical vein endothelial cells, as the quantities and proteins composition of exosomes varied with ALPL expression. Mechanistically, loss of function of ALPL enhanced ATP release. Additional ATP, in turn, led to markedly elevated level of ATP receptor P2X7, which consequently promoted exosomes secretion, resulting in a decreased capacity to promote angiogenesis. Conversely, inhibition of P2X7 increased the angiogenic induction capacity by preventing excessive release of anti-angiogenic exosomes in ALPL deficient-hBMMSCs.ConclusionThe ALPL–ATP axis regulates the pro-angiogenic ability of hBMMSCs by controlling exosomes secretion through the P2X7 receptor. Thus, P2X7 may be proved as an effective therapeutic target for accelerating neovascularization in ALPL–deficient bone defects.

Walking the 'design-build-test-learn' cycle: flux analysis and genetic engineering reveal the pliability of plant central metabolism.

This article is a Commentary on Morley et al. (2023), 239: 1834–1851.

The negative impact of shade on photosynthetic efficiency in sugarcane may reflect a metabolic bottleneck

Plants with C4 metabolism normally have higher photosynthetic rates than C3 ones. As a result, several of the most productive species known are NADP-ME C4 grasses, such as sugarcane and maize. However, the advantages of the C4 cycle are most evident under high light as the CO2 concentrating mechanism comes at the expense of additional ATP. Recent works have suggested a negative impact of shading across NADP-ME C4 grasses, causing a downregulation of maximal quantum efficiency of CO2 assimilation (ϕCO2,max). The mechanisms behind the loss in photosynthetic efficiency and whether these results apply for other C4 crops species and within germplasm of a species remain unclear. We analysed the photosynthetic acclimation to shade in four sugarcane genotypes with contrasting yield. To find out whether the effects of leaf history, i.e., shading a leaf developed under full sunlight and exposed later on to shade differ from a leaf fully developed under shading, these two types of leaves were evaluated. Shaded sugarcane plants showed decreased CO2 assimilation efficiency compared to plants grown under full sunlight. Based on the fluorescence measurements, it seems that this reduction coincided with a more reduced QA redox state, which could point to a metabolic limitation downstream of the light-dependent reactions. The results were similar for all genotypes and were observed regardless of whether leaves developed under shade or under full sunlight conditions and exposed subsequently to shade, suggesting that light is the main factor affecting photosynthetic efficiency. This study reinforces the notion that this negative impact of shade could reflect a common bottleneck across NADP-ME C4 grasses.

V̇O2 (non-)linear increase in ramp-incremental exercise vs. V̇O2 slow component in constant-power exercise: Underlying mechanisms

A computer model of the skeletal muscle bioenergetic system involving the Pi double-threshold mechanism of muscle fatigue was used to study the V̇O2 (non-)linear increase in time in ramp-incremental exercise as compared to the V̇O2 slow component in constant-power exercise. The Pi double-threshold mechanism applies to both constant-power and ramp-incremental exercise. The additional ATP usage is initiated at a significantly higher ATP usage activity (power output), determining the moderate/heavy exercise border, in ramp-incremental, than in constant-power exercise. A significantly lowered additional ATP usage activity or elevated glycolysis stimulation at the highest power outputs in ramp-incremental exercise in relation to constant-power exercise can additionally explain the much smaller (or zero) V̇O2 non-linearity in ramp-incremental exercise, than V̇O2 slow component in constant-power exercise. The V̇O2 (non-)linearity in ramp-incremental exercise and V̇O2 slow component in constant-power exercise is a derivative of a balance between the additional ATP usage and ATP production by anaerobic glycolysis.

Two cyclic electron flows around photosystem I differentially participate in C4 photosynthesis.

C4 plants assimilate CO2 more efficiently than C3 plants because of their C4 cycle that concentrates CO2. However, the C4 cycle requires additional ATP molecules, which may be supplied by cyclic electron flow (CEF) around photosystem I. One CEF route, which depends on a chloroplast NADH dehydrogenase-like (NDH) complex, is suggested to be crucial for C4 plants despite the low activity in C3 plants. The other route depends on proton gradient regulation 5 (PGR5) and PGR5-like photosynthetic phenotype 1 (PGRL1) and is considered a major CEF route to generate the proton gradient across the thylakoid membrane in C3 plants. However, its contribution to C4 photosynthesis is still unclear. In this study, we investigated the contribution of the two CEF routes to the NADP-malic enzyme subtype of C4 photosynthesis in Flaveria bidentis. We observed that suppressing the NDH-dependent route drastically delayed growth and decreased the CO2 assimilation rate to approximately 30% of the wild-type rate. On the other hand, suppressing the PGR5/PGRL1-dependent route did not affect plant growth and resulted in a CO2 assimilation rate that was approximately 80% of the wild-type rate. Our data indicate that the NDH-dependent CEF substantially contributes to the NADP-malic enzyme subtype of C4 photosynthesis and that the PGR5/PGRL1-dependent route cannot complement the NDH-dependent route in F. bidentis. These findings support the fact that during C4 evolution, photosynthetic electron flow may have been optimized to provide the energy required for C4 photosynthesis.

