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The online version of this article (doi:10.1186/s12915-017-0445-8) contains supplementary material, which is available to authorized users.
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Affiliation(s)
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1
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Zannini F, Herrmann JM, Couturier J, Rouhier N. Oxidation of Arabidopsis thaliana COX19 Using the Combined Action of ERV1 and Glutathione. Antioxidants (Basel) 2023; 12:1949. [PMID: 38001802 PMCID: PMC10669224 DOI: 10.3390/antiox12111949] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/06/2023] [Revised: 10/26/2023] [Accepted: 10/30/2023] [Indexed: 11/26/2023] Open
Abstract
Protein import and oxidative folding within the intermembrane space (IMS) of mitochondria relies on the MIA40-ERV1 couple. The MIA40 oxidoreductase usually performs substrate recognition and oxidation and is then regenerated by the FAD-dependent oxidase ERV1. In most eukaryotes, both proteins are essential; however, MIA40 is dispensable in Arabidopsis thaliana. Previous complementation experiments have studied yeast mia40 mutants expressing a redox inactive, but import-competent versions of yeast Mia40 using A. thaliana ERV1 (AtERV1) suggest that AtERV1 catalyzes the oxidation of MIA40 substrates. We assessed the ability of both yeast and Arabidopsis MIA40 and ERV1 recombinant proteins to oxidize the apo-cytochrome reductase CCMH and the cytochrome c oxidase assembly protein COX19, a typical MIA40 substrate, in the presence or absence of glutathione, using in vitro cysteine alkylation and cytochrome c reduction assays. The presence of glutathione used at a physiological concentration and redox potential was sufficient to support the oxidation of COX19 by AtERV1, providing a likely explanation for why MIA40 is not essential for the import and oxidative folding of IMS-located proteins in Arabidopsis. The results point to fundamental biochemical differences between Arabidopsis and yeast ERV1 in catalyzing protein oxidation.
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Affiliation(s)
- Flavien Zannini
- Université de Lorraine, INRAE, IAM, F-54000 Nancy, France; (F.Z.); (J.C.)
| | - Johannes M. Herrmann
- Cell Biology, University of Kaiserslautern, RPTU, 67663 Kaiserslautern, Germany;
| | - Jérémy Couturier
- Université de Lorraine, INRAE, IAM, F-54000 Nancy, France; (F.Z.); (J.C.)
| | - Nicolas Rouhier
- Université de Lorraine, INRAE, IAM, F-54000 Nancy, France; (F.Z.); (J.C.)
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2
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Verma AK, Sharma A, Subramaniyam N, Gandhi CR. Augmenter of liver regeneration: Mitochondrial function and steatohepatitis. J Hepatol 2022; 77:1410-1421. [PMID: 35777586 DOI: 10.1016/j.jhep.2022.06.019] [Citation(s) in RCA: 16] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/04/2022] [Revised: 05/24/2022] [Accepted: 06/09/2022] [Indexed: 12/15/2022]
Abstract
Augmenter of liver regeneration (ALR), a ubiquitous fundamental life protein, is expressed more abundantly in the liver than other organs. Expression of ALR is highest in hepatocytes, which also constitutively secrete it. ALR gene transcription is regulated by NRF2, FOXA2, SP1, HNF4α, EGR-1 and AP1/AP4. ALR's FAD-linked sulfhydryl oxidase activity is essential for protein folding in the mitochondrial intermembrane space. ALR's functions also include cytochrome c reductase and protein Fe/S maturation activities. ALR depletion from hepatocytes leads to increased oxidative stress, impaired ATP synthesis and apoptosis/necrosis. Loss of ALR's functions due to homozygous mutation causes severe mitochondrial defects and congenital progressive multiorgan failure, suggesting that individuals with one functional ALR allele might be susceptible to disorders involving compromised mitochondrial function. Genetic ablation of ALR from hepatocytes induces structural and functional mitochondrial abnormalities, dysregulation of lipid homeostasis and development of steatohepatitis. High-fat diet-fed ALR-deficient mice develop non-alcoholic steatohepatitis (NASH) and fibrosis, while hepatic and serum levels of ALR are lower than normal in human NASH and NASH-cirrhosis. Thus, ALR deficiency may be a critical predisposing factor in the pathogenesis and progression of NASH.
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Affiliation(s)
- Alok Kumar Verma
- Division of Gastroenterology, Hepatology and Nutrition, Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA; Cincinnati VA Medical Center, Cincinnati, Ohio, USA
| | - Akanksha Sharma
- Division of Gastroenterology, Hepatology and Nutrition, Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA; Cincinnati VA Medical Center, Cincinnati, Ohio, USA
| | - Nithyananthan Subramaniyam
- Division of Gastroenterology, Hepatology and Nutrition, Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA
| | - Chandrashekhar R Gandhi
- Division of Gastroenterology, Hepatology and Nutrition, Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA; Cincinnati VA Medical Center, Cincinnati, Ohio, USA; Department of Surgery, University of Cincinnati, Cincinnati, Ohio, USA.
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3
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Backes S, Garg SG, Becker L, Peleh V, Glockshuber R, Gould SB, Herrmann JM. Development of the Mitochondrial Intermembrane Space Disulfide Relay Represents a Critical Step in Eukaryotic Evolution. Mol Biol Evol 2019; 36:742-756. [PMID: 30668797 DOI: 10.1093/molbev/msz011] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/26/2023] Open
Abstract
The mitochondrial intermembrane space evolved from the bacterial periplasm. Presumably as a consequence of their common origin, most proteins of these compartments are stabilized by structural disulfide bonds. The molecular machineries that mediate oxidative protein folding in bacteria and mitochondria, however, appear to share no common ancestry. Here we tested whether the enzymes Erv1 and Mia40 of the yeast mitochondrial disulfide relay could be functionally replaced by corresponding components of other compartments. We found that the sulfhydryl oxidase Erv1 could be replaced by the Ero1 oxidase or the protein disulfide isomerase from the endoplasmic reticulum, however at the cost of respiration deficiency. In contrast to Erv1, the mitochondrial oxidoreductase Mia40 proved to be indispensable and could not be replaced by thioredoxin-like enzymes, including the cytoplasmic reductase thioredoxin, the periplasmic dithiol oxidase DsbA, and Pdi1. From our studies we conclude that the profound inertness against glutathione, its slow oxidation kinetics and its high affinity to substrates renders Mia40 a unique and essential component of mitochondrial biogenesis. Evidently, the development of a specific mitochondrial disulfide relay system represented a crucial step in the evolution of the eukaryotic cell.
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Affiliation(s)
- Sandra Backes
- Cell Biology, University of Kaiserslautern, Kaiserslautern, Germany
| | - Sriram G Garg
- Molecular Evolution, Heinrich-Heine-University of Dusseldorf, Dusseldorf, Germany
| | - Laura Becker
- Cell Biology, University of Kaiserslautern, Kaiserslautern, Germany
| | - Valentina Peleh
- Cell Biology, University of Kaiserslautern, Kaiserslautern, Germany
| | - Rudi Glockshuber
- Molecular Biology and Biophysics, ETH Zürich, Zürich, Switzerland
| | - Sven B Gould
- Molecular Evolution, Heinrich-Heine-University of Dusseldorf, Dusseldorf, Germany
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4
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Erdogan AJ, Ali M, Habich M, Salscheider SL, Schu L, Petrungaro C, Thomas LW, Ashcroft M, Leichert LI, Roma LP, Riemer J. The mitochondrial oxidoreductase CHCHD4 is present in a semi-oxidized state in vivo. Redox Biol 2018; 17:200-206. [PMID: 29704824 PMCID: PMC6007816 DOI: 10.1016/j.redox.2018.03.014] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2018] [Accepted: 03/22/2018] [Indexed: 11/30/2022] Open
Abstract
Disulfide formation in the mitochondrial intermembrane space is an essential process catalyzed by a disulfide relay machinery. In mammalian cells, the key enzyme in this machinery is the oxidoreductase CHCHD4/Mia40. Here, we determined the in vivo CHCHD4 redox state, which is the major determinant of its cellular activity. We found that under basal conditions, endogenous CHCHD4 redox state in cultured cells and mouse tissues was predominantly oxidized, however, degrees of oxidation in different tissues varied from 70% to 90% oxidized. To test whether differences in the ratio between CHCHD4 and ALR might explain tissue-specific differences in the CHCHD4 redox state, we determined the molar ratio of both proteins in different mouse tissues. Surprisingly, ALR is superstoichiometric over CHCHD4 in most tissues. However, the levels of CHCHD4 and the ratio of ALR over CHCHD4 appear to correlate only weakly with the redox state, and although ALR is present in superstoichiometric amounts, it does not lead to fully oxidized CHCHD4.
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Affiliation(s)
- Alican J Erdogan
- Department of Chemistry, Institute of Biochemistry, University of Cologne, Zuelpicher Str. 47a, 50674 Cologne, Germany
| | - Muna Ali
- Department of Chemistry, Institute of Biochemistry, University of Cologne, Zuelpicher Str. 47a, 50674 Cologne, Germany; Department of Biology, Cellular Biochemistry, University of Kaiserslautern, Erwin-Schroedinger-Str. 13, 67663 Kaiserslautern, Germany
| | - Markus Habich
- Department of Chemistry, Institute of Biochemistry, University of Cologne, Zuelpicher Str. 47a, 50674 Cologne, Germany
| | - Silja L Salscheider
- Department of Chemistry, Institute of Biochemistry, University of Cologne, Zuelpicher Str. 47a, 50674 Cologne, Germany
| | - Laura Schu
- Department of Biology, Cellular Biochemistry, University of Kaiserslautern, Erwin-Schroedinger-Str. 13, 67663 Kaiserslautern, Germany
| | - Carmelina Petrungaro
- Department of Chemistry, Institute of Biochemistry, University of Cologne, Zuelpicher Str. 47a, 50674 Cologne, Germany
| | - Luke W Thomas
- Department of Medicine, University of Cambridge, Cambridge Biomedical Campus, Cambridge, UK
| | - Margaret Ashcroft
- Department of Medicine, University of Cambridge, Cambridge Biomedical Campus, Cambridge, UK
| | - Lars I Leichert
- Institute for Biochemistry and Pathobiochemistry - Microbial Biochemistry, Ruhr-Universität Bochum, 44797 Bochum, Germany
| | - Leticia Prates Roma
- Biophysics Department, Center for Integrative Physiology and Molecular Medicine, Saarland University, 66421 Homburg, Saar, Germany
| | - Jan Riemer
- Department of Chemistry, Institute of Biochemistry, University of Cologne, Zuelpicher Str. 47a, 50674 Cologne, Germany.
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5
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Gupta P, Venugopal SK. Augmenter of liver regeneration: A key protein in liver regeneration and pathophysiology. Hepatol Res 2018; 48:587-596. [PMID: 29633440 DOI: 10.1111/hepr.13077] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/15/2018] [Revised: 03/10/2018] [Accepted: 03/29/2018] [Indexed: 12/22/2022]
Abstract
Liver is constantly exposed to pathogens, viruses, chemicals, and toxins, and several of them cause injury, leading to the loss of liver mass and sometimes resulting in cirrhosis and cancer. Under physiological conditions, liver can regenerate if the loss of cells is less than the proliferation of hepatocytes. If the loss is more than the proliferation, the radical treatment available is liver transplantation. Due to this reason, the search for an alternative therapeutic agent has been the focus of liver research. Liver regeneration is regulated by several growth factors; one of the key factors is augmenter of liver regeneration (ALR). Involvement of ALR has been reported in crucial processes such as oxidative phosphorylation, maintenance of mitochondria and mitochondrial biogenesis, and regulation of autophagy and cell proliferation. Augmenter of liver regeneration has been observed to be involved in liver regeneration by not only overcoming cell cycle inhibition but by maintaining the stem cell pool as well. These observations have created curiosity regarding the possible role of ALR in maintenance of liver health. Thus, this review brings a concise presentation of the work done in areas exploring the role of ALR in normal liver physiology and in liver health maintenance by fighting liver diseases, such as liver failure, non-alcoholic fatty liver disease/non-alcoholic steatohepatitis, viral infections, cirrhosis, and hepatocellular carcinoma.
