Mitochondrial Ca2+ uptake through the Ca2+ uniporter supports cell functions, including oxidative metabolism, while meeting tissue-specific calcium signaling patterns and energy needs. The molecular ...mechanisms underlying tissue-specific control of the uniporter are unknown. Here, we investigated a possible role for tissue-specific stoichiometry between the Ca2+-sensing regulators (MICUs) and pore unit (MCU) of the uniporter. Low MICU1:MCU protein ratio lowered the Ca2+ threshold for Ca2+ uptake and activation of oxidative metabolism but decreased the cooperativity of uniporter activation in heart and skeletal muscle compared to liver. In MICU1-overexpressing cells, MICU1 was pulled down by MCU proportionally to MICU1 overexpression, suggesting that MICU1:MCU protein ratio directly reflected their association. Overexpressing MICU1 in the heart increased MICU1:MCU ratio, leading to liver-like mitochondrial Ca2+ uptake phenotype and cardiac contractile dysfunction. Thus, the proportion of MICU1-free and MICU1-associated MCU controls these tissue-specific uniporter phenotypes and downstream Ca2+ tuning of oxidative metabolism.
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•Abundance of MICU1 relative to MCU directly reflects their association•Proportion of MICU1-bound MCU is limited by tissue-specific MICU1 availability•MICU1:MCU ratio affects mitochondrial Ca2+ uptake in liver and muscle•Liver-like MICU1:MCU ratio in heart leads to contractile dysfunction
Paillard et al. report that the relative abundance of the pore-forming protein of the mitochondrial Ca2+ uniporter (MCU) and its Ca2+-sensing regulator (MICU1) define the proportion of MCU complexes with or without MICU1. This ratio is central to programming tissue-specific mitochondrial Ca2+ uptake phenotypes in the heart and liver.
Calcium is a universal signal in all eukaryotes, but the mechanism for encoding calcium signatures remains largely unknown. Calcium oscillations control pollen tube growth and fertilization in ...flowering plants, serving as a model for dissecting the molecular machines that mediate calcium fluctuations. We report that pollen-tube-specific cyclic nucleotide-gated channels (CNGC18, CNGC8, and CNGC7) together with calmodulin 2 (CaM2) constitute a molecular switch that either opens or closes the calcium channel depending on cellular calcium levels. Under low calcium, calcium-free calmodulin 2 (Apo-CaM2) interacts with CNGC18-CNGC8 complex, leading to activation of the influx channel and consequently increasing cytosolic calcium levels. Calcium-bound CaM2 dissociates from CNGC18/8 heterotetramer, closing the channel and initiating a downturn of cellular calcium levels. We further reconstituted the calcium oscillator in HEK293 cells, supporting the model that Ca2+-CaM-dependent regulation of CNGC channel activity provides an auto-regulatory feedback mechanism for calcium oscillations during pollen tube growth.
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•Plant CNGC subunit CNGC8 interacts with CNGC18, forming an inactive heterotetramer•Ca2+-free CaM2 interacts with CNGC18-CNGC8 heterotetramer, activating the channel•Calcium-loaded CaM2 dissociates from CNGC18-CNGC8, inactivating the channel•CNGC18/8-CaM2 interactions encode calcium oscillations in pollen tube growth
Calcium oscillations are universal signals for cell growth regulation in eukaryotes. Pan, Chai, Gao et al. identify and reconstitute a calcium oscillator from the Arabidopsis pollen tube. The oscillator consists of two CNGC subunits and a calmodulin that together drive oscillatory channel activity depending on the calcium levels in the cell.