Identification of a chemical probe for lipid kinase phosphatidylinositol-5-phosphate 4-kinase gamma (PI5P4Kγ)

Phosphatidylinositol-5-phosphate 4-kinase gamma (PI5P4Kγ), which phosphorylates phosphatidylinositol-5-monophosphate (PI(5)P), is a human lipid kinase with intriguing roles in inflammation, T cell activation, autophagy regulation, immunity, heart failure, and several cancers. To provide a high-quality chemical tool that would enable additional characterization of this protein, we designed and evaluated a potent, selective, and cell-active inhibitor of human PI5P4Kγ. We describe the use of the PI5P4Kγ NanoBRET assay to generate structure–activity relationships (SAR), support chemical probe (2) design, and identify a structurally related negative control (4). We have characterized the binding of our chemical probe to PI5P4Kγ using orthogonal assay formats reliant on competition with an ATP-competitive reagent. Based on our results in these assays, additional ATP titration studies, and published co-crystal structures with structurally related compounds, we hypothesize that 2 binds in the ATP active site of PI5P4Kγ. Kinome-wide profiling complemented by further off-target screening confirmed the selectivity of both our chemical probe and negative control. When a breast cancer cell line (MCF-7) was treated with compound 2, increased mTORC1 signaling was observed, demonstrating that efficacious binding of 2 to PI5P4Kγ in cells results in activation of a negative feedback loop also reported in PI5P4Kγ knockout mice.

Regulation of light energy conversion between linear and cyclic electron flow within photosystem II controlled by the plastoquinone/quinol redox poise.

Ultrapurified Photosystem II complexes crystalize as uniform microcrystals (PSIIX) of unprecedented homogeneity that allow observation of details previously unachievable, including the longest sustained oscillations of flash-induced O2 yield over > 200 flashes and a novel period-4.7 water oxidation cycle. We provide new evidence for a molecular-based mechanism for PSII-cyclic electron flow that accounts for switching from linear to cyclic electron flow within PSII as the downstream PQ/PQH2 pool reduces in response to metabolic needs and environmental input. The model is supported by flash oximetry of PSIIX as the LEF/CEF switch occurs, Fourier analysis of O2 flash yields, and Joliot-Kok modeling. The LEF/CEF switch rebalances the ratio of reductant energy (PQH2) to proton gradient energy (H+o/H+i) created by PSII photochemistry. Central to this model is the requirement for a regulatory site (QC) with two redox states in equilibrium with the dissociable secondary electron carrier site QB. Both sites are controlled by electrons and protons. Our evidence fits historical LEF models wherein light-driven water oxidation delivers electrons (from QA-) and stromal protons through QB to generate plastoquinol, the terminal product of PSII-LEF in vivo. The new insight is the essential regulatory role of QC. This site senses both the proton gradient (H+o/H+i) and the PQ pool redox poise via e-/H+ equilibration with QB. This information directs switching to CEF upon population of the protonated semiquinone in the Qc site (Q-H+)C, while the WOC is in the reducible S2 or S3 states. Subsequent photochemical primary charge separation (P+QA-) forms no (QH2)B, but instead undergoes two-electron backward transition in which the QC protons are pumped into the lumen, while the electrons return to the WOC forming (S1/S2). PSII-CEF enables production of additional ATP needed to power cellular processes including the terminal carboxylation reaction and in some cases PSI-dependent CEF.

Teleostean fishes may have developed an efficient Na+ uptake for adaptation to the freshwater system.