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Affiliation(s)
- Parul Gupta
- Faculty of Life Sciences and Biotechnology, South Asian University, New Delhi, India
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6
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Zhang C, An W. Progress in research of augmenter of liver regeneration. Shijie Huaren Xiaohua Zazhi 2017; 25:3171-3179. [DOI: 10.11569/wcjd.v25.i36.3171] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Augmenter of liver regeneration (ALR), also known as hepatic stimulatory substance or hepatopoietin, is expressed ubiquitously in all organs, and exclusively in hepatocytes in the liver. Over the past decade, research indicates that ALR is able to promote growth of hepatocytes in the regenerating or injured liver, and plays an important role in hepatocyte transplantation, the pathogenesis of fulminant hepatitis, liver regeneration, and the development of hepatocellular carcinoma.
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Affiliation(s)
- Chao Zhang
- Department of Cell Biology, Basic Medical College, Capital Medical University, Beijing 100191, China
| | - Wei An
- Department of Cell Biology, Basic Medical College, Capital Medical University, Beijing 100191, China
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7
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Peleh V, Zannini F, Backes S, Rouhier N, Herrmann JM. Erv1 of Arabidopsis thaliana can directly oxidize mitochondrial intermembrane space proteins in the absence of redox-active Mia40. BMC Biol 2017; 15:106. [PMID: 29117860 PMCID: PMC5679390 DOI: 10.1186/s12915-017-0445-8] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2017] [Accepted: 10/19/2017] [Indexed: 11/20/2022] Open
Abstract
Background Many proteins of the mitochondrial intermembrane space (IMS) contain structural disulfide bonds formed by the mitochondrial disulfide relay. In fungi and animals, the sulfhydryl oxidase Erv1 ‘generates’ disulfide bonds that are passed on to the oxidoreductase Mia40, which oxidizes substrate proteins. A different structural organization of plant Erv1 proteins compared to that of animal and fungal orthologs was proposed to explain its inability to complement the corresponding yeast mutant. Results Herein, we have revisited the biochemical and functional properties of Arabidopsis thaliana Erv1 by both in vitro reconstituted activity assays and complementation of erv1 and mia40 yeast mutants. These mutants were viable, however, they showed severe defects in the biogenesis of IMS proteins. The plant Erv1 was unable to oxidize yeast Mia40 and rather even blocked its activity. Nevertheless, it was able to mediate the import and folding of mitochondrial proteins. Conclusions We observed that plant Erv1, unlike its homologs in fungi and animals, can promote protein import and oxidative protein folding in the IMS independently of the oxidoreductase Mia40. In accordance to the absence of Mia40 in many protists, our study suggests that the mitochondrial disulfide relay evolved in a stepwise reaction from an Erv1-only system to which Mia40 was added in order to improve substrate specificity.
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The mitochondrial disulfide relay evolved in a step-wise manner from an Erv1-only system.
- Valentina Peleh
- Cell Biology, University of Kaiserslautern, Erwin-Schrödinger-Strasse 13, 67663, Kaiserslautern, Germany
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- Flavien Zannini
- Unité Mixte de Recherches 1136 Interactions Arbres-Microorganismes, Université de Lorraine/INRA, Faculté des sciences et technologies, 54500 Vandoeuvre-lès-Nancy, Nancy, France
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- Sandra Backes
- Cell Biology, University of Kaiserslautern, Erwin-Schrödinger-Strasse 13, 67663, Kaiserslautern, Germany
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- Nicolas Rouhier
- Unité Mixte de Recherches 1136 Interactions Arbres-Microorganismes, Université de Lorraine/INRA, Faculté des sciences et technologies, 54500 Vandoeuvre-lès-Nancy, Nancy, France.
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- Johannes M Herrmann
- Cell Biology, University of Kaiserslautern, Erwin-Schrödinger-Strasse 13, 67663, Kaiserslautern, Germany.
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Abstract
Cysteine thiols are among the most reactive functional groups in proteins, and their pairing in disulfide linkages is a common post-translational modification in proteins entering the secretory pathway. This modest amino acid alteration, the mere removal of a pair of hydrogen atoms from juxtaposed cysteine residues, contrasts with the substantial changes that characterize most other post-translational reactions. However, the wide variety of proteins that contain disulfides, the profound impact of cross-linking on the behavior of the protein polymer, the numerous and diverse players in intracellular pathways for disulfide formation, and the distinct biological settings in which disulfide bond formation can take place belie the simplicity of the process. Here we lay the groundwork for appreciating the mechanisms and consequences of disulfide bond formation in vivo by reviewing chemical principles underlying cysteine pairing and oxidation. We then show how enzymes tune redox-active cofactors and recruit oxidants to improve the specificity and efficiency of disulfide formation. Finally, we discuss disulfide bond formation in a cellular context and identify important principles that contribute to productive thiol oxidation in complex, crowded, dynamic environments.
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Affiliation(s)
- Deborah Fass
- Department of Structural Biology, Weizmann Institute of Science , Rehovot 7610001, Israel
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- Colin Thorpe
- Department of Chemistry and Biochemistry, University of Delaware , Newark, Delaware 19716, United States
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Thangamani S, Maland M, Mohammad H, Pascuzzi PE, Avramova L, Koehler CM, Hazbun TR, Seleem MN. Repurposing Approach Identifies Auranofin with Broad Spectrum Antifungal Activity That Targets Mia40-Erv1 Pathway.
Front Cell Infect Microbiol 2017;
7:4. [PMID:
28149831 PMCID:
PMC5241286 DOI:
10.3389/fcimb.2017.00004]
[Citation(s) in RCA: 67] [Impact Index Per Article: 8.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/04/2016] [Accepted: 01/03/2017] [Indexed: 12/24/2022] Open
Abstract
Current antifungal therapies have limited effectiveness in treating invasive fungal infections. Furthermore, the development of new antifungal is currently unable to keep pace with the urgent demand for safe and effective new drugs. Auranofin, an FDA-approved drug for the treatment of rheumatoid arthritis, inhibits growth of a diverse array of clinical isolates of fungi and represents a new antifungal agent with a previously unexploited mechanism of action. In addition to auranofin's potent antifungal activity against planktonic fungi, this drug significantly reduces the metabolic activity of Candida cells encased in a biofilm. Unbiased chemogenomic profiling, using heterozygous S. cerevisiae deletion strains, combined with growth assays revealed three probable targets for auranofin's antifungal activity—mia40, acn9, and coa4. Mia40 is of particular interest given its essential role in oxidation of cysteine rich proteins imported into the mitochondria. Biochemical analysis confirmed auranofin targets the Mia40-Erv1 pathway as the drug inhibited Mia40 from interacting with its substrate, Cmc1, in a dose-dependent manner similar to the control, MB-7. Furthermore, yeast mitochondria overexpressing Erv1 were shown to exhibit resistance to auranofin as an increase in Cmc1 import was observed compared to wild-type yeast. Further in vivo antifungal activity of auranofin was examined in a Caenorhabditis elegans animal model of Cryptococcus neoformans infection. Auranofin significantly reduced the fungal load in infected C. elegans. Collectively, the present study provides valuable evidence that auranofin has significant promise to be repurposed as a novel antifungal agent and may offer a safe, effective, and quick supplement to current approaches for treating fungal infections.
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Affiliation(s)
- Shankar Thangamani
- Department of Comparative Pathobiology, College of Veterinary Medicine, Purdue University West Lafayette, IN, USA
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- Matthew Maland
- Department of Chemistry and Biochemistry and the Molecular Biology Institute, University of California, Los Angeles Los Angeles, CA, USA
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- Haroon Mohammad
- Department of Comparative Pathobiology, College of Veterinary Medicine, Purdue University West Lafayette, IN, USA
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- Pete E Pascuzzi
- Purdue University Libraries, Purdue UniversityWest Lafayette, IN, USA; Department of Biochemistry, Purdue UniversityWest Lafayette, IN, USA
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- Larisa Avramova
- Bindley Bioscience Center, Purdue University West Lafayette, IN, USA
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- Carla M Koehler
- Department of Chemistry and Biochemistry and the Molecular Biology Institute, University of California, Los Angeles Los Angeles, CA, USA
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- Tony R Hazbun
- Bindley Bioscience Center, Purdue UniversityWest Lafayette, IN, USA; Department of Medicinal Chemistry and Molecular Pharmacology, College of Pharmacy, Purdue UniversityWest Lafayette, IN, USA
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- Mohamed N Seleem
- Department of Comparative Pathobiology, College of Veterinary Medicine, Purdue UniversityWest Lafayette, IN, USA; Purdue Institute for Inflammation, Immunology, and Infectious DiseasesWest Lafayette, IN, USA
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Balogh T, Lőrincz T, Stiller I, Mandl J, Bánhegyi G, Szarka A. The Level of ALR is Regulated by the Quantity of Mitochondrial DNA.
Pathol Oncol Res 2015;
22:431-7. [PMID:
26584568 DOI:
10.1007/s12253-015-0020-y]
[Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/03/2015] [Accepted: 11/16/2015] [Indexed: 01/08/2023]
Abstract
Augmenter of liver regeneration (ALR) contributes to mitochondrial biogenesis, maintenance and to the physiological operation of mitochondria. The depletion of ALR has been widely studied and had serious consequences on the mitochondrial functions. However the inverse direction, the effect of the depletion of mitochondrial electron transfer chain and mtDNA on ALR expression has not been investigated yet. Thus mtDNA depleted, ρ(0) cell line was prepared to investigate the role of mitochondrial electron transfer chain and mtDNA on ALR expression. The depletion of mtDNA has not caused any difference at mRNA level, but at protein level the expression of ALR has been markedly increased. The regulatory role of ATP and ROS levels could be ruled out because the treatment of the parental cell line with different respiratory inhibitors and uncoupling agent could not provoke any changes in the protein level of ALR. The effect of mtDNA depletion on the protein level of ALR has been proved not to be liver specific, since the phenomenon could be observed in the case of two other, non-hepatic cell lines. It seems the level of mtDNA and/or its products may have regulatory role on the protein level of ALR. The up-regulation of ALR can be a part of the adaptive response in ρ(0) cells that preserves the structural integrity and the transmembrane potential despite the absence of protein components encoded by the mtDNA.
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Affiliation(s)
- Tibor Balogh
- Department of Applied Biotechnology and Food Science, Laboratory of Biochemistry and Molecular Biology, Budapest University of Technology and Economics, Szent Gellért tér 4, Budapest, 1111, Hungary
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- Tamás Lőrincz
- Department of Applied Biotechnology and Food Science, Laboratory of Biochemistry and Molecular Biology, Budapest University of Technology and Economics, Szent Gellért tér 4, Budapest, 1111, Hungary
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- Ibolya Stiller
- Pathobiochemistry Research Group of Hungarian Academy of Sciences, Semmelweis University, P.O. Box 260, Budapest, 1444, Hungary
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- József Mandl
- Department of Medical Chemistry, Molecular Biology and Pathobiochemistry Pathobiochemistry, Semmelweis University, POB 260, Budapest, 1444, Hungary
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- Gábor Bánhegyi
- Department of Medical Chemistry, Molecular Biology and Pathobiochemistry Pathobiochemistry, Semmelweis University, POB 260, Budapest, 1444, Hungary
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- András Szarka
- Department of Applied Biotechnology and Food Science, Laboratory of Biochemistry and Molecular Biology, Budapest University of Technology and Economics, Szent Gellért tér 4, Budapest, 1111, Hungary. .,Pathobiochemistry Research Group of Hungarian Academy of Sciences, Semmelweis University, P.O. Box 260, Budapest, 1444, Hungary.
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11
Ozer HK, Dlouhy AC, Thornton JD, Hu J, Liu Y, Barycki JJ, Balk J, Outten CE. Cytosolic Fe-S Cluster Protein Maturation and Iron Regulation Are Independent of the Mitochondrial Erv1/Mia40 Import System.
