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Ascorbic acid depletion and scorbutic extracellular matrix in mice deficient in endoplasmic reticulum thiol oxidases

ABSTRACT: Endoplasmic reticulum (ER) thiol oxidases initiate a disulfide relay to oxidatively-fold secreted proteins. We found that combined loss-of-function mutations in genes encoding the ER thiol oxidases ERO1alpha, ERO1beta and PRDX4, compromised the extracellular matrix in mice and interfered with the intracellular maturation of procollagen. These severe abnormalities were associated with an unexpectedly-modest delay in disulfide bond formation in secreted proteins but a profound, five-fold lower procollagen 4 hydroxyproline content and enhanced cysteinyl sulfenic acid modification of ER proteins. Tissue ascorbic acid content was lower in mutant mice and ascorbic acid supplementation improved procollagen maturation and lowered sulfenic acid content, in vivo. In vitro, the presence of a sulfenic acid donor accelerated the oxidative inactivation of ascorbate by an H2O2 generating system. Compromised ER disulfide relay thus exposes protein thiols to competing oxidation to sulfenic acid, resulting in depletion of ascorbic acid, impaired procollagen proline 4-hydroxylation and a non-canonical form of scurvy. double and triple mutants and wild type

ORGANISM(S): Mus musculus

SUBMITTER: Brian Lam

PROVIDER: E-GEOD-39492 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Ascorbic acid depletion and scorbutic extracellular matrix in mice deficient in endoplasmic reticulum thiol oxidases

Project description:Endoplasmic reticulum (ER) thiol oxidases initiate a disulfide relay to oxidatively-fold secreted proteins. We found that combined loss-of-function mutations in genes encoding the ER thiol oxidases ERO1alpha, ERO1beta and PRDX4, compromised the extracellular matrix in mice and interfered with the intracellular maturation of procollagen. These severe abnormalities were associated with an unexpectedly-modest delay in disulfide bond formation in secreted proteins but a profound, five-fold lower procollagen 4 hydroxyproline content and enhanced cysteinyl sulfenic acid modification of ER proteins. Tissue ascorbic acid content was lower in mutant mice and ascorbic acid supplementation improved procollagen maturation and lowered sulfenic acid content, in vivo. In vitro, the presence of a sulfenic acid donor accelerated the oxidative inactivation of ascorbate by an H2O2 generating system. Compromised ER disulfide relay thus exposes protein thiols to competing oxidation to sulfenic acid, resulting in depletion of ascorbic acid, impaired procollagen proline 4-hydroxylation and a non-canonical form of scurvy.

2012-09-15 | GSE39492 | GEO

Mus musculus

Project description:Ascorbic acid depletion and scorbutic extracellular matrix in mice deficient in endoplasmic reticulum thiol oxidases

| PRJNA170982 | ENA

Expression data from control and Ascorbic Acid (AA) stimulated Mc-3T3-E1 osteoblasts

Project description:Despite advances in investigating functional aspects of osteoblast (OB) differentiation, especially studies on how bone proteins are deposited and mineralized, there has been little research on the intracellular trafficking of bone proteins during OB differentiation. Collagen synthesis and secretion is markedly upregulated upon Ascorbic Acid (AA) stimulation. Understanding the mechanism by which collagen is mobilized in specialized OB cells is important for both basic cell biology and diseases involving defects in bone secretion and deposition. RabGTPases are major regulators on protein trafficking throughout the cell. In this study, we identified the Rab GTPases that are upregulated during 5-day AA differentiation of OBs using microarray analysis, namely Rab1, Rab3d and Rab27b. We used microarrays to detail the global programme of gene expression underlying procollagen production and trafficking and identified up-regulated genes during this process. Control Mc3T3-E1 cells and 5 day Ascorbic Acid stimulated cells were processed for RNA extraction and hybridization on Affymetrix microarrays.

2012-05-01 | E-GEOD-37676 | biostudies-arrayexpress

Model of heart failure induced by mild thiol-blocking with cystamine

Project description:Depletion of cardiac ATP content is a characteristic feature of heart failure in patients and experimental animal models. To analyze the impact of insufficient ATP supply on heart function we inhibited cellular respiration by disulfide poisoning with the mild thiol-blocking agent, cystamine. We chose 4 month-old apolipoprotein E (apoE)-deficient mice, which are highly vulnerable to increased oxygen and ATP demands. After 4 weeks of cystamine treatment (300 mg/kg in drinking water), echocardiography and histology analyses demonstrated that apoE-deficient mice had developed heart failure with cardiac dilation. The microarray gene expression study of heart tissue from cystamine-treated apoE-deficient mice relative to untreated mice confirmed the development of heart failure showing up-regulation heart failure-specific genes by mild thiol-blocking with cystamine.