Understanding Na+ uptake mechanisms in vertebrates has been a research priority since vertebrate ancestors were thought to originate from hyperosmotic marine habitats to the hypoosmotic freshwater system. Given the evolutionary success of osmoregulator teleosts, these freshwater conquerors from the marine habitats are reasonably considered to develop the traits of absorbing Na+ from the Na+-poor circumstances for ionic homeostasis. However, in teleosts, the loss of epithelial Na+ channel (ENaC) has long been a mystery and an issue under debate in the evolution of vertebrates. In this study, we evaluate the idea that energetic efficiency in teleosts may have been improved by selection for ENaC loss and an evolved energy-saving alternative, the Na+/H+ exchangers (NHE3)-mediated Na+ uptake/NH4 + excretion machinery. The present study approaches this question from the lamprey, a pioneer invader of freshwater habitats, initially developed ENaC-mediated Na+ uptake driven by energy-consuming apical H+-ATPase (VHA) in the gills, similar to amphibian skin and external gills. Later, teleosts may have intensified ammonotelism to generate larger NH4 + outward gradients that facilitate NHE3-mediated Na+ uptake against an unfavorable Na+ gradient in freshwater without consuming additional ATP. Therefore, this study provides a fresh starting point for expanding our understanding of vertebrate ion regulation and environmental adaptation within the framework of the energy constraint concept.

Sensitivity of V̇O2max, critical power and V̇O2 on-kinetics to O2 concentration/delivery and other factors in skeletal muscle

Relative sensitivities of the maximal oxygen uptake (V̇O2max), critical power (CP) and transition time of the primary phase II of the V̇O2 on-kinetics (t0.63) to selected factors in skeletal muscle are simulated using a computer model of the skeletal muscle bioenergetic system. In normoxia, V̇O2max is significantly positively sensitive to peak Pi (Pipeak, Pi at which exercise is terminated because of fatigue), OXPHOS (oxidative phosphorylation) activity (kOX) and ESA (each-step-activation) intensity (AOXmax), and negatively sensitive to the accessible phosphate (and total creatine) pool (Pacc). CP is additionally moderately positively sensitive to critical Pi (Picrit, Pi at which the additional ATP usage appears) and negatively sensitive to the additional ATP usage activity (kadd). t0.63 is significantly positively sensitive to Pacc and negatively to kOX and AOXmax. The positive sensitivity of V̇O2max and CP, and negative sensitivity of t0.63 to O2 concentration/delivery is low at normoxic and hyperoxic O2 levels, but increases significantly at severely hypoxic O2 levels.

Skeletal muscle biochemical origin of exercise intensity domains and their relation to whole-body V̇O2 kinetics.

This article presents the biochemical intra-skeletal-muscle basis of exercise intensity domains: moderate (M), heavy (H), very heavy (VH) and severe (S). Threshold origins are mediated by a ‘Pi double-threshold’ mechanism of muscle fatigue, which assumes (1) additional ATP usage, underlying muscle V̇O2 and metabolite slow components, is initiated when inorganic phosphate (Pi) exceeds a critical value (Picrit); (2) exercise is terminated because of fatigue, when Pi reaches a peak value (Pipeak); and (3) the Pi increase and additional ATP usage increase mutually stimulate each other forming a positive feedback. M/H and H/VH borders are defined by Pi on-kinetics in relation to Picrit and Pipeak. The values of the ATP usage activity, proportional to power output (PO), for the M/H, H/VH and VH/S borders are lowest in untrained muscle and highest in well-trained muscle. The metabolic range between the M/H and H/VH border (or ‘H space’) decreases with muscle training, while the difference between the H/VH and VH/S border (or ‘VH space’) is only weakly dependent on training status. The absolute magnitude of the muscle V̇O2 slow-component, absent in M exercise, rises gradually with PO to a maximal value in H exercise, and then decreases with PO in VH and S exercise. Simulations of untrained, physically active and well-trained muscle demonstrate that the muscle M/H border need not be identical to the whole-body M/H border determined from pulmonary V̇O2 on-kinetics and blood lactate, while suggesting that the biochemical origins of the H/VH border reside within skeletal muscle and correspond to whole-body critical power.

Which UV wavelength is the most effective for chlorine-resistant bacteria in terms of the impact of activity, cell membrane and DNA?

The widespread use of chlorine disinfectants in drinking water has accelerated the selection of chlorine-resistant bacteria (CRB). The presence of CRB in drinking water (during treatment, distribution, and consumption) has a profound impact on public safety. Ultraviolet (UV) irradiation is an effective control technology for CRB. Nevertheless, the relative efficacies of different UV light wavelengths (traditional mercury lamps, ultraviolet light-emitting diodes (UV-LED 265 nm and 285 nm), and 222 nm krypton-chlorine excilamps) in the control and inactivation of various CRB have not been investigated. In this study, the impacts of different UV light sources on the inactivation rate and microbial cell structure of CRB were evaluated. The results showed that the order of inactivation efficacy was as follows: UV-LED 265 nm > low-pressure ultraviolet (LPUV) ≈ medium-pressure ultraviolet (MPUV) ≈ 222 nm > UV-LED 285 nm. CRB was inactivated via DNA damage and additional ATP decline under UV-LED 265 nm, UV-LED 285 nm and LPUV irradiation, of which UV-LED 265 nm was the most effective based on fluence and inactivation rate constants. The inactivation mechanisms revealed that the 222 nm wavelength was significantly different from other wavelengths; it stimulated the production of a large number of reactive oxygen species (ROS) which resulted in bacterial cell membrane damage, ATP direct decline, and a small number of DNA lesions. Multispectral MPUV induced both of the foregoing mechanisms but mainly damaged DNA. The present study provides empirical evidence for the reasonable application of UV disinfection technology in the treatment of water and wastewater and their conduits and delivery systems.