J Biol Chem 2015;
290:27829-40. [PMID:
26396185 DOI:
10.1074/jbc.m115.682179]
[Citation(s) in RCA: 19] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2015] [Indexed: 01/08/2023] Open
Abstract
The sulfhydryl oxidase Erv1 partners with the oxidoreductase Mia40 to import cysteine-rich proteins in the mitochondrial intermembrane space. In Saccharomyces cerevisiae, Erv1 has also been implicated in cytosolic Fe-S protein maturation and iron regulation. To investigate the connection between Erv1/Mia40-dependent mitochondrial protein import and cytosolic Fe-S cluster assembly, we measured Mia40 oxidation and Fe-S enzyme activities in several erv1 and mia40 mutants. Although all the erv1 and mia40 mutants exhibited defects in Mia40 oxidation, only one erv1 mutant strain (erv1-1) had significantly decreased activities of cytosolic Fe-S enzymes. Further analysis of erv1-1 revealed that it had strongly decreased glutathione (GSH) levels, caused by an additional mutation in the gene encoding the glutathione biosynthesis enzyme glutamate cysteine ligase (GSH1). To address whether Erv1 or Mia40 plays a role in iron regulation, we measured iron-dependent expression of Aft1/2-regulated genes and mitochondrial iron accumulation in erv1 and mia40 strains. The only strain to exhibit iron misregulation is the GSH-deficient erv1-1 strain, which is rescued with addition of GSH. Together, these results confirm that GSH is critical for cytosolic Fe-S protein biogenesis and iron regulation, whereas ruling out significant roles for Erv1 or Mia40 in these pathways.
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Affiliation(s)
- Hatice K Ozer
- From the Department of Chemistry and Biochemistry, University of South Carolina, Columbia, South Carolina 29208
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- Adrienne C Dlouhy
- From the Department of Chemistry and Biochemistry, University of South Carolina, Columbia, South Carolina 29208
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- Jeremy D Thornton
- the John Innes Centre and University of East Anglia, Norwich Research Park, Norwich NR4 7UH, United Kingdom, and
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- Jingjing Hu
- From the Department of Chemistry and Biochemistry, University of South Carolina, Columbia, South Carolina 29208
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- Yilin Liu
- the Department of Biochemistry and the Redox Biology Center, University of Nebraska, Lincoln, Nebraska 68588
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- Joseph J Barycki
- the Department of Biochemistry and the Redox Biology Center, University of Nebraska, Lincoln, Nebraska 68588
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- Janneke Balk
- the John Innes Centre and University of East Anglia, Norwich Research Park, Norwich NR4 7UH, United Kingdom, and
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- Caryn E Outten
- From the Department of Chemistry and Biochemistry, University of South Carolina, Columbia, South Carolina 29208,
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12
Abstract
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Mitochondria are fundamental intracellular organelles with key
roles in important cellular processes like energy production, Fe/S
cluster biogenesis, and homeostasis of lipids and inorganic ions.
Mitochondrial dysfunction is consequently linked to many human pathologies
(cancer, diabetes, neurodegeneration, stroke) and apoptosis. Mitochondrial
biogenesis relies on protein import as most mitochondrial proteins
(about 10–15% of the human proteome) are imported after their
synthesis in the cytosol. Over the last several years many mitochondrial
translocation pathways have been discovered. Among them, the import
pathway that targets proteins to the intermembrane space (IMS) stands
out as it is the only one that couples import to folding and oxidation
and results in the covalent modification of the incoming precursor
that adopt internal disulfide bonds in the process (the MIA pathway).
The discovery of this pathway represented a significant paradigm shift
as it challenged the prevailing dogma that the endoplasmic reticulum
is the only compartment of eukaryotic cells where oxidative folding
can occur.
The concept of the oxidative folding pathway was
first proposed
on the basis of folding and import data for the small Tim proteins
that have conserved cysteine motifs and must adopt intramolecular
disulfides after import so that they are retained in the organelle.
The introduction of disulfides in the IMS is catalyzed by Mia40 that
functions as a chaperone inducing their folding. The sulfhydryl oxidase
Erv1 generates the disulfide pairs de novo using either molecular
oxygen or, cytochrome c and other proteins as terminal
electron acceptors that eventually link this folding process to respiration.
The solution NMR structure of Mia40 (and supporting biochemical experiments)
showed that Mia40 is a novel type of disulfide donor whose recognition
capacity for its substrates relies on a hydrophobic binding cleft
found adjacent to a thiol active CPC motif. Targeting of the substrates
to this pathway is guided by a novel type of IMS targeting signal
called ITS or MISS. This consists of only 9 amino acids, found upstream
or downstream of a unique Cys that is primed for docking to Mia40
when the substrate is accommodated in the Mia40 binding cleft. Different
routes exist to complete the folding of the substrates and their final
maturation in the IMS. Identification of new Mia40 substrates (some
even without the requirement of their cysteines) reveals an expanded
chaperone-like activity of this protein in the IMS. New evidence on
the targeting of redox active proteins like thioredoxin, glutaredoxin,
and peroxiredoxin into the IMS suggests the presence of redox-dependent
regulatory mechanisms of the protein folding and import process in
mitochondria. Maintenance of redox balance in mitochondria is crucial
for normal cell physiology and depends on the cross-talk between the
various redox signaling processes and the mitochondrial oxidative
folding pathway.
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Affiliation(s)
- Amelia Mordas
- Institute
of Molecular Cell and Systems Biology, College of Medical Veterinary
and Life Sciences, University of Glasgow, Glasgow G12 8QQ, United Kingdom
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- Kostas Tokatlidis
- Institute
of Molecular Cell and Systems Biology, College of Medical Veterinary
and Life Sciences, University of Glasgow, Glasgow G12 8QQ, United Kingdom
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13
Han LH, Dong LY, Yu H, Sun GY, Wu Y, Gao J, Thasler W, An W. Deceleration of liver regeneration by knockdown of augmenter of liver regeneration gene is associated with impairment of mitochondrial DNA synthesis in mice.
Am J Physiol Gastrointest Liver Physiol 2015;
309:G112-22. [PMID:
25977511 DOI:
10.1152/ajpgi.00435.2014]
[Citation(s) in RCA: 17] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/09/2014] [Accepted: 05/11/2015] [Indexed: 01/31/2023]
Abstract
Hepatic stimulator substance, also known as augmenter of liver regeneration (ALR), is a novel hepatic mitogen that stimulates liver regeneration after partial hepatectomy (PH). Recent work has indicated that a lack of ALR expression inhibited liver regeneration in rats, and the mechanism seems to be related to increased cell apoptosis. The mitochondria play an important role during liver regeneration. Adequate ATP supply, which is largely dependent on effective mitochondrial biogenesis, is essential for progress of liver regeneration. However, ALR gene expression during liver regeneration, particularly its function with mitochondrial DNA synthesis, remains poorly understood. In this study, ALR expression in hepatocytes of mice was suppressed with ALR short-hairpin RNA interference or ALR deletion (knockout, KO). The ALR-defective mice underwent PH, and the liver was allowed to regenerate for 1 wk. Analysis of liver growth and its correlation with mitochondrial biogenesis showed that both ALR mRNA and protein levels increased robustly in control mice with a maximum at days 3 and 4 post-PH. However, ALR knockdown inhibited hepatic DNA synthesis and decelerated liver regeneration after PH. Furthermore, both in the ALR-knockdown and ALR-KO mice, expression of mitochondrial transcription factor A and peroxisome proliferator-activated receptor-γ coactivator-1α were reduced, resulting in impaired mitochondrial biogenesis. In conclusion, ALR is apparently required to ensure appropriate liver regeneration following PH in mice, and deletion of the ALR gene may delay liver regeneration in part due to impaired mitochondrial biogenesis.
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Affiliation(s)
- Li-hong Han
- Department of Cell Biology and Municipal Laboratory of Liver Protection and Regulation of Regeneration, Capital Medical University, Beijing, China; and
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- Ling-yue Dong
- Department of Cell Biology and Municipal Laboratory of Liver Protection and Regulation of Regeneration, Capital Medical University, Beijing, China; and
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- Hao Yu
- Department of Cell Biology and Municipal Laboratory of Liver Protection and Regulation of Regeneration, Capital Medical University, Beijing, China; and
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- Guang-yong Sun
- Department of Cell Biology and Municipal Laboratory of Liver Protection and Regulation of Regeneration, Capital Medical University, Beijing, China; and
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- Yuan Wu
- Department of Cell Biology and Municipal Laboratory of Liver Protection and Regulation of Regeneration, Capital Medical University, Beijing, China; and
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- Jian Gao
- Department of Cell Biology and Municipal Laboratory of Liver Protection and Regulation of Regeneration, Capital Medical University, Beijing, China; and
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- Wei An
- Department of Cell Biology and Municipal Laboratory of Liver Protection and Regulation of Regeneration, Capital Medical University, Beijing, China; and
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14
A novel chitin binding crayfish molar tooth protein with elasticity properties.
PLoS One 2015;
10:e0127871. [PMID:
26010981 PMCID:
PMC4444123 DOI:
10.1371/journal.pone.0127871]
[Citation(s) in RCA: 9] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2015] [Accepted: 04/21/2015] [Indexed: 12/04/2022] Open
Abstract
The molar tooth of the crayfish Cherax quadricarinatus is part of the mandible, and is covered by a layer of apatite (calcium phosphate). This tooth sheds and is regenerated during each molting cycle together with the rest of the exoskeleton. We discovered that molar calcification occurs at the pre-molt stage, unlike calcification of the rest of the new exoskeleton. We further identified a novel molar protein from C. quadricarinatus and cloned its transcript from the molar-forming epithelium. We termed this protein Cq-M13. The temporal level of transcription of Cq-M13 in an NGS library of molar-forming epithelium at different molt stages coincides with the assembly and mineralization pattern of the molar tooth. The predicted protein was found to be related to the pro-resilin family of cuticular proteins. Functionally, in vivo silencing of the transcript caused molt cycle delay and a recombinant version of the protein was found to bind chitin and exhibited elastic properties.
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15
Szarka A, Bánhegyi G. Oxidative folding: recent developments.
Biomol Concepts 2015;
2:379-90. [PMID:
25962043 DOI:
10.1515/bmc.2011.038]
[Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/30/2011] [Accepted: 07/21/2011] [Indexed: 01/29/2023] Open
Abstract
Disulfide bond formation in proteins is an effective tool of both structure stabilization and redox regulation. The prokaryotic periplasm and the endoplasmic reticulum of eukaryotes were long considered as the only compartments for enzyme mediated formation of stable disulfide bonds. Recently, the mitochondrial intermembrane space has emerged as the third protein-oxidizing compartment. The classic view on the mechanism of oxidative folding in the endoplasmic reticulum has also been reshaped by new observations. Moreover, besides the structure stabilizing function, reversible disulfide bridge formation in some proteins of the endoplasmic reticulum, seems to play a regulatory role. This review briefly summarizes the present knowledge of the redox systems supporting oxidative folding, emphasizing recent developments.
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16
Abstract
ALR is a mystic protein. It has a so called "long" 22 kDa and a "short" 15 kDa forms. It has been described after partial hepatectomy and it has just been considered as a key protein of liver regeneration. At the beginning of the 21st century it has been revealed that the "long" form is localized in the mitochondrial intermembrane space and it is an element of the mitochondrial protein import and disulphide relay system. Several proteins of the substrates of the mitochondrial disulphide relay system are necessary for the proper function of the mitochondria, thus any mutation of the ALR gene leads to mitochondrial diseases. The "short" form of ALR functions as a secreted extracellular growth factor and it promotes the protection, regeneration and proliferation of hepatocytes. The results gained on the recently generated conditional ALR mutant mice suggest that ALR can play an important role in the pathogenesis of alcoholic and non-alcoholic steatosis. Since the serum level of ALR is modified in several liver diseases it can be a promising marker molecule in laboratory diagnostics.