Mitochondrial Ca2+ uptake tailors the strength of stimulation of plasma membrane phospholipase C–coupled receptors to that of cellular bioenergetics. However, how Ca2+ uptake by the mitochondrial ...Ca2+ uniporter (MCU) shapes receptor-evoked interorganellar Ca2+ signaling is unknown. Here, we used CRISPR/Cas9 gene knockout, subcellular Ca2+ imaging, and mathematical modeling to show that MCU is a universal regulator of intracellular Ca2+ signaling across mammalian cell types. MCU activity sustains cytosolic Ca2+ signaling by preventing Ca2+-dependent inactivation of store-operated Ca2+ release–activated Ca2+ channels and by inhibiting Ca2+ extrusion. Paradoxically, MCU knockout (MCU-KO) enhanced cytosolic Ca2+ responses to store depletion. Physiological agonist stimulation in MCU-KO cells led to enhanced frequency of cytosolic Ca2+ oscillations, endoplasmic reticulum Ca2+ refilling, nuclear translocation of nuclear factor for activated T cells transcription factors, and cell proliferation, without altering inositol-1,4,5-trisphosphate receptor activity. Our data show that MCU has dual counterbalancing functions at the cytosol–mitochondria interface, whereby the cell-specific MCU-dependent cytosolic Ca2+ clearance and buffering capacity of mitochondria reciprocally regulate interorganellar Ca2+ transfer and nuclear factor for activated T cells nuclear translocation during receptor-evoked signaling. These findings highlight the critical dual function of the MCU not only in the acute Ca2+ buffering by mitochondria but also in shaping endoplasmic reticulum and cytosolic Ca2+ signals that regulate cellular transcription and function.
The suprachiasmatic nucleus (SCN) of the hypothalamus orchestrates daily rhythms of physiology and behavior in mammals. Its circadian (∼24 hr) oscillations of gene expression and electrical activity ...are generated intrinsically and can persist indefinitely in temporal isolation. This robust and resilient timekeeping is generally regarded as a product of the intrinsic connectivity of its neurons. Here we show that neurons constitute only one “half” of the SCN clock, the one metabolically active during circadian daytime. In contrast, SCN astrocytes are active during circadian nighttime, when they suppress the activity of SCN neurons by regulating extracellular glutamate levels. This glutamatergic gliotransmission is sensed by neurons of the dorsal SCN via specific pre-synaptic NMDA receptor assemblies containing NR2C subunits. Remarkably, somatic genetic re-programming of intracellular clocks in SCN astrocytes was capable of remodeling circadian behavioral rhythms in adult mice. Thus, SCN circuit-level timekeeping arises from interdependent and mutually supportive astrocytic-neuronal signaling.
•SCN neurons are active during circadian day, but SCN astrocytes are active at night•Astrocytes direct circadian cycles of extracellular glutamate to inhibit SCN neurons•Astrocyte-derived inhibition is mediated by NMDAR2C complexes on dorsal SCN neurons•Genetic re-programming of the clock in SCN astrocytes reshapes circadian behavior
Neurons of the suprachiasmatic nucleus (SCN) are responsible for circadian pacemaking in mammals. Here, Brancaccio et al. demonstrate that SCN astrocytes also possess pacemaking properties and unravel how they set circadian tempo by reciprocal timing with their neuronal partners.
Living systems maintain or increase local order by working against the second law of thermodynamics. Thermodynamic consistency is restored as they consume free energy, thereby increasing the net ...entropy of their environment. Recently introduced estimators for the entropy production rate have provided major insights into the efficiency of important cellular processes. In experiments, however, many degrees of freedom typically remain hidden to the observer, and, in these cases, existing methods are not optimal. Here, by reformulating the problem within an optimization framework, we are able to infer improved bounds on the rate of entropy production from partial measurements of biological systems. Our approach yields provably optimal estimates given certain measurable transition statistics. In contrast to prevailing methods, the improved estimator reveals nonzero entropy production rates even when nonequilibrium processes appear time symmetric and therefore may pretend to obey detailed balance. We demonstrate the broad applicability of this framework by providing improved bounds on the energy consumption rates in a diverse range of biological systems including bacterial flagella motors, growing microtubules, and calcium oscillations within human embryonic kidney cells.
