Crosstalk and barriers between the electron carriers of the endoplasmic reticulum.

SIGNIFICANCE The lumen of the endoplasmic reticulum (ER) constitutes a separate compartment with a special proteome and metabolome. The characteristic redox environment required for the optimal functioning of local pathways is defined by the redox couples of the main electron carriers. These molecules, glutathione, pyridine nucleotides, and ascorbic acid, are present within the ER, but their composition, concentration, and redox state are characteristically different from those observed in other subcellular compartments. Spatial and kinetic barriers contribute to the generation and maintenance of this special redox environment. RECENT ADVANCES The ER redox has usually been considered from the perspective of oxidative protein folding, one of the major functions of the ER. Thus, the lumen has been described as a relatively oxidizing subcellular compartment. CRITICAL ISSUES The ER redoxome has been scantily mapped. However, recent observations suggest that the redox systems in reduced and oxidized states are present simultaneously. The concerted actions of transmembrane uptake processes and local oxidoreductases as well as the absence of specific transport and enzyme activities maintain the oxidized state of the thiol-disulfide systems and the reduced state of the pyridine nucleotide redox systems. These states are prerequisites for the normal redox reactions localized in the ER. FUTURE DIRECTIONS An outline of the interactions between the major electron carriers of the ER will contribute to a better understanding of human diseases related to ER redox homeostasis

Transit of H2O2 across the endoplasmic reticulum membrane is not sluggish

Cellular metabolism provides various sources of hydrogen peroxide (H2O2) in different organelles and compartments. The suitability of H2O2 as an intracellular signaling molecule therefore also depends on its ability to pass cellular membranes. The propensity of the membranous boundary of the endoplasmic reticulum (ER) to let pass H2O2 has been discussed controversially. In this essay, we challenge the recent proposal that the ER membrane constitutes a simple barrier for H2O2 diffusion and support earlier data showing that (i) ample H2O2 permeability of the ER membrane is a prerequisite for signal transduction, (ii) aquaporin channels are crucially involved in the facilitation of H2O2 permeation, and (iii) a proper experimental framework not prone to artifacts is necessary to further unravel the role of H2O2 permeation in signal transduction and organelle biology. © 2016 Elsevier Inc

The glucose-6-phosphate transport is not mediated by a glucose-6-phosphate/phosphate exchange in liver microsomes

A phosphate-linked antiporter activity of the glucose-6-phosphate transporter (G6PT) has been recently described in liposomes including the reconstituded transporter protein. We directly investigated the mechanism of glucose-6-phosphate (G6P) transport in rat liver microsomal vesicles. Pre-loading with inorganic phosphate (Pi) did not stimulate G6P or Pi microsomal inward transport. Pi efflux from pre-loaded microsomes could not be enhanced by G6P or Pi addition. Rapid G6P or Pi influx was registered by light-scattering in microsomes not containing G6P or Pi. The G6PT inhibitor, S3483, blocked G6P transport irrespectively of experimental conditions. We conclude that hepatic G6PT functions as an uniporter

The glucose-6-phosphate transport is not mediated by a glucose-6-phosphate/phosphate exchange in liver microsomes

A phosphate-linked antiporter activity of the glucose-6-phosphate transporter (G6PT) has been recently described in liposomes including the reconstituded transporter protein. We directly investigated the mechanism of glucose-6-phosphate (G6P) transport in rat liver microsomal vesicles. Preloading with inorganic phosphate (Pi) did not stimulate G6P or Pi microsomal inward transport. Pi efflux from pre-loaded microsomes could not be enhanced by G6P or Pi addition. Rapid G6P or Pi influx was registered by light-scattering in microsomes not containing G6P or Pi. The G6PT inhibitor, S3483, blocked G6P transport irrespectively of experimental conditions. We conclude that hepatic G6PT functions as an uniporter

Arterial tortuosity syndrome : an ascorbate compartmentalization disorder?

Critical Issues: Although several hypotheses have been forwarded, the molecular mechanisms linking disrupted GLUT10 activity with arterial malformations are largely unknown. Recent Advances: The vascular and systemic manifestations and natural history of ATS patients have been largely delineated. GLUT10 was identified as an intracellular transporter of dehydroascorbic acid, which contributes to collagen and elastin cross-linking in the endoplasmic reticulum, redox homeostasis in the mitochondria, and global and gene-specific methylation/hydroxymethylation affecting epigenetic regulation in the nucleus. We revise here the current knowledge on ATS and the role of GLUT10 within the compartmentalization of ascorbate in physiological and diseased states. Future Directions: Centralization of clinical, treatment, and outcome data will enable better management for ATS patients. Establishment of representative animal disease models could facilitate the study of pathomechanisms underlying ATS. This might be relevant for other forms of vascular dysplasia, such as isolated aneurysm formation, hypertensive vasculopathy, and neovascularization. Antioxid. Redox Signal. 00, 000-000