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Affiliation(s)
- Tibor Balogh
- Budapesti Műszaki és Gazdaságtudományi Egyetem, Vegyészmérnöki és Biomérnöki Kar Alkalmazott Biotechnológia és Élelmiszer-tudományi Tanszék, Biokémiai és Molekuláris Biológiai Laboratórium Budapest
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- András Szarka
- Budapesti Műszaki és Gazdaságtudományi Egyetem, Vegyészmérnöki és Biomérnöki Kar Alkalmazott Biotechnológia és Élelmiszer-tudományi Tanszék, Biokémiai és Molekuláris Biológiai Laboratórium Budapest Semmelweis Egyetem, Általános Orvostudományi Kar Orvosi Vegytani Molekuláris Biológiai és Patobiokémiai Intézet Budapest Pf. 260 1444
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17
Gandhi CR, Chaillet JR, Nalesnik MA, Kumar S, Dangi A, Demetris AJ, Ferrell R, Wu T, Divanovic S, Stankeiwicz T, Shaffer B, Stolz DB, Harvey SAK, Wang J, Starzl TE. Liver-specific deletion of augmenter of liver regeneration accelerates development of steatohepatitis and hepatocellular carcinoma in mice.
Gastroenterology 2015;
148:379-391.e4. [PMID:
25448926 PMCID:
PMC4802363 DOI:
10.1053/j.gastro.2014.10.008]
[Citation(s) in RCA: 78] [Impact Index Per Article: 7.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/21/2014] [Revised: 10/06/2014] [Accepted: 10/07/2014] [Indexed: 12/22/2022]
Abstract
BACKGROUND & AIMS
Augmenter of liver regeneration (ALR, encoded by GFER) is a widely distributed pleiotropic protein originally identified as a hepatic growth factor. However, little is known about its roles in hepatic physiology and pathology. We created mice with liver-specific deletion of ALR to study its function.
METHODS
We developed mice with liver-specific deletion of ALR (ALR-L-KO) using the albumin-Cre/LoxP system. Liver tissues were collected from ALR-L-KO mice and ALR(floxed/floxed) mice (controls) and analyzed by histology, reverse-transcription polymerase chain reaction, immunohistochemistry, electron microscopy, and techniques to measure fibrosis and lipids. Liver tissues from patients with and without advanced liver disease were determined by immunoblot analysis.
RESULTS
Two weeks after birth, livers of ALR-L-KO mice contained low levels of ALR and adenosine triphosphate (ATP); they had reduced mitochondrial respiratory function and increased oxidative stress, compared with livers from control mice, and had excessive steatosis, and hepatocyte apoptosis. Levels of carbamyl-palmitoyl transferase 1a and ATP synthase subunit ATP5G1 were reduced in livers of ALR-L-KO mice, indicating defects in mitochondrial fatty acid transport and ATP synthesis. Electron microscopy showed mitochondrial swelling with abnormalities in shapes and numbers of cristae. From weeks 2-4 after birth, levels of steatosis and apoptosis decreased in ALR-L-KO mice, and numbers of ALR-expressing cells increased, along with ATP levels. However, at weeks 4-8 after birth, livers became inflamed, with hepatocellular necrosis, ductular proliferation, and fibrosis; hepatocellular carcinoma developed by 1 year after birth in nearly 60% of the mice. Hepatic levels of ALR were also low in ob/ob mice and alcohol-fed mice with liver steatosis, compared with controls. Levels of ALR were lower in liver tissues from patients with advanced alcoholic liver disease and nonalcoholic steatohepatitis than in control liver tissues.
CONCLUSIONS
We developed mice with liver-specific deletion of ALR, and showed that it is required for mitochondrial function and lipid homeostasis in the liver. ALR-L-KO mice provide a useful model for investigating the pathogenesis of steatohepatitis and its complications.
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Affiliation(s)
- Chandrashekhar R Gandhi
- Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio; Department of Surgery, University of Cincinnati, Cincinnati, Ohio; Cincinnati VA Medical Center, Cincinnati, Ohio; Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh, Pennsylvania.
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- J Richard Chaillet
- Department of Microbiology and Molecular Genetics, University of Pittsburgh, Pennsylvania
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- Michael A Nalesnik
- Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh, Pennsylvania; Department of Pathology, University of Pittsburgh, Pennsylvania
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- Sudhir Kumar
- Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio
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- Anil Dangi
- Department of Surgery, University of Cincinnati, Cincinnati, Ohio; Cincinnati VA Medical Center, Cincinnati, Ohio
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- A Jake Demetris
- Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh, Pennsylvania; Department of Pathology, University of Pittsburgh, Pennsylvania
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- Robert Ferrell
- School of Public Health, University of Pittsburgh, Pennsylvania
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- Tong Wu
- Department of Pathology and Laboratory Medicine, Tulane University School of Medicine, New Orleans, Louisiana
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- Senad Divanovic
- Division of Immunobiology, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio
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- Traci Stankeiwicz
- Division of Immunobiology, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio
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- Benjamin Shaffer
- Department of Microbiology and Molecular Genetics, University of Pittsburgh, Pennsylvania
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- Donna B Stolz
- Department of Cell Biology, University of Pittsburgh, Pennsylvania
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- Jiang Wang
- Department of Pathology and Laboratory Medicine, University of Cincinnati, Cincinnati, Ohio
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- Thomas E Starzl
- Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh, Pennsylvania
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18
Murcha MW, Kmiec B, Kubiszewski-Jakubiak S, Teixeira PF, Glaser E, Whelan J. Protein import into plant mitochondria: signals, machinery, processing, and regulation.
JOURNAL OF EXPERIMENTAL BOTANY 2014;
65:6301-35. [PMID:
25324401 DOI:
10.1093/jxb/eru399]
[Citation(s) in RCA: 60] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/23/2023]
Abstract
The majority of more than 1000 proteins present in mitochondria are imported from nuclear-encoded, cytosolically synthesized precursor proteins. This impressive feat of transport and sorting is achieved by the combined action of targeting signals on mitochondrial proteins and the mitochondrial protein import apparatus. The mitochondrial protein import apparatus is composed of a number of multi-subunit protein complexes that recognize, translocate, and assemble mitochondrial proteins into functional complexes. While the core subunits involved in mitochondrial protein import are well conserved across wide phylogenetic gaps, the accessory subunits of these complexes differ in identity and/or function when plants are compared with Saccharomyces cerevisiae (yeast), the model system for mitochondrial protein import. These differences include distinct protein import receptors in plants, different mechanistic operation of the intermembrane protein import system, the location and activity of peptidases, the function of inner-membrane translocases in linking the outer and inner membrane, and the association/regulation of mitochondrial protein import complexes with components of the respiratory chain. Additionally, plant mitochondria share proteins with plastids, i.e. dual-targeted proteins. Also, the developmental and cell-specific nature of mitochondrial biogenesis is an aspect not observed in single-celled systems that is readily apparent in studies in plants. This means that plants provide a valuable model system to study the various regulatory processes associated with protein import and mitochondrial biogenesis.
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Affiliation(s)
- Monika W Murcha
- Australian Research Council Centre of Excellence in Plant Energy Biology, University of Western Australia, 35 Stirling Highway, Crawley, Western Australia, 6009, Australia
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- Beata Kmiec
- Department of Biochemistry and Biophysics, Stockholm University, Arrhenius Laboratories for Natural Sciences, SE-10691 Stockholm, Sweden
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- Szymon Kubiszewski-Jakubiak
- Australian Research Council Centre of Excellence in Plant Energy Biology, University of Western Australia, 35 Stirling Highway, Crawley, Western Australia, 6009, Australia
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- Pedro F Teixeira
- Department of Biochemistry and Biophysics, Stockholm University, Arrhenius Laboratories for Natural Sciences, SE-10691 Stockholm, Sweden
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- Elzbieta Glaser
- Department of Biochemistry and Biophysics, Stockholm University, Arrhenius Laboratories for Natural Sciences, SE-10691 Stockholm, Sweden
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- James Whelan
- Australian Research Council Centre of Excellence in Plant Energy Biology, School of Life Science, La Trobe University, Bundoora, Victoria, 3086, Australia
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19
Khandoga A, Mende K, Iskandarov E, Rosentreter D, Schelcher C, Reifart J, Jauch KW, Thasler WE. Augmenter of liver regeneration attenuates inflammatory response in the postischemic mouse liver in vivo.
J Surg Res 2014;
192:187-94. [DOI:
10.1016/j.jss.2014.05.026]
[Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2014] [Revised: 05/02/2014] [Accepted: 05/13/2014] [Indexed: 12/12/2022]
20
Yu HY, Zhu MH, Xiang DR, Li J, Sheng JF. High expression of 23 kDa protein of augmenter of liver regeneration (ALR) in human hepatocellular carcinoma.
Onco Targets Ther 2014;
7:887-93. [PMID:
24940072 PMCID:
PMC4051792 DOI:
10.2147/ott.s61531]
[Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Background
Augmenter of liver regeneration (ALR) is an important polypeptide that participates in the process of liver regeneration. Two forms of ALR proteins are expressed in hepatocytes. Previous data have shown that ALR is essential for cell survival and has potential antimetastatic properties in hepatocellular carcinoma (HCC).
Aims
The study aimed to evaluate the expression levels of two forms of ALR proteins in HCC and their possible significance in HCC development.
Methods
Balb/c mouse monoclonal antibody against ALR protein was prepared in order to detect the ALR protein in HCC by Western blotting and immunohistochemistry. ALR mRNA expression levels were measured by real-time polymerase chain reaction in HCC tissues and compared to paracancerous liver tissues in 22 HCC patients.
Results
ALR mRNA expression in HCC liver tissues (1.51×106 copies/μL) was higher than in paracancerous tissues (1.04×104 copies/μL). ALR protein expression was also enhanced in HCC liver tissues. The enhanced ALR protein was shown to be 23 kDa by Western blotting. Immunohistochemical analysis showed that the 23 kDa ALR protein mainly existed in the hepatocyte cytosol.
Conclusion
The 23 kDa ALR protein was highly expressed in HCC and may play an important role in hepatocarcinogenesis.
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Affiliation(s)
- Hai-Ying Yu
- State Key Laboratory of Infectious Disease and Department of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China
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- Man-Hua Zhu
- State Key Laboratory of Infectious Disease and Department of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China
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- Dai-Rong Xiang
- State Key Laboratory of Infectious Disease and Department of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China
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- Jun Li
- State Key Laboratory of Infectious Disease and Department of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China
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- Ji-Fang Sheng
- State Key Laboratory of Infectious Disease and Department of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China
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21
Kallergi E, Kalef-Ezra E, Karagouni-Dalakoura K, Tokatlidis K. Common Players in Mitochondria Biogenesis and Neuronal Protection Against Stress-Induced Apoptosis.
Neurochem Res 2013;
39:546-55. [DOI:
10.1007/s11064-013-1109-x]
[Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2013] [Revised: 07/01/2013] [Accepted: 07/08/2013] [Indexed: 10/26/2022]
22
Chatzi A, Tokatlidis K. The mitochondrial intermembrane space: a hub for oxidative folding linked to protein biogenesis.
Antioxid Redox Signal 2013;
19:54-62. [PMID:
22901034 DOI:
10.1089/ars.2012.4855]
[Citation(s) in RCA: 26] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
SIGNIFICANCE
The introduction of disulfide bonds in proteins of the mitochondrial intermembrane space (IMS) is fundamental for their folding and assembly. This oxidative folding process depends on the disulfide donor/import receptor Mia40 and the flavin adenine dinucleotide oxidase Erv1 and concerns proteins involved in mitochondrial biogenesis, respiratory complex assembly, and metal transfer.
RECENT ADVANCES
The recently determined structural basis of the interaction between Mia40 and some substrates provides a framework for the electron transfer process. A possible proofreading role for the cellular reductant glutathione has been proposed, while other studies suggest the association of Mia40 and Erv1 in dynamic multiprotein complexes in the IMS.