Background: Skeletal muscle cells continuously generate reactive oxygen species (ROS). Excessive ROS can affect lipids resulting in lipid peroxidation (LPO). Here we investigated the effects of ...myotube intracellular calcium-induced signaling eliciting contractions on the LPO induction and the impact of LPO-product 4-hydroxynonenal (4-HNE) on physiology/pathology of myotubes using C2C12 myoblasts.
Methods: C2C12 myoblasts were differentiated into myotubes, stimulated with caffeine and analyzed for the induction of LPO and formation of 4-HNE protein adducts. Further effects of 4-HNE on mitochondrial bioenergetics, NADH level, mitochondrial density and expression of mitochondrial metabolism genes were determined.
Results: Short and long-term caffeine stimulation of myotubes promoted superoxide production, LPO and formation of 4-HNE protein adducts. Furthermore, low 4-HNE concentrations had no effect on myotube viability and cellular redox homeostasis, while concentrations from 10 μM and above reduced myotube viability and significantly disrupted homeostasis. A time and dose-dependent 4-HNE effect on superoxide production and mitochondrial NADH-autofluorescence was observed. Finally, 4-HNE had strong impact on maximal respiration, spare respiratory capacity, ATP production, coupling efficiency of mitochondria and mitochondrial density.
Conclusion: Data presented in this work make evident for the first time that pathological 4-HNE levels elicit damaging effects on skeletal muscle cells while acute exposure to physiological 4-HNE induces transient adaptation.
General significance: This work suggests an important role of 4-HNE on the regulation of myotube's mitochondrial metabolism and cellular energy production. It further signifies the importance of skeletal muscle cells hormesis in response to acute stress in order to maintain essential biological functions.
•Caffeine induces does-dependent calcium oscillations and promotes superoxide production in myotubes.•Caffeine induces lipid peroxidation and 4-hydroxynonenal formation in myotubes.•The 4-hydroxynonenal regulates mitochondrial metabolism in skeletal muscle cells.•Acute exposure to physiological 4-HNE induces transient adaptation of skeletal muscle cells.
The accumulation of amyloid β peptide (Aβ) in the brain is hypothesized to be the major factor driving Alzheimer’s disease (AD) pathogenesis. Mounting evidence suggests that astrocytes are the ...primary target of Aβ neurotoxicity. Aβ is known to interfere with multiple calcium fluxes, thus disrupting the calcium homeostasis regulation of astrocytes, which are likely to produce calcium oscillations. Ca
2+
dyshomeostasis has been observed to precede the appearance of clinical symptoms of AD; however, it is experimentally very difficult to investigate the interactions of many mechanisms. Given that Ca
2+
disruption is ubiquitously involved in AD progression, it is likely that focusing on Ca
2+
dysregulation may serve as a potential therapeutic approach to preventing or treating AD, while current hypotheses concerning AD have so far failed to yield curable therapies. For this purpose, we derive and investigate a concise mathematical model for Aβ-mediated multi-pathway astrocytic intracellular Ca
2+
dynamics. This model accounts for how Aβ affects various fluxes contributions through voltage-gated calcium channels, Aβ-formed channels and ryanodine receptors. Bifurcation analysis of Aβ level, which reflected the corresponding progression of the disease, revealed that Aβ significantly induced the increasing Ca
2+
i
and frequency of calcium oscillations. The influence of inositol 1,4,5-trisphosphate production (IP
3
) is also investigated in the presence of Aβ as well as the impact of changes in resting membrane potential. In turn, the Ca
2+
flux can be considerably changed by exerting specific interventions, such as ion channel blockers or receptor antagonists. By doing so, a “combination therapy” targeting multiple pathways simultaneously has finally been demonstrated to be more effective. This study helps to better understand the effect of Aβ, and our findings provide new insight into the treatment of AD.
