Uniporter Research Articles

Role of mitochondrial calcium uniporter in mitophagy in SH-SY5Y cells subjected to oxygen-glucose deprivation and restoration

Objective To evaluate the role of mitochondrial calcium uniporter (MCU) in mitophagy in SH-SY5Y cells subjected to oxygen-glucose deprivation and restoration (OGD/R). Methods SH-SY5Y cells were cultured in vitro, seeded in 96-well plates at a density of 2×105 cells/ml, and randomly divided into 4 groups (n=6 each) using a random number table method: control group (group C), group OGD/R, OGD/R plus MCU inhibitor group (group OGD/R + Ru360) and MCU inhibitor group (group Ru360). Cells were cultured in normal culture medium in group C. Cells were subjected to O2-glucose deprivation for 6 h followed by restoration of O2-glucose supply for 24 h in group OGD/R.In group OGD/R+ Ru360, Ru360 at a final concentration of 10 μmol/L was added at 30 min before O2-glucose deprivation, and the other treatments were similar to those previously described in group OGD/R.Ru360 was added at a final concentration of 10 μmol/L, and 30 min later cells were cultured under normoxic conditions in group Ru360.At 24 h of restoration of O2-glucose supply, cell counting kit-8 assay was used to detect the cell survival rate, JC-1 assay was used to detect mitochondrial membrane potential (MMP), the ultrastructure of cells was observed with a transmission electron microscope, and the expression of p62, Tom20 and Beclin-1 was detected by Western blot. Results Compared with group C, no significant change was found in each parameter in group Ru360 (P>0.05), the cell survival rate and MMP were significantly decreased, the expression of Tom20 and p62 was down-regulated, Beclin-1 expression was up-regulated (P<0.01), the mitochondria swelled, and mitochondrial autophagosomes were increased in group OGD/R.Compared with group OGD/R, the cell survival rate and MMP were significantly increased, the expression of Tom20 and p62 was up-regulated, Beclin-1 expression was down-regulated (P<0.01), the mitochondrial morphology kept well, and mitochondrial autophagosomes were decreased. Conclusion MCU is involved in the process of mitophagy in SH-SY5Y cells subjected to OGD/R. Key words: Mitochondria; Ion transport; Cell hypoxia; Autophagy

A channel for calcium

Structural Biology Maintaining the correct balance of calcium concentrations between the cytosol and the mitochondria is essential for cellular physiology. A calcium-selective channel called the mitochondrial calcium uniporter (MCU) mediates calcium entry into mitochondria. Yoo et al. report the high-resolution structure of MCU from Neurospora crassa. The channel is formed by four MCU protomers with differing symmetry between the soluble and membrane domains. The structure, together with mutagenesis, suggests that two acidic rings inside the channel provide the selectivity for calcium. Science , this issue p. [506][1] [1]: /lookup/doi/10.1126/science.aar4056

Structural Events in a Bacterial Uniporter Leading to Translocation of Glucose Inside the Cytosol

Structural Events in a Bacterial Uniporter Leading to Translocation of Glucose Inside the Cytosol

Role of mitochondrial calcium uniporter in lung ischemia-reperfusion injury in rats

Objective To evaluate the role of mitochondrial calcium uniporter in lung ischemia-reperfusion(I/R)injury in rats. Methods Thirty healthy male Wistar rats, weighing 250-300 g, were randomly divided into 3 groups(n=10 each)using a random number table: sham operation group(group S), group I/R and ruthenium red(mitochondrial calcium uniporter inhibitor)group(group RR). Lung I/R was produced by clamping the left pulmonary hilum for 45 min followed by 2 h reperfusion in I/R and RR groups.In addition, ruthenium red 2.5 mg/kg was injected via the tail vein at 1 h before occlusion of the left pulmonary hilum in RR group.Arterial blood samples were obtained at 2 h of reperfusion for blood gas analysis, and oxygenation index(PaO2/FiO2)was calculated.The rats were sacrificed, and then the lungs were removed for determination of wet/dry lung weight ratio(W/D ratio). Results Compared with group S, oxygenation index was significantly decreased, and W/D ratio was increased in I/R and RR groups.Oxygenation index was significantly higher, and W/D ratio was lower in group RR than in group I/R. Conclusion The mechanism underlying lung I/R injury is related to the opening of mitochondrial calcium uniporter in rats. Key words: Mitochondrial membrane transport proteins; Reperfusion injury; Lung

The uniporter: From newly identified parts to function

The uniporter: From newly identified parts to function

The calcium uniporter regulates the permeability transition pore in isolated cortical mitochondria.