CRITICAL ISSUES
The association of Mia40 with Erv1 and substrates in large multiprotein complexes is critical. Completion of substrate folding by additional disulfide bonds after initial binding to Mia40 remains unclear. Furthermore, a more general role for Mia40 in recognizing substrates targeted to other compartments, or even without specific cysteine motifs, remains an intriguing possibility.
FUTURE DIRECTIONS
Dissecting a regulatory role of intramitochondrial protein complex organization and small redox-active molecules will be crucial for understanding oxidative folding in the IMS. This should have an impact on the physiology of human cells, as disease-linked mutations of key components of this process have been manifested, and their expression in stem cells appears crucial for development.
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Affiliation(s)
- Afroditi Chatzi
- Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology Hellas IMBB-FORTH, Heraklion, Greece
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23
Napoli E, Wong S, Hung C, Ross-Inta C, Bomdica P, Giulivi C. Defective mitochondrial disulfide relay system, altered mitochondrial morphology and function in Huntington's disease.
Hum Mol Genet 2013;
22:989-1004. [PMID:
23197653 PMCID:
PMC8482967 DOI:
10.1093/hmg/dds503]
[Citation(s) in RCA: 66] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2012] [Revised: 11/07/2012] [Accepted: 11/26/2012] [Indexed: 01/09/2024] Open
Abstract
A number of studies have been conducted that link mitochondrial dysfunction (MD) to Huntington's disease (HD); however, contradicting results had resulted in a lack of a clear mechanism that links expression of mutant Huntingtin protein and MD. Mouse homozygous (HM) and heterozygous (HT) mutant striatal cells with two or one allele encoding for a mutant huntingtin protein with 111 polyGln repeats showed a significant impairment of the mitochondrial disulfide relay system (MDRS). This system (consisting of two proteins, Gfer and Mia40) is involved in the mitochondrial import of Cys-rich proteins. The Gfer-to-Mia40 ratio was significantly altered in HM cells compared with controls, along with the expression of mitochondrial proteins considered substrates of the MDRS. In progenitors and differentiated neuron-like HM cells, impairment of MDRS were accompanied by deficient oxidative phosphorylation, Complex I, IV and V activities, decreased mtDNA copy number and transcripts, accumulation of mtDNA deletions and changes in mitochondrial morphology, consistent with other MDRS-deficient biological models, thus providing a framework for the energy deficits observed in this HD model. The majority (>90%) of the mitochondrial outcomes exhibited a gene-dose dependency with the expression of mutant Htt. Finally, decreases in the mtDNA copy number, along with the accumulation of mtDNA deletions, provide a mechanism for the progressive neurodegeneration observed in HD patients.
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Affiliation(s)
- Eleonora Napoli
- Department of Molecular Biosciences, University of California
Davis, Davis, CA 95616, USA
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- Sarah Wong
- Department of Molecular Biosciences, University of California
Davis, Davis, CA 95616, USA
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- Connie Hung
- Department of Molecular Biosciences, University of California
Davis, Davis, CA 95616, USA
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- Catherine Ross-Inta
- Department of Molecular Biosciences, University of California
Davis, Davis, CA 95616, USA
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- Prithvi Bomdica
- Department of Molecular Biosciences, University of California
Davis, Davis, CA 95616, USA
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- Cecilia Giulivi
- Department of Molecular Biosciences, University of California
Davis, Davis, CA 95616, USA
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24
Banci L, Bertini I, Cefaro C, Ciofi-Baffoni S, Gajda K, Felli IC, Gallo A, Pavelkova A, Kallergi E, Andreadaki M, Katrakili N, Pozidis C, Tokatlidis K. An intrinsically disordered domain has a dual function coupled to compartment-dependent redox control.
J Mol Biol 2013;
425:594-608. [PMID:
23207295 DOI:
10.1016/j.jmb.2012.11.032]
[Citation(s) in RCA: 19] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2012] [Revised: 11/03/2012] [Accepted: 11/25/2012] [Indexed: 11/20/2022]
Abstract
The functional role of unstructured protein domains is an emerging field in the frame of intrinsically disordered proteins. The involvement of intrinsically disordered domains (IDDs) in protein targeting and biogenesis processes in mitochondria is so far not known. Here, we have characterized the structural/dynamic and functional properties of an IDD of the sulfhydryl oxidase ALR (augmenter of liver regeneration) located in the intermembrane space of mitochondria. At variance to the unfolded-to-folded structural transition of several intrinsically disordered proteins, neither substrate recognition events nor redox switch of its shuttle cysteine pair is linked to any such structural change. However, this unstructured domain performs a dual function in two cellular compartments: it acts (i) as a mitochondrial targeting signal in the cytosol and (ii) as a crucial recognition site in the disulfide relay system of intermembrane space. This domain provides an exciting new paradigm for IDDs ensuring two distinct functions that are linked to intracellular organelle targeting.
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Affiliation(s)
- Lucia Banci
- Magnetic Resonance Center (CERM), University of Florence, Via Luigi Sacconi 6, 50019 Sesto Fiorentino, Florence, Italy.
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25
Gad W, Nair MG, Van Belle K, Wahni K, De Greve H, Van Ginderachter JA, Vandenbussche G, Endo Y, Artis D, Messens J. The quiescin sulfhydryl oxidase (hQSOX1b) tunes the expression of resistin-like molecule alpha (RELM-α or mFIZZ1) in a wheat germ cell-free extract.
PLoS One 2013;
8:e55621. [PMID:
23383248 PMCID:
PMC3561318 DOI:
10.1371/journal.pone.0055621]
[Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2012] [Accepted: 01/02/2013] [Indexed: 11/19/2022] Open
Abstract
BACKGROUND
Although disulfide bond formation in proteins is one of the most common types of post-translational modifications, the production of recombinant disulfide-rich proteins remains a challenge. The most popular host for recombinant protein production is Escherichia coli, but disulfide-rich proteins are here often misfolded, degraded, or found in inclusion bodies.
METHODOLOGY/PRINCIPAL FINDINGS
We optimize an in vitro wheat germ translation system for the expression of an immunological important eukaryotic protein that has to form five disulfide bonds, resistin-like alpha (mFIZZ1). Expression in combination with human quiescin sulfhydryl oxidase (hQSOX1b), the disulfide bond-forming enzyme of the endoplasmic reticulum, results in soluble, intramolecular disulfide bonded, monomeric, and biological active protein. The mFIZZ1 protein clearly suppresses the production of the cytokines IL-5 and IL-13 in mouse splenocytes cultured under Th2 permissive conditions.
CONCLUSION/SIGNIFICANCE
The quiescin sulfhydryl oxidase hQSOX1b seems to function as a chaperone and oxidase during the oxidative folding. This example for mFIZZ1 should encourage the design of an appropriate thiol/disulfide oxidoreductase-tuned cell free expression system for other challenging disulfide rich proteins.
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Affiliation(s)
- Wael Gad
- Brussels Center for Redox Biology, Brussels, Belgium
- Department of Structural Biology, Vlaams Instituut voor Biotechnologie, Brussels, Belgium
- Structural Biology Brussels, Vrije Universiteit Brussel, Brussels, Belgium
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- Meera G. Nair
- Department of Microbiology and Institute for Immunology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America
- Division of Biomedical Sciences, School of Medicine, University of California Riverside, Riverside, California, United States of America
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- Karolien Van Belle
- Brussels Center for Redox Biology, Brussels, Belgium
- Department of Structural Biology, Vlaams Instituut voor Biotechnologie, Brussels, Belgium
- Structural Biology Brussels, Vrije Universiteit Brussel, Brussels, Belgium
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- Khadija Wahni
- Brussels Center for Redox Biology, Brussels, Belgium
- Department of Structural Biology, Vlaams Instituut voor Biotechnologie, Brussels, Belgium
- Structural Biology Brussels, Vrije Universiteit Brussel, Brussels, Belgium
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- Henri De Greve
- Department of Structural Biology, Vlaams Instituut voor Biotechnologie, Brussels, Belgium
- Structural Biology Brussels, Vrije Universiteit Brussel, Brussels, Belgium
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- Jo A. Van Ginderachter
- Lab of Cellular and Molecular Immunology, Vrije Universiteit Brussel, Brussels, Belgium
- Myeloid Cell Immunology Lab, Vlaams Instituut voor Biotechnologie, Brussels, Belgium
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- Guy Vandenbussche
- Centre de Biologie Structurale et de Bioinformatique, Structure et Fonction des Membranes Biologiques, Université Libre de Bruxelles, Brussels, Belgium
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- Yaeta Endo
- Cell Free Science and Technology Research Center, Ehime University, Matsuyama, Japan
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- David Artis
- Department of Microbiology and Institute for Immunology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America
- Department of Pathobiology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America
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- Joris Messens
- Brussels Center for Redox Biology, Brussels, Belgium
- Department of Structural Biology, Vlaams Instituut voor Biotechnologie, Brussels, Belgium
- Structural Biology Brussels, Vrije Universiteit Brussel, Brussels, Belgium
- * E-mail:
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26
Fraga H, Ventura S. Oxidative folding in the mitochondrial intermembrane space in human health and disease.
Int J Mol Sci 2013;
14:2916-27. [PMID:
23364613 PMCID:
PMC3588022 DOI:
10.3390/ijms14022916]
[Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2012] [Revised: 01/21/2013] [Accepted: 01/23/2013] [Indexed: 11/22/2022] Open
Abstract
Oxidative folding in the mitochondrial intermembrane space (IMS) is a key cellular event associated with the folding and import of a large and still undetermined number of proteins. This process is catalyzed by an oxidoreductase, Mia40 that is able to recognize substrates with apparently little or no homology. Following substrate oxidation, Mia40 is reduced and must be reoxidized by Erv1/Alr1 that consequently transfers the electrons to the mitochondrial respiratory chain. Although our understanding of the physiological relevance of this process is still limited, an increasing number of pathologies are being associated with the impairment of this pathway; especially because oxidative folding is fundamental for several of the proteins involved in defense against oxidative stress. Here we review these aspects and discuss recent findings suggesting that oxidative folding in the IMS is modulated by the redox state of the cell.
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Affiliation(s)
- Hugo Fraga
- Department of Biochemistry and Molecular Biology, Autonomous University of Barcelona, Bellaterra E-08193, Spain
- Authors to whom correspondence should be addressed; E-Mails: (H.F.); (S.V.); Tel.: +34-93-581-2154 (H.F.); +34-93-586-8956 (S.V.); Fax: +34-93-581-1264 (H.F. & S.V.)
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- Salvador Ventura
- Department of Biochemistry and Molecular Biology, Autonomous University of Barcelona, Bellaterra E-08193, Spain
- Institute of Biotechnology and Biomedicine, Autonomous University of Barcelona, Bellaterra E-08193, Spain
- Authors to whom correspondence should be addressed; E-Mails: (H.F.); (S.V.); Tel.: +34-93-581-2154 (H.F.); +34-93-586-8956 (S.V.); Fax: +34-93-581-1264 (H.F. & S.V.)
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27
Vodovotz Y, Prelich J, Lagoa C, Barclay D, Zamora R, Murase N, Gandhi CR. Augmenter of liver regeneration (ALR) is a novel biomarker of hepatocellular stress/inflammation: in vitro, in vivo and in silico studies.