Mevalonate Pathway-mediated ER Homeostasis Is Required for Haploid Stability in Human Somatic Cells

The somatic haploidy is unstable in diplontic animals, but cellular processes determining haploid stability remain elusive. Here, we found that inhibition of mevalonate pathway by pitavastatin, a widely used cholesterol-lowering drug, drastically destabilized the haploid state in HAP1 cells. Interestingly, cholesterol supplementation did not restore haploid stability in pitavastatin-treated cells, and cholesterol inhibitor U18666A did not phenocopy haploid destabilization. These results ruled out the involvement of cholesterol in haploid stability. Besides cholesterol perturbation, pitavastatin induced endoplasmic reticulum (ER) stress, the suppression of which by a chemical chaperon significantly restored haploid stability in pitavastatin-treated cells. Our data demonstrate the involvement of the mevalonate pathway in the stability of the haploid state in human somatic cells through managing ER stress, highlighting a novel link between ploidy and ER homeostatic control

Ero1α regulates Ca(2+) fluxes at the endoplasmic reticulum-mitochondria interface (MAM)

AIMS The endoplasmic reticulum (ER) is involved in many functions, including protein folding, redox homeostasis, and Ca(2+) storage and signaling. To perform these multiple tasks, the ER is composed of distinct, specialized subregions, amongst which mitochondrial-associated ER membranes (MAM) emerge as key signaling hubs. How these multiple functions are integrated with one another in living cells remains unclear. RESULTS Here we show that Ero1α, a key controller of oxidative folding and ER redox homeostasis, is enriched in MAM and regulates Ca(2+) fluxes. Downregulation of Ero1α by RNA interference inhibits mitochondrial Ca(2+) fluxes and modifies the activity of mitochondrial Ca(2+) uniporters. The overexpression of redox active Ero1α increases passive Ca(2+) efflux from the ER, lowering [Ca(2+)](ER) and mitochondrial Ca(2+) fluxes in response to IP3 agonists. INNOVATION The unexpected observation that Ca(2+) fluxes are affected by either increasing or decreasing the levels of Ero1α reveals a pivotal role for this oxidase in the early secretory compartment and implies a strict control of its amounts. CONCLUSIONS Taken together, our results indicate that the levels, subcellular localization, and activity of Ero1α coordinately regulate Ca(2+) and redox homeostasis and signaling in the early secretory compartment

Ero1α regulates Ca(2+) fluxes at the endoplasmic reticulum-mitochondria interface (MAM)

AIMS The endoplasmic reticulum (ER) is involved in many functions, including protein folding, redox homeostasis, and Ca(2+) storage and signaling. To perform these multiple tasks, the ER is composed of distinct, specialized subregions, amongst which mitochondrial-associated ER membranes (MAM) emerge as key signaling hubs. How these multiple functions are integrated with one another in living cells remains unclear. RESULTS Here we show that Ero1α, a key controller of oxidative folding and ER redox homeostasis, is enriched in MAM and regulates Ca(2+) fluxes. Downregulation of Ero1α by RNA interference inhibits mitochondrial Ca(2+) fluxes and modifies the activity of mitochondrial Ca(2+) uniporters. The overexpression of redox active Ero1α increases passive Ca(2+) efflux from the ER, lowering [Ca(2+)](ER) and mitochondrial Ca(2+) fluxes in response to IP3 agonists. INNOVATION The unexpected observation that Ca(2+) fluxes are affected by either increasing or decreasing the levels of Ero1α reveals a pivotal role for this oxidase in the early secretory compartment and implies a strict control of its amounts. CONCLUSIONS Taken together, our results indicate that the levels, subcellular localization, and activity of Ero1α coordinately regulate Ca(2+) and redox homeostasis and signaling in the early secretory compartment

Transit of H₂O₂ across the endoplasmic reticulum membrane is not sluggish

Abstract Cellular metabolism provides various sources of hydrogen peroxide (H₂O₂) in different organelles and compartments. The suitability of H₂O₂ as an intracellular signaling molecule therefore also depends on its ability to pass cellular membranes. The propensity of the membranous boundary of the endoplasmic reticulum (ER) to let pass H₂O₂ has been discussed controversially. In this essay, we challenge the recent proposal that the ER membrane constitutes a simple barrier for H₂O₂ diffusion and support earlier data showing that (i) ample H₂O₂ permeability of the ER membrane is a prerequisite for signal transduction, (ii) aquaporin channels are crucially involved in the facilitation of H₂O₂ permeation, and (iii) a proper experimental framework not prone to artifacts is necessary to further unravel the role of H₂O₂ permeation in signal transduction and organelle biology