Dissection of respiratory and cyclic electron transport in Synechocystis sp. PCC 6803.

Cyclic electron transport (CET) is an attractive hypothesis for regulating photosynthetic electron transport and producing the additional ATP in oxygenic phototrophs. The concept of CET has been established in the last decades, and it is proposed to function in the progenitor of oxygenic photosynthesis, cyanobacteria. The in vivo activity of CET is frequently evaluated either from the redox state of the reaction center chlorophyll in photosystem (PS) I, P700, in the absence of PSII activity or by comparing PSI and PSII activities through the P700 redox state and chlorophyll fluorescence, respectively. The evaluation of CET activity, however, is complicated especially in cyanobacteria, where CET shares the intersystem chain, including plastoquinone, cytochrome b6/f complex, plastocyanin, and cytochrome c6, with photosynthetic linear electron transport (LET) and respiratory electron transport (RET). Here we sought to distinguish the in vivo electron transport rates in RET and CET in the cyanobacterium Synechocystis sp. PCC 6803. The reduction rate of oxidized P700 (P700+) decreased to less than 10% when PSII was inhibited, indicating that PSII is the dominant electron source to PSI but P700+ is also reduced by electrons derived from other sources. The oxidative pentose phosphate (OPP) pathway functions as the dominant electron source for RET, which was found to be inhibited by glycolaldehyde (GA). In the condition where the OPP pathway and respiratory terminal oxidases were inhibited by GA and KCN, the P700+ reduction rate was less than 1% of that without any inhibitors. This study indicate that the electron transport to PSI when PSII is inhibited is dominantly derived from the OPP pathway in Synechocystis sp. PCC 6803.

Biobased PET from lignin using an engineered cis, cis-muconate-producing Pseudomonas putida strain with superior robustness, energy and redox properties

Polyethylene terephthalate (PET), the most common synthetic polyester today, is largely produced from fossil resources, contributing to global warming. Consequently, sustainable sources must be developed to meet the increasing demand for this useful polymer. Here, we demonstrate a cascaded value chain that provides green PET from lignin, the world's most underutilized renewable, via fermentative production of cis, cis-muconate (MA) from lignin-based aromatics as a central step. Catechol, industrially the most relevant but apparently also a highly toxic lignin-related aromatic, strongly inhibited MA-producing Pseudomonas putida MA-1. Assessed by 13C metabolic flux analysis, the microbe substantially redirected its carbon core fluxes, resulting in enhanced NADPH supply for stress defense but causing additional ATP costs. The reconstruction of MA production in a genome-reduced P. putida chassis yielded novel producers with superior pathway fluxes and enhanced robustness to catechol and a wide range of other aromatics. Using the advanced producer P. putida MA-10 catechol, MA could be produced in a fed-batch process from catechol (plus glucose as additional growth substrate) up to an attractive titer of 74 g L−1 and a space-time-yield of 1.4 g L−1 h−1. In terms of co-consumed sugar, the further streamlined strain MA-11 achieved the highest yield of 1.4 mol MA (mol glucose)−1, providing a striking economic advantage. Following fermentative production, bio-based MA was purified and used to chemically synthetize the PET monomer terephthalic acid and the comonomer diethylene glycol terephthalic acid through five steps, which finally enabled the first green PET from lignin.

Low Cancer Incidence in Naked Mole-Rats May Be Related to Their Inability to Express the Warburg Effect.