Mol Med 2013;
18:1421-9. [PMID:
23073658 PMCID:
PMC3563711 DOI:
10.2119/molmed.2012.00183]
[Citation(s) in RCA: 25] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2012] [Accepted: 10/09/2012] [Indexed: 12/15/2022] Open
Abstract
The liver is a central organ involved in inflammatory processes, including the elaboration of acute-phase proteins. Augmenter of liver regeneration (ALR) protein, expressed and secreted by hepatocytes, promotes liver regeneration and maintains viability of hepatocytes. ALR also stimulates secretion of inflammatory cytokines (tumor necrosis factor [TNF]-α and interleukin [IL]-6) and nitric oxide from Kupffer cells. We hypothesized that ALR may be involved in modulating inflammation induced by various stimuli. We found that hepatic ALR levels are elevated at 24 h, before or about the same time as an increase in the mRNA expression of TNF-α and IL-6, after portacaval shunt surgery in rats. Serum ALR also increased, but significantly only on d 4 when pathological changes in the liver become apparent. In rats, serum ALR was elevated after intraperitoneal administration of lipopolysaccharide alone and in a model of gram-negative sepsis. Serum ALR increased before alanine aminotransferase (ALT) in endotoxemia and in the same general time frame as TNF-α and IL-6 in the bacterial sepsis model. Furthermore, mathematical prediction of tissue damage correlated strongly with alterations in serum ALR in a mouse model of hemorrhagic shock. In vitro, monomethyl sulfonate, TNF-α, actinomycin D and lipopolysaccharide all caused increased release of ALR from rat hepatocytes, which preceded the loss of cell viability and/or inhibition of DNA synthesis. ALR may thus serve as a potential diagnostic marker of hepatocellular stress and/or acute inflammatory conditions.
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Affiliation(s)
- Yoram Vodovotz
- Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America
- Center for Inflammation and Regenerative Modeling, McGowan Institute for Regenerative Medicine, Pittsburgh, Pennsylvania, United States of America
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- John Prelich
- VA Pittsburgh Healthcare System, Pittsburgh, Pennsylvania, United States of America
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- Claudio Lagoa
- Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America
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- Derek Barclay
- Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America
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- Ruben Zamora
- Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America
- Center for Inflammation and Regenerative Modeling, McGowan Institute for Regenerative Medicine, Pittsburgh, Pennsylvania, United States of America
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- Noriko Murase
- Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America
- Thomas E. Starzl Transplantation Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America
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- Chandrashekhar R Gandhi
- Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America
- Department of Pathology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America
- Thomas E. Starzl Transplantation Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America
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28
Sztolsztener ME, Brewinska A, Guiard B, Chacinska A. Disulfide bond formation: sulfhydryl oxidase ALR controls mitochondrial biogenesis of human MIA40.
Traffic 2012. [PMID:
23186364 DOI:
10.1111/tra.12030]
[Citation(s) in RCA: 45] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
The conserved MIA pathway is responsible for the import and oxidative folding of proteins destined for the intermembrane space of mitochondria. In contrast to a wealth of information obtained from studies with yeast, the function of the MIA pathway in higher eukaryotes has remained enigmatic. Here, we took advantage of the molecular understanding of the MIA pathway in yeast and designed a model of the human MIA pathway. The yeast model for MIA consists of two critical components, the disulfide bond carrier Mia40 and sulfhydryl oxidase Erv1/ALR. Human MIA40 and ALR substituted for their yeast counterparts in the essential function for the oxidative biogenesis of mitochondrial intermembrane space proteins. In addition, the sulfhydryl oxidases ALR/Erv1 were found to be involved in the mitochondrial localization of human MIA40. Furthermore, the defective accumulation of human MIA40 in mitochondria underlies a recently identified disease that is caused by amino acid exchange in ALR. Thus, human ALR is an important factor that controls not only the ability of MIA40 to bind and oxidize protein clients but also the localization of human MIA40 in mitochondria.
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Affiliation(s)
- Malgorzata E Sztolsztener
- International Institute of Molecular and Cell Biology, Laboratory of Mitochondrial Biogenesis Warsaw, 02-109, Poland
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29
Gandhi CR. Augmenter of liver regeneration.
FIBROGENESIS & TISSUE REPAIR 2012;
5:10. [PMID:
22776437 PMCID:
PMC3519801 DOI:
10.1186/1755-1536-5-10]
[Citation(s) in RCA: 54] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/26/2012] [Accepted: 06/26/2012] [Indexed: 11/10/2022]
Abstract
'Augmenter of liver regeneration' (ALR) (also known as hepatic stimulatory substance or hepatopoietin) was originally found to promote growth of hepatocytes in the regenerating or injured liver. ALR is expressed ubiquitously in all organs, and exclusively in hepatocytes in the liver. ALR, a survival factor for hepatocytes, exhibits significant homology with ERV1 (essential for respiration and viability) protein that is essential for the survival of the yeast, Saccharomyces cerevisiae. ALR comprises 198 to 205 amino acids (approximately 22 kDa), but is post-translationally modified to three high molecular weight species (approximately 38 to 42 kDa) found in hepatocytes. ALR is present in mitochondria, cytosol, endoplasmic reticulum, and nucleus. Mitochondrial ALR may be involved in oxidative phosphorylation, but also functions as sulfhydryl oxidase and cytochrome c reductase, and causes Fe/S maturation of proteins. ALR, secreted by hepatocytes, stimulates synthesis of TNF-α, IL-6, and nitric oxide in Kupffer cells via a G-protein coupled receptor. While the 22 kDa rat recombinant ALR does not stimulate DNA synthesis in hepatocytes, the short form (15 kDa) of human recombinant ALR was reported to be equipotent as or even stronger than TGF-α or HGF as a mitogen for hepatocytes. Altered serum ALR levels in certain pathological conditions suggest that it may be a diagnostic marker for liver injury/disease. Although ALR appears to have multiple functions, the knowledge of its role in various organs, including the liver, is extremely inadequate, and it is not known whether different ALR species have distinct functions. Future research should provide better understanding of the expression and functions of this enigmatic molecule.
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30
Stojanovski D, Bragoszewski P, Chacinska A. The MIA pathway: a tight bond between protein transport and oxidative folding in mitochondria.
BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 2012;
1823:1142-50. [PMID:
22579494 DOI:
10.1016/j.bbamcr.2012.04.014]
[Citation(s) in RCA: 64] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/21/2012] [Revised: 04/25/2012] [Accepted: 04/26/2012] [Indexed: 11/29/2022]
Abstract
Many newly synthesized proteins obtain disulfide bonds in the bacterial periplasm, the endoplasmic reticulum (ER) and the mitochondrial intermembrane space. The acquisition of disulfide bonds is critical for the folding, assembly and activity of these proteins. Spontaneous oxidation of thiol groups is inefficient in vivo, therefore cells have developed machineries that catalyse the oxidation of substrate proteins. The identification of the machinery that mediates this process in the intermembrane space of mitochondria, known as MIA (mitochondrial intermembrane space assembly), provided a unique mechanism of protein transport. The MIA machinery introduces disulfide bonds into incoming intermembrane space precursors and thus tightly couples the process of precursor translocation to precursor oxidation. We discuss our current understanding of the MIA pathway and the mechanisms that oversee thiol-exchange reactions in mitochondria.
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Affiliation(s)
- Diana Stojanovski
- La Trobe Institute for Molecular Sciences, 3086 Melbourne, Australia
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31
Duncan O, Murcha MW, Whelan J. Unique components of the plant mitochondrial protein import apparatus.
BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 2012;
1833:304-13. [PMID:
22406071 DOI:
10.1016/j.bbamcr.2012.02.015]
[Citation(s) in RCA: 44] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/04/2012] [Revised: 02/21/2012] [Accepted: 02/23/2012] [Indexed: 10/28/2022]
Abstract
The basic mitochondrial protein import apparatus was established in the earliest eukaryotes. Over the subsequent course of evolution and the divergence of the plant, animal and fungal lineages, this basic import apparatus has been modified and expanded in order to meet the specific needs of protein import in each kingdom. In the plant kingdom, the arrival of the plastid complicated the process of protein trafficking and is thought to have given rise to the evolution of a number of unique components that allow specific and efficient targeting of mitochondrial proteins from their site of synthesis in the cytosol, to their final location in the organelle. This includes the evolution of two unique outer membrane import receptors, plant Translocase of outer membrane 20 kDa subunit (TOM20) and Outer membrane protein of 64 kDa (OM64), the loss of a receptor domain from an ancestral import component, Translocase of outer membrane 22 kDa subunit (TOM22), evolution of unique features in the disulfide relay system of the inter membrane space, and the addition of an extra membrane spanning domain to another ancestral component of the inner membrane, Translocase of inner membrane 17 kDa subunit (TIM17). Notably, many of these components are encoded by multi-gene families and exhibit differential sub-cellular localisation and functional specialisation. This article is part of a Special Issue entitled: Protein Import and Quality Control in Mitochondria and Plastids.
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Affiliation(s)
- Owen Duncan
- ARC Centre of Excellence in Plant Energy Biology, Bayliss Building M316, The University of Western Australia, 35 Stirling Hwy, Crawley, WA 6009, Australia
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32
Li Y, Farooq M, Sheng D, Chandramouli C, Lan T, Mahajan NK, Kini RM, Hong Y, Lisowsky T, Ge R. Augmenter of liver regeneration (alr) promotes liver outgrowth during zebrafish hepatogenesis.
PLoS One 2012;
7:e30835. [PMID:
22292055 PMCID:
PMC3266923 DOI:
10.1371/journal.pone.0030835]
[Citation(s) in RCA: 28] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2011] [Accepted: 12/29/2011] [Indexed: 02/06/2023] Open
Abstract
Augmenter of Liver Regeneration (ALR) is a sulfhydryl oxidase carrying out fundamental functions facilitating protein disulfide bond formation. In mammals, it also functions as a hepatotrophic growth factor that specifically stimulates hepatocyte proliferation and promotes liver regeneration after liver damage or partial hepatectomy. Whether ALR also plays a role during vertebrate hepatogenesis is unknown. In this work, we investigated the function of alr in liver organogenesis in zebrafish model. We showed that alr is expressed in liver throughout hepatogenesis. Knockdown of alr through morpholino antisense oligonucleotide (MO) leads to suppression of liver outgrowth while overexpression of alr promotes liver growth. The small-liver phenotype in alr morphants results from a reduction of hepatocyte proliferation without affecting apoptosis. When expressed in cultured cells, zebrafish Alr exists as dimer and is localized in mitochondria as well as cytosol but not in nucleus or secreted outside of the cell. Similar to mammalian ALR, zebrafish Alr is a flavin-linked sulfhydryl oxidase and mutation of the conserved cysteine in the CxxC motif abolishes its enzymatic activity. Interestingly, overexpression of either wild type Alr or enzyme-inactive Alr(C131S) mutant promoted liver growth and rescued the liver growth defect of alr morphants. Nevertheless, alr(C131S) is less efficacious in both functions. Meantime, high doses of alr MOs lead to widespread developmental defects and early embryonic death in an alr sequence-dependent manner. These results suggest that alr promotes zebrafish liver outgrowth using mechanisms that are dependent as well as independent of its sulfhydryl oxidase activity. This is the first demonstration of a developmental role of alr in vertebrate. It exemplifies that a low-level sulfhydryl oxidase activity of Alr is essential for embryonic development and cellular survival. The dose-dependent and partial suppression of alr expression through MO-mediated knockdown allows the identification of its late developmental role in vertebrate liver organogenesis.
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Affiliation(s)
- Yan Li
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
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- Muhammad Farooq
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
- Department of Zoology, College of Science, King Saud University, Riyadh, Kingdom of Saudi Arabia
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- Donglai Sheng
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
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- Chanchal Chandramouli
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
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- Tian Lan
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
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- Nilesh K. Mahajan
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
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- R. Manjunatha Kini
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
- Department of Biochemistry and Molecular Biology, Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia, United States of America
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- Yunhan Hong
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
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- Ruowen Ge
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
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33
Structure of a baculovirus sulfhydryl oxidase, a highly divergent member of the erv flavoenzyme family.