2010-12-03 | GSE25768 | GEO

Model of heart failure induced by mild thiol-blocking with cystamine

2010-12-03 | E-GEOD-25768 | biostudies-arrayexpress

Chondrodysplasia-inducing COL2A1 p.Gly1170Ser causes an ER storage defect without associated unfolded protein response in chondronoids

Project description:Collagenopathies are a group of clinically diverse disorders caused by defects in collagen folding and secretion. For example, mutations in the gene encoding collagen type-II (COL2A1), the primary collagen in cartilage, can lead to chondrodysplasias of various severities. One example is the Gly1170Ser substitution in procollagen-II, which causes precocious osteoarthritis and Legg-Calvé-Perthes disease. Here, we develop and characterize a novel induced pluripotent stem cell-based cartilage model of this disease, including both hetero- and homozygous genotypes. Biochemical characterization reveals that Gly1170Ser procollagen-II is notably slow to fold and secrete. Instead, procollagen-II accumulates intracellularly, consistent with an endoplasmic reticulum (ER) storage disorder. Intriguingly, though perhaps due to the pathologic substitution occurring within a triple-helical domain that lacks hydrophobic character, this intra-ER protein accumulation is not recognized by cellular stress responses, such as the unfolded protein response. Interactome studies showed that Gly1170Ser procollagen-II interacts to a greater extent with certain ER chaperones and modifying enzymes, consistent with its slow folding. These findings provide mechanistic elucidation into the etiology of this disease. Moreover, the expandable cartilage model developed here provides a valuable platform to rapidly screen and develop therapeutic strategies that can restore procollagen folding and secretion in this collagenopathy and others.

2024-07-02 | GSE244375 | GEO

Ascorbic acid reprogrames epigenome and epitranscriptome by recycling Fe (III) into Fe (II) in the catalytic cycle of dioxygenases (RNA-Seq)

Project description:Ascorbic acid has been reported to stimulate DNA iterative oxidase TET enzymes, Jumonji C-domain-containinghistone demethylase and potentially RNA m6A demethylase FTO and ALKBH5 as a cofactor. Although ascorbic acid has been widely investigated in reprogramming DNA and histone methylation status in vitro, in cell lines and mouse models, its specific role in the catalytic cycle of dioxygenases remains enigmatic. Here we systematically investigated the stimulation of ascorbic towards TET2, ALKBH3, histone demethylases and FTO. We find that ascorbic acid reprograms epitranscrip-tome by erasing the hypermethylated m6A sites. Biochemistry and Electron spin resonance (ESR) assays demonstrate that ascorbic acid enters the active pocket of dioxygenases, reduces Fe (III), either incorporated upon protein synthesis or generated upon rebounding the hydroxyl radical during oxidation, into Fe (II). Finally, we propose a new model for the catalytic cycle of dioxygenases by adding in the essential dynamic cofactor, ascorbic acid. Ascorbic acid refreshes and regenerates inactive dioxygenase through recycling Fe (III) into Fe (II) in a dynamic “hit-and-run” manner.

2024-02-01 | GSE242759 | GEO

Ascorbic acid reprogrames epigenome and epitranscriptome by recycling Fe (III) into Fe (II) in the catalytic cycle of dioxygenases (m6A)

2024-02-01 | GSE242758 | GEO

Ascorbic acid induces transient global active chromatin remodelling, providing clues for its anti-cancer effect

Project description:In addition to the role of antioxidant, ascorbic acid (reducing Vitamin C) is an important cofactor for Fe2+ and α-ketoglutarate (α-KG) dependent dioxygenases (Fe2+/α-KGDDs) that comprise many diverse enzymes, including collagen prolyl hydroxylases, jmjC (Jumonji C) domain containing histone demethylases, Ten-eleven translocation (TET) 5-methyl cytosine (5mC) dioxygenases, and N6-methyl adenosine (m6A) demethylase FTO and ALKBH5. Ascorbic acid was reported to induce global epigenetic reprogramming. Here we optimized the library construction flow chart of single-stranded DNA profiling method KAS-seq and utilized KAS-seq to profile transient chromatin states changes upon ascorbic acid treatment for 10 min. We identified several critical pathways affected by ascorbic acid treatment, providing some clues for explaining the reported positive impact of anti-cancer, anti-depression, and anti-obesity for taking ascorbic acid.

2023-07-25 | GSE237843 | GEO

Expression data from control and Ascorbic Acid (AA) stimulated Mc-3T3-E1 osteoblasts

2012-05-02 | GSE37676 | GEO