Metabolic flexibility in mammals enables stressed tissues to generate additional ATP by converting large amounts of glucose into lactic acid; however, this process can cause transient local or systemic acidosis. Certain mammals are adapted to extreme environments and are capable of enhanced metabolic flexibility as a specialized adaptation to challenging habitat niches. For example, naked mole-rats (NMRs) are a fossorial and hypoxia-tolerant mammal whose metabolic responses to environmental stressors markedly differ from most other mammals. When exposed to hypoxia, NMRs exhibit robust hypometabolism but develop minimal acidosis. Furthermore, and despite a very long lifespan relative to other rodents, NMRs have a remarkably low cancer incidence. Most advanced cancers in mammals display increased production of lactic acid from glucose, irrespective of oxygen availability. This hallmark of cancer is known as the Warburg effect (WE). Most malignancies acquire this metabolic phenotype during their somatic evolution, as the WE benefits tumor growth in several ways. We propose that the peculiar metabolism of the NMR makes development of the WE inherently difficult, which might contribute to the extraordinarily low cancer rate in NMRs. Such an adaptation of NMRs to their subterranean environment may have been facilitated by modified biochemical responses with a stronger inhibition of the production of CO2 and lactic acid by a decreased extracellular pH. Since this pH-inhibition could be deeply hard-wired in their metabolic make-up, it may be difficult for malignant cells in NMRs to acquire the WE-phenotype that facilitates cancer growth in other mammals. In the present commentary, we discuss this idea and propose experimental tests of our hypothesis.

CRAC channel inhibitors in pancreatic pathologies.

CRAC channel inhibitors in pancreatic pathologies.

Mechanisms of the effect of oxidative phosphorylation deficiencies on the skeletal muscle bioenergetic system in patients with mitochondrial myopathies.

Simulations carried out using a previously developed model of the skeletal muscle bioenergetic system, involving the "inorganic phosphate (Pi) double-threshold" mechanism of muscle fatigue, lead to the conclusion that a decrease in the oxidative phosphorylation (OXPHOS) activity, caused by mutations in mitochondrial or nuclear DNA, is the main mechanism underlying the changes in the kinetic properties of the system in mitochondrial myopathies (MM). These changes generally involve the very-heavy-exercise-like behavior and exercise termination because of fatigue at low work intensities. In particular, a sufficiently large (at a given work intensity) decrease in OXPHOS activity leads to slowing of the primary phase II of the oxygen uptake (V̇o2) on-kinetics, decrease in maximal V̇o2 (V̇o2max), appearance of the slow component of the V̇o2 on-kinetics, exercise intolerance, and lactic acidosis at relatively low power outputs encountered in experimental studies in patients with MM. Thus, the "Pi double-threshold" mechanism of muscle fatigue is able to account, at least semiquantitatively, for various kinetic effects of inborn OXPHOS deficiencies of the skeletal muscle bioenergetic system. Exercise can be potentially lengthened and V̇o2max elevated in patients with MM through an increase in peak Pi (Pipeak), at which exercise is terminated because of fatigue. Generally, a mechanism underlying the kinetic effects of OXPHOS deficiencies on the skeletal muscle bioenergetic system in MM is proposed that was absent in the literature.NEW & NOTEWORTHY A mechanism of the OXPHOS deficiencies-induced changes of the skeletal muscle bioenergetic system in patients with mitochondrial myopathies (MM), namely, appearance of the slow component of the V̇o2 on-kinetics at relatively low work intensities, slowed primary phase II of the V̇o2 on-kinetics, lowered V̇o2max, and lactic acidosis is proposed. It involves a decrease in OXPHOS activity acting through the "Pi double-threshold" mechanism of muscle fatigue comprising initiation of the additional ATP usage and termination of exercise.

Exploiting the metabolic energy demands of drug efflux pumps provides a strategy to overcome multidrug resistance in cancer

BackgroundP-glycoprotein (P-gp) is a prevalent resistance mediator and it requires considerable cellular energy to ensure ATP dependent efflux of anticancer drugs. The glycolytic pathway generates the majority of catabolic energy in cancer cells; however, the high rates of P-gp activity places added strain on its inherently limited capacity to generate ATP. This is particularly relevant for compounds such as verapamil that are believed to trap P-gp in a futile transport process that requires continuing ATP consumption. Ultimately, this leads to cell death and the hypersensitivity of resistant cells to verapamil is termed collateral sensitivity. ResultsWe show that the addition of verapamil to resistant cells produces a prominent reduction in ATP levels that supports the idea of disrupted energy homeostasis. Even in the absence of verapamil, P-gp expressing cells display near maximal rates of glycolysis and oxidative phosphorylation, which prevents an adequate response to the demand for ATP to sustain transport activity. Moreover, the near perpetually maximal rate of oxidative phosphorylation in the presence of verapamil resulted in elevated levels of reactive oxygen species that affect cell survival and underscore collateral sensitivity. ConclusionsOur results demonstrate that the strained metabolic profiles of P-gp expressing resistant cancer cells can be overwhelmed by additional ATP demands. General significanceConsequently, collateral sensitising drugs may overcome the resistant phenotype by exploiting, rather than inhibiting, the energy demanding activity of pumps such as P-gp.