J Virol 2011;
85:9406-13. [PMID:
21752922 DOI:
10.1128/jvi.05149-11]
[Citation(s) in RCA: 17] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/24/2023] Open
Abstract
Genomes of nucleocytoplasmic large DNA viruses (NCLDVs) encode enzymes that catalyze the formation of disulfide bonds between cysteine amino acid residues in proteins, a function essential for the proper assembly and propagation of NCLDV virions. Recently, a catalyst of disulfide formation was identified in baculoviruses, a group of large double-stranded DNA viruses considered phylogenetically distinct from NCLDVs. The NCLDV and baculovirus disulfide catalysts are flavin adenine dinucleotide (FAD)-binding sulfhydryl oxidases related to the cellular Erv enzyme family, but the baculovirus enzyme, the product of the Ac92 gene in Autographa californica multiple nucleopolyhedrovirus (AcMNPV), is highly divergent at the amino acid sequence level. The crystal structure of the Ac92 protein presented here shows a configuration of the active-site cysteine residues and bound cofactor similar to that observed in other Erv sulfhydryl oxidases. However, Ac92 has a complex quaternary structural arrangement not previously seen in cellular or viral enzymes of this family. This novel assembly comprises a dimer of pseudodimers with a striking 40-degree kink in the interface helix between subunits. The diversification of the Erv sulfhydryl oxidase enzymes in large double-stranded DNA viruses exemplifies the extreme degree to which these viruses can push the boundaries of protein family folds.
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34
Abstract
The identification of protein disulfide isomerase, almost 50 years ago, opened the way to the study of oxidative protein folding. Oxidative protein folding refers to the composite process by which a protein recovers both its native structure and its native disulfide bonds. Pathways that form disulfide bonds have now been unraveled in the bacterial periplasm (disulfide bond protein A [DsbA], DsbB, DsbC, DsbG, and DsbD), the endoplasmic reticulum (protein disulfide isomerase and Ero1), and the mitochondrial intermembrane space (Mia40 and Erv1). This review summarizes the current knowledge on disulfide bond formation in both prokaryotes and eukaryotes and highlights the major problems that remain to be solved.
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Affiliation(s)
- Matthieu Depuydt
- de Duve Institute, Université catholique de Louvain, Brussels, Belgium
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35
Shouldice SR, Heras B, Walden PM, Totsika M, Schembri MA, Martin JL. Structure and function of DsbA, a key bacterial oxidative folding catalyst.
Antioxid Redox Signal 2011;
14:1729-60. [PMID:
21241169 DOI:
10.1089/ars.2010.3344]
[Citation(s) in RCA: 86] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/17/2023]
Abstract
Since its discovery in 1991, the bacterial periplasmic oxidative folding catalyst DsbA has been the focus of intense research. Early studies addressed why it is so oxidizing and how it is maintained in its less stable oxidized state. The crystal structure of Escherichia coli DsbA (EcDsbA) revealed that the oxidizing periplasmic enzyme is a distant evolutionary cousin of the reducing cytoplasmic enzyme thioredoxin. Recent significant developments have deepened our understanding of DsbA function, mechanism, and interactions: the structure of the partner membrane protein EcDsbB, including its complex with EcDsbA, proved a landmark in the field. Studies of DsbA machineries from bacteria other than E. coli K-12 have highlighted dramatic differences from the model organism, including a striking divergence in redox parameters and surface features. Several DsbA structures have provided the first clues to its interaction with substrates, and finally, evidence for a central role of DsbA in bacterial virulence has been demonstrated in a range of organisms. Here, we review current knowledge on DsbA, a bacterial periplasmic protein that introduces disulfide bonds into diverse substrate proteins and which may one day be the target of a new class of anti-virulence drugs to treat bacterial infection.
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Affiliation(s)
- Stephen R Shouldice
- Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
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36
Endo T, Yamano K, Kawano S. Structural basis for the disulfide relay system in the mitochondrial intermembrane space.
Antioxid Redox Signal 2010;
13:1359-73. [PMID:
20136511 DOI:
10.1089/ars.2010.3099]
[Citation(s) in RCA: 39] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 10/19/2022]
Abstract
Mitochondria contain two biological membranes. Although reducing agents can diffuse from the cytosol into the intermembrane space (IMS) between the outer and inner mitochondrial membranes, the IMS has a dedicated disulfide relay system to introduce disulfide bonds into mainly small and soluble proteins. This system consists of two essential proteins, a disulfide carrier Tim40/Mia40 and a flavin-dependent sulfhydryl oxidase Erv1, high-resolution structures that have recently become available. Tim40/Mia40 transfers disulfide bonds to newly imported IMS proteins by dithiol/disulfide exchange reactions involving mixed disulfide intermediates. Tight folding by introduction of disulfide bonds prevents egress of these small IMS proteins, resulting in their selective retention in the compartment. After disulfide transfer from Tim40/Mia40 to substrate proteins, Tim40/Mia40 is reoxidized again by Erv1, which is then oxidized by electron transfer to either cytochrome c or molecular oxygen. Here we review the recent advancement of the knowledge on the mechanism of the disulfide relay system in the mitochondrial IMS, especially shedding light on the structural aspects of its components.
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Affiliation(s)
- Toshiya Endo
- Department of Chemistry, Nagoya University, Japan.
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37
Sideris DP, Tokatlidis K. Oxidative protein folding in the mitochondrial intermembrane space.
Antioxid Redox Signal 2010;
13:1189-204. [PMID:
20214493 DOI:
10.1089/ars.2010.3157]
[Citation(s) in RCA: 71] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/13/2022]
Abstract
Disulfide bond formation is a crucial step for oxidative folding and necessary for the acquisition of a protein's native conformation. Introduction of disulfide bonds is catalyzed in specialized subcellular compartments and requires the coordinated action of specific enzymes. The intermembrane space of mitochondria has recently been found to harbor a dedicated machinery that promotes the oxidative folding of substrate proteins by shuttling disulfide bonds. The newly identified oxidative pathway consists of the redox-regulated receptor Mia40 and the sulfhydryl oxidase Erv1. Proteins destined to the intermembrane space are trapped by a disulfide relay mechanism that involves an electron cascade from the incoming substrate to Mia40, then on to Erv1, and finally to molecular oxygen via cytochrome c. This thiol-disulfide exchange mechanism is essential for the import and for maintaining the structural stability of the incoming precursors. In this review we describe the mechanistic parameters that define the interaction and oxidation of the substrate proteins in light of the recent publications in the mitochondrial oxidative folding field.
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Affiliation(s)
- Dionisia P Sideris
- Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology Hellas (IMBB-FORTH), Heraklion, Crete, Greece
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38
Hakim M, Fass D. Cytosolic disulfide bond formation in cells infected with large nucleocytoplasmic DNA viruses.
Antioxid Redox Signal 2010;
13:1261-71. [PMID:
20136503 DOI:
10.1089/ars.2010.3128]
[Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/17/2023]
Abstract
Proteins that have evolved to contain stabilizing disulfide bonds generally fold in a membrane-delimited compartment in the cell [i.e., the endoplasmic reticulum (ER) or the mitochondrial intermembrane space (IMS)]. These compartments contain sulfhydryl oxidase enzymes that catalyze the pairing and oxidation of cysteine residues. In contrast, most proteins in a healthy cytosol are maintained in reduced form through surveillance by NADPH-dependent reductases and the lack of sulfhydryl oxidases. Nevertheless, one of the core functionalities that unify the broad and diverse set of nucleocytoplasmic large DNA viruses (NCLDVs) is the ability to catalyze disulfide formation in the cytosol. The substrates of this activity are proteins that contribute to the assembly, structure, and infectivity of the virions. If the last common ancestor of NCLDVs was present during eukaryogenesis as has been proposed, it is interesting to speculate that viral disulfide bond formation pathways may have predated oxidative protein folding in intracellular organelles.
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Affiliation(s)
- Motti Hakim
- Department of Structural Biology, Weizmann Institute of Science, Rehovot, Israel
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39
Carrie C, Giraud E, Duncan O, Xu L, Wang Y, Huang S, Clifton R, Murcha M, Filipovska A, Rackham O, Vrielink A, Whelan J. Conserved and novel functions for Arabidopsis thaliana MIA40 in assembly of proteins in mitochondria and peroxisomes.
J Biol Chem 2010;
285:36138-48. [PMID:
20829360 DOI:
10.1074/jbc.m110.121202]
[Citation(s) in RCA: 91] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
The disulfide relay system of the mitochondrial intermembrane space has been extensively characterized in Saccharomyces cerevisiae. It contains two essential components, Mia40 and Erv1. The genome of Arabidopsis thaliana contains a single gene for each of these components. Although insertional inactivation of Erv1 leads to a lethal phenotype, inactivation of Mia40 results in no detectable deleterious phenotype. A. thaliana Mia40 is targeted to and accumulates in mitochondria and peroxisomes. Inactivation of Mia40 results in an alteration of several proteins in mitochondria, an absence of copper/zinc superoxide dismutase (CSD1), the chaperone for superoxide dismutase (Ccs1) that inserts copper into CSD1, and a decrease in capacity and amount of complex I. In peroxisomes the absence of Mia40 leads to an absence of CSD3 and a decrease in abnormal inflorescence meristem 1 (Aim1), a β-oxidation pathway enzyme. Inactivation of Mia40 leads to an alteration of the transcriptome of A. thaliana, with genes encoding peroxisomal proteins, redox functions, and biotic stress significantly changing in abundance. Thus, the mechanistic operation of the mitochondrial disulfide relay system is different in A. thaliana compared with other systems, and Mia40 has taken on new roles in peroxisomes and mitochondria.
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Affiliation(s)
- Chris Carrie
- Australian Research Council Centre of Excellence in Plant Energy Biology, University of Western Australia, Crawley, Western Australia 6009, Australia
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40
Dayoub R, Groitl P, Dobner T, Bosserhoff AK, Schlitt HJ, Weiss TS. Foxa2 (HNF-3beta) regulates expression of hepatotrophic factor ALR in liver cells.
Biochem Biophys Res Commun 2010;
395:465-70. [PMID:
20382118 DOI:
10.1016/j.bbrc.2010.04.023]
[Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2010] [Accepted: 04/03/2010] [Indexed: 12/11/2022]
Abstract
Liver regeneration is a multistep and well-orchestrated process which is initiated by injuries such as tissue loss, infectious or toxic insults. Augmenter of liver regeneration (ALR) is a hepatotrophic growth factor which has been shown to stimulate hepatic regeneration after partial hepatectomy and therefore seems to be regulated during the regenerative process in the liver. Our aim was to analyze how ALR is regulated in hepatic tissues and which transcription factors might regulate its tissue-specific expression. Promoter studies of ALR (-733/+527 bp) revealed potential regulatory elements for various transcription factors like Foxa2, IL-6 RE-BP and C/EBPbeta. Analysis of the promoter activity by performing luciferase assays revealed that co-transfection with Foxa2 significantly induced the activity of ALR promoter in HepG2 cells. EMSA and Supershift analysis using anti-Foxa2 antibody confirmed the specific binding of Foxa2 to ALR promoter and this binding was inducible when the cells were simultaneously stimulated with IL-6. The increased binding after activation with IL-6 and/or Foxa2 was confirmed by elevated ALR protein levels using Western blot technique. In addition, we could not detect any binding of C/EBPbeta and IL-6 RE-BP to the promoter of ALR. In conclusion, these results indicate that ALR is regulated by Foxa2, and this regulation may be amplified by IL-6.
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Affiliation(s)
- Rania Dayoub
- Center for Liver Cell Research, University Medical Center Regensburg, Germany
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41
Gandhi CR, Murase N, Starzl TE. Cholera toxin-sensitive GTP-binding protein-coupled activation of augmenter of liver regeneration (ALR) receptor and its function in rat kupffer cells.