A simple network model consisting of a pyramidal neuron, an interneuron, and an astrocyte is constructed to simulate epileptiform discharges, focusing on the role of the interneuron in the ...pathological state. Simulation results show that with the change of the parameters related to abnormal glutamate degradation, the system can be transformed from bursting discharges or subthreshold oscillations to seizure-like discharges containing depolarization block. Meanwhile, the proposal of optogenetics has made it possible to target specific cells to modulate seizures, however, discoveries remain to be made regarding the specific effects limited by light mechanisms, stimulation patterns, and other factors. Hence, based on the constructed model, firstly, the experimental phenomenon that different types of light mechanisms are required to target the interneuron to control seizures under different situations is verified, and further, the effect of blue light targeting the astrocyte on seizure thresholds is revealed. The results demonstrate that the choice of stimulation frequency for seizure control varies in different situations, but the pulse width must be larger to be more conducive to control. In particular, the inhibitory photostimulation may change bursting discharges into spike discharges or subthreshold oscillations, in addition to eliminating the depolarization block part of the bursting discharges. Due to the slow-scale variation of calcium dynamics, stimulation with the same duty cycle does not have a consistent effect on thresholds for the appearance of epileptiform discharges. More importantly, by means of dynamical changes in the calcium signal near the bifurcation point from oscillation to resting, the effect of different stimulation patterns on the onset threshold can be explained. Our results may provide theoretical insight for the application of optogenetics in epileptic disorders caused by abnormal astrocyte function.
•Two possible epileptiform discharges due to astrocytic abnormality are simulated.•Epileptiform discharges can be removed as different lights act on the interneuron.•Same duty cycle blue light on the astrocyte may affect seizure thresholds diversely.•Effect of blue light on the astrocyte is explained by transition of calcium signals.
Astrocytes are important for information processing in the brain and they achieve this by fine‐tuning neuronal communication via continuous uptake and release of biochemical modulators of ...neurotransmission and synaptic plasticity. Often overlooked are their important functions in mechanosensation. Indeed, astrocytes can detect pathophysiological changes in the mechanical properties of injured, ageing, or degenerating brain tissue. We have recently shown that astrocytes surrounding mechanically‐stiff amyloid plaques upregulate the mechanosensitive ion channel, Piezo1. Moreover, ageing transgenic Alzheimer's rats harboring a chronic peripheral bacterial infection displayed enhanced Piezo1 expression in amyloid plaque‐reactive astrocytes of the hippocampus and cerebral cortex. Here, we have shown that the bacterial endotoxin, lipopolysaccharide (LPS), also upregulates Piezo1 in primary mouse cortical astrocyte cultures in vitro. Activation of Piezo1, via the small molecule agonist Yoda1, enhanced Ca2+ influx in both control and LPS‐stimulated astrocytes. Moreover, Yoda1 augmented intracellular Ca2+ oscillations but decreased subsequent Ca2+ influx in response to adenosine triphosphate (ATP) stimulation. Neither blocking nor activating Piezo1 affected cell viability. However, LPS‐stimulated astrocyte cultures exposed to the Piezo1 activator, Yoda1, migrated significantly slower than reactive astrocytes treated with the mechanosensitive channel‐blocking peptide, GsMTx4. Furthermore, our data show that activating Piezo1 channels inhibits the release of cytokines and chemokines, such as IL‐1β, TNFα, and fractalkine (CX3CL1), from LPS‐stimulated astrocyte cultures. Taken together, our results suggest that astrocytic Piezo1 upregulation may act to dampen neuroinflammation and could be a useful drug target for neuroinflammatory disorders of the brain.
Main Points
Activation of Piezo1 channels in LPS‐stimulated astrocytes induces calcium influx and augments intracellular calcium oscillations.
Piezo1 activation inhibits the release of pro‐inflammatory cytokines, IL‐1β and TNFα, from LPS‐stimulated astrocytes.
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