To investigate the influence of the mitochondrial calcium uniporter on the mitochondrial permeability transition pore, the present study observed mitochondrial morphology in cortical neurons isolated from adult rats using transmission electron microscopy, and confirmed the morphology and activity of isolated mitochondria by detecting succinic dehydrogenase and monoamine oxidase, two mitochondrial enzymes. Isolated mitochondria were treated with either ruthenium red, an inhibitor of the uniporter, spermine, an activator of the uniporter, or in combination with cyclosporin A, an inhibitor of the mitochondrial permeability transition pore. Results showed that ruthenium red inhibited CaCl2-induced mitochondrial permeability transition pore opening, spermine enhanced opening, and cyclosporin A attenuated the effects of spermine. Results demonstrated that the mitochondrial calcium uniporter plays a role in regulating the mitochondrial permeability transition pore in mitochondria isolated from the rat brain cortex.

Diazoxide preconditioning protects cerebrum against ischemia/reperfusion injury via mitochondrial calcium uniporter

Objective To investigate whether diazoxide preconditioning protects cerebrum against i(s)chemia reperfusion injury via mitochondrial calcium uniporter(MCU)or not.Methods The ischemia reperfusion model was performed by thread occlusion of middle cerebral artery for 2 h,and then 24 h reperfusion.Fifty-two rats were randomly divided into four groups:group A (sham-operated group),group B(ischemia/reperfusion group),group C(diazoxide group),group D(diazozide-spermine group).After 24 h reperfusion,neuropsychological score,the mitochondrial calcium content([Ca2+]m),,the activity of superoxide dismutase (SOD)and glutathione peroxidase(GSH-Px),the content of malondialdehyde(MDA)and nitric oxide(NO)in ischemic hemisphere were observed in above groups respectively.Results Compared with group B,diazoxide preconditioning in group C improved neuropsychological score(13.1 ±0.8,P＜0.01),improved the activity of SOD(143 ±-18)U/mgprot and GSH-Px(84 ±8)U/mgprot(P＜0.05),and decreased the content of MDA(0.799 ±0.092)nmol/mgprot and NO(0.216±0.138)nmol/mgprot(P＜0.05)and[Ca2+]m (293±7)nmol/mgprot(P＜0.01).Spermine in group D attenuated the beneficial effects of diazoxide preconditioning in group C.Compared with group C,spermine attenuated neuropsychological score(8.0±0.7,P＜0.01)and the activity of SOD(128±7)U/mgprot and GSH-Px(73 ±6)U/mgprot(P＜0.05),and improved[Ca2+]m(322±10)nmol/mgprot(P＜0.01)and the content of MDA (0.955±0.141)nmol/mgprot and NO(0.575±0.292)nmol/mgprot(P＜0.05).Conclusions The effects of diazoxide preconditioning against cerebral ischemia reperfusion injury by attenuating[Ca2+]m and peroxidation injury in rats may involve in the suppression of MCU during reperfusion. Key words: Diazoxide; Spermine; Mitochondria; Brain ischemia; Reperfusion

Altered regulation of cardiac contraction and relaxation by Ca2+ in heart failure

Myocardial contractile dysfunction in congestive heart failure is characterized by a decrease in force developed and retardation of relaxation. These alterations are mainly due to those in intracellular Ca(2 +) transients (CaT) . CaT are regulated by a number of functional proteins, including sarcolemmal L-type Ca(2 +) channels, Na(+)/Ca(2 +) exchanger and Ca(2 +) ATPase, sarcoplasmic reticulum Ca(2 +) ATPase (SERCA 2 ) , phospholamban and ryanodine receptors, and mitochondrial Ca(2 +) uniporter. Changes in expression and function of these regulatory proteins that occur in the course of increasing severity of heart failure are responsible for the characteristic changes in force development and relaxation observed under pathophysiological conditions in congestive heart failure.

Is the mitochondrial Ca 2+ uniporter a voltage-modulated transport pathway?

The influence of membrane potential (Δψ) on Ca 2+ transport through the Ca 2+ uniporter (UP) was investigated in fura-2-loaded rat heart mitochondria at physiologically relevant-submicromolar-external [Ca 2+]. In the absence of Δψ the UP could not mediate Ca 2+ uptake even when an 8-fold external (∼500 nM) to internal (∼60 nM) [Ca 2+] gradient was present and charge compensation was provided by acetate and the protonophore, CCCP. A small (∼−120 mV) and transient Δψ (generated by valinomycin) resulted in a rise in matrix [Ca 2+] only when external [Ca 2+] exceeded 150 nM. At physiologically high (∼−180 mV) and stable Δψ this threshold value for Ca 2+ uptake dropped to 15 nM. The results indicate that (1) at physiological [Ca 2+] m Δψ in addition to being a component of Δμ Ca 2+ seems to be necessary for providing a transport-competent conformation for the UP; and (2) below a threshold [Ca 2+] m the UP cannot operate even in the presence of a high electric driving force.