J Cell Physiol 2010;
222:365-73. [PMID:
19859909 PMCID:
PMC3034370 DOI:
10.1002/jcp.21957]
[Citation(s) in RCA: 26] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Mitogenic effect of augmenter of liver regeneration (ALR), a protein produced and released by hepatocytes, on hepatocytes in vivo but not in vitro suggests that the effect is mediated by nonparenchymal cells. Since mediators produced by Kupffer cells are implicated in hepatic regeneration, we investigated receptor for ALR and its functions in rat Kupffer cells. Kupffer cells were isolated from rat liver by enzymatic digestion and centrifugal elutriation. Radioligand ([(125)I] ALR) receptor binding, ALR-induced GTP/G-protein association, and nitric oxide (NO), tumor necrosis factor (TNF)-alpha, and interleukin-6 (IL-6) synthesis were determined. High-affinity receptor for ALR, belonging to the G-protein family, with K(d) of 1.25 +/- 0.18 nM and B(max) of 0.26 +/- 0.02 fmol/microg DNA was identified. ALR stimulated NO, TNF-alpha, and IL-6 synthesis via cholera toxin-sensitive G-protein, as well as p38-MAPK activity and nuclear translocation of NFkappaB. While inhibitor of NFkappaB (MG132) inhibited ALR-induced NO synthesis, MG132 and p38-MAPK inhibitor (SB203580) abrogated ALR-induced TNF-alpha and IL-6 synthesis. ALR also prevented the release of mediator(s) from Kupffer cells that cause inhibition of DNA synthesis in hepatocytes. Administration of ALR to 40% partially hepatectomized rats increased expression of TNF-alpha, IL-6, and inducible nitric oxide synthase (iNOS) and caused augmentation of hepatic regeneration. These results demonstrate specific G-protein coupled binding of ALR and its function in Kupffer cells and suggest that mediators produced by ALR-stimulated Kupffer cells may elicit physiologically important effects on hepatocytes.
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Affiliation(s)
- Chandrashekhar R. Gandhi
- VA Pittsburgh Healthcare System, Pittsburgh, Pennsylvania
- Department of Surgery, Thomas E. Starzl Transplantation Institute, University of Pittsburgh, Pittsburgh, Pennsylvania
- Department of Pathology, Thomas E. Starzl Transplantation Institute, University of Pittsburgh, Pittsburgh, Pennsylvania
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- Noriko Murase
- Department of Surgery, Thomas E. Starzl Transplantation Institute, University of Pittsburgh, Pittsburgh, Pennsylvania
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- Thomas E. Starzl
- Department of Surgery, Thomas E. Starzl Transplantation Institute, University of Pittsburgh, Pittsburgh, Pennsylvania
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42
Chen S, Liu D, Finley RL, Greenberg ML. Loss of mitochondrial DNA in the yeast cardiolipin synthase crd1 mutant leads to up-regulation of the protein kinase Swe1p that regulates the G2/M transition.
J Biol Chem 2010;
285:10397-407. [PMID:
20086012 DOI:
10.1074/jbc.m110.100784]
[Citation(s) in RCA: 36] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/28/2023] Open
Abstract
The anionic phospholipid cardiolipin and its precursor phosphatidylglycerol are synthesized and localized in the mitochondrial inner membrane of eukaryotes. They are required for structural integrity and optimal activities of a large number of mitochondrial proteins and complexes. Previous studies showed that loss of anionic phospholipids leads to cell inviability in the absence of mitochondrial DNA. However, the mechanism linking loss of anionic phospholipids to petite lethality was unclear. To elucidate the mechanism, we constructed a crd1Deltarho degrees mutant, which is viable and mimics phenotypes of pgs1Delta in the petite background. We found that loss of cardiolipin in rho degrees cells leads to elevated expression of Swe1p, a morphogenesis checkpoint protein. Moreover, the retrograde pathway is activated in crd1Deltarho degrees cells, most likely due to the exacerbation of mitochondrial dysfunction. Interestingly, the expression of SWE1 is dependent on retrograde regulation as elevated expression of SWE1 is suppressed by deletion of RTG2 or RTG3. Taken together, these findings indicate that activation of the retrograde pathway leads to up-regulation of SWE1 in crd1Deltarho degrees cells. These results suggest that anionic phospholipids are required for processes that are essential for normal cell division in rho degrees cells.
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Affiliation(s)
- Shuliang Chen
- Department of Biological Sciences, Wayne State University, Detroit, Michigan 48202, USA
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43
Ke F, Zhao Z, Zhang Q. Cloning, expression and subcellular distribution of a Rana grylio virus late gene encoding ERV1 homologue.
Mol Biol Rep 2009;
36:1651-9. [PMID:
18819018 DOI:
10.1007/s11033-008-9365-6]
[Citation(s) in RCA: 12] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2008] [Accepted: 09/11/2008] [Indexed: 10/21/2022]
Abstract
An essential for respiration and viability (ERV1) homologue, 88R, was cloned and characterized from Rana grylio virus (RGV). Database searches found its homologues in all sequenced iridoviruses, and sequence alignment revealed a highly conserved motif shared by all ERV1 family proteins: Cys-X-X-Cys. RT-PCR and western blot analysis revealed that 88R begins to transcribe and translate at 6 h postinfection (p.i.) and remains detectable at 48 h p.i. during RGV infection course. Furthermore, using drug inhibition analysis by a de novo protein synthesis inhibitor and a viral DNA replication inhibitor, RGV 88R was classified as a late (L) viral gene during the in vitro infection. 88R-EGFP fusion protein was observed in both the cytoplasm and nucleus of pEGFP-N3-88R transfected EPC cells. Although result of immunofluorescence is similar, 88R protein was not detected in viromatrix. Moreover, function of RGV 88R on virus replication were evaluated by RNAi assay. Nevertheless, effect of knockdown of RGV 88R expression on virus replication was not detected in cultured fish cell lines. Collectively, current data indicate that RGV 88R was a late gene of iridovirus encoding protein that distributed both the cytoplasm and nucleus.
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Affiliation(s)
- Fei Ke
- State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Graduate School of the Chinese Academy of Sciences, Wuhan, China
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44
Deponte M, Hell K. Disulphide Bond Formation in the Intermembrane Space of Mitochondria.
J Biochem 2009;
146:599-608. [DOI:
10.1093/jb/mvp133]
[Citation(s) in RCA: 46] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
45
Daithankar VN, Farrell SR, Thorpe C. Augmenter of liver regeneration: substrate specificity of a flavin-dependent oxidoreductase from the mitochondrial intermembrane space.
Biochemistry 2009;
48:4828-37. [PMID:
19397338 PMCID:
PMC2730831 DOI:
10.1021/bi900347v]
[Citation(s) in RCA: 59] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023]
Abstract
Augmenter of liver regeneration (ALR) is both a growth factor and a sulfhydryl oxidase that binds FAD in an unusual helix-rich domain containing a redox-active CxxC disulfide proximal to the flavin ring. In addition to the cytokine form of ALR (sfALR) that circulates in serum, a longer form, lfALR, is believed to participate in oxidative trapping of reduced proteins entering the mitochondrial intermembrane space (IMS). This longer form has an 80-residue N-terminal extension containing an additional, distal, CxxC motif. This work presents the first enzymological characterization of human lfALR. The N-terminal region conveys no catalytic advantage toward the oxidation of the model substrate dithiothreitol (DTT). In addition, a C71A or C74A mutation of the distal disulfide does not increase the turnover number toward DTT. Unlike Erv1p, the yeast homologue of lfALR, static spectrophotometric experiments with the human oxidase provide no evidence of communication between distal and proximal disulfides. An N-terminal His-tagged version of human Mia40, a resident oxidoreductase of the IMS and a putative physiological reductant of lfALR, was subcloned and expressed in Escherichia coli BL21 DE3 cells. Mia40, as isolated, shows a visible spectrum characteristic of an Fe-S center and contains 0.56 +/- 0.02 atom of iron per subunit. Treatment of Mia40 with guanidine hydrochloride and triscarboxyethylphosphine hydrochloride during purification removed this chromophore. The resulting protein, with a reduced CxC motif, was a good substrate of lfALR. However, neither sfALR nor lfALR mutants lacking the distal disulfide could oxidize reduced Mia40 efficiently. Thus, catalysis involves a flow of reducing equivalents from the reduced CxC motif of Mia40 to distal and then proximal CxxC motifs of lfALR to the flavin ring and, finally, to cytochrome c or molecular oxygen.
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Affiliation(s)
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- Scott R. Farrell
- Department of Chemistry and Biochemistry, University of Delaware, Newark, Delaware 19716
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- Colin Thorpe
- Department of Chemistry and Biochemistry, University of Delaware, Newark, Delaware 19716
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46
Bihlmaier K, Mesecke N, Kloeppel C, Herrmann JM. The disulfide relay of the intermembrane space of mitochondria: an oxygen-sensing system?
Ann N Y Acad Sci 2009;
1147:293-302. [PMID:
19076451 DOI:
10.1196/annals.1427.005]
[Citation(s) in RCA: 20] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
The intermembrane space of mitochondria contains many proteins that lack classical mitochondrial targeting sequences. Instead, these proteins often show characteristic patterns of cysteine residues that are critical for their accumulation in the organelle. Import of these proteins is catalyzed by two essential components, Mia40 and Erv1. Mia40 is a protein in the intermembrane space that directly binds newly imported proteins via disulfide bonds. By reorganization of these bonds, intramolecular disulfide bonds are formed in the imported proteins, which are thereby released from Mia40 into the intermembrane space. Because folded proteins are unable to traverse the import pore of the outer membrane, this leads to a permanent location of these proteins within the mitochondria. During this reaction, Mia40 becomes reduced and needs to be re-oxidized to regain its activity. Oxidation of Mia40 is carried out by Erv1, a conserved flavine adenine dinucleotide (FAD)-binding sulfhydryl oxidase. Erv1 directly interacts with Mia40 and shuttles electrons from reduced Mia40 to oxidized cytochrome c, from whence they flow through cytochrome oxidase to molecular oxygen. The connection of the disulfide relay with the respiratory chain not only significantly increases the efficiency of the oxidase activity, but also prevents the formation of potentially deleterious hydrogen peroxide. The oxidative activity of Erv1 strongly depends on the oxygen concentration in mitochondria. Erv1, therefore, may function as a molecular switch that adapts mitochondrial activities to the oxygen levels in the cell.
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Affiliation(s)
- Karl Bihlmaier
- Cell Biology, University of Kaiserslautern, Kaiserslautern, Germany
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47
Terziyska N, Grumbt B, Kozany C, Hell K. Structural and Functional Roles of the Conserved Cysteine Residues of the Redox-regulated Import Receptor Mia40 in the Intermembrane Space of Mitochondria.
J Biol Chem 2009;
284:1353-63. [DOI:
10.1074/jbc.m805035200]
[Citation(s) in RCA: 65] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
48
Thiol oxidation in bacteria, mitochondria and chloroplasts: Common principles but three unrelated machineries?
BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 2009;
1793:71-7. [DOI:
10.1016/j.bbamcr.2008.05.001]
[Citation(s) in RCA: 45] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/12/2008] [Revised: 04/25/2008] [Accepted: 05/05/2008] [Indexed: 11/18/2022]
49
Hell K, Neupert W. Oxidative Protein Folding in Mitochondria.
OXIDATIVE FOLDING OF PEPTIDES AND PROTEINS 2008. [DOI:
10.1039/9781847559265-00067]
[Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/30/2023]
Affiliation(s)
- Kai Hell
- Adolf-Butenandt-Institut für Physiologische Chemie, Ludwig-Maximilians-Universität München Butenandtstrasse 5 81377 München Germany
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- Walter Neupert
- Adolf-Butenandt-Institut für Physiologische Chemie, Ludwig-Maximilians-Universität München Butenandtstrasse 5 81377 München Germany
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50
Down-regulation of hepatic nuclear factor 4alpha on expression of human hepatic stimulator substance via its action on the proximal promoter in HepG2 cells.
Biochem J 2008;
415:111-21. [PMID:
18513187 DOI:
10.1042/bj20080221]
[Citation(s) in RCA: 13] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022]
Abstract
hHSS (human hepatic stimulator substance) stimulates hepatocyte growth. To understand the mechanism controlling hHSS expression, we analysed the proximal promoter activity and identified two regulatory regions (-212/-192 and -152/-132) that were important for transcription in HepG2 cells. Using the luciferase reporter assay, gel-shift experiments and ChIP (chromatin immunoprecipitation), we found that the transcription factors HNF4alpha (hepatocyte nuclear factor 4alpha) and Sp1 (stimulating protein-1) were essential for hHSS promoter activity and could directly bind to regions -209/-204 and -152/-145 respectively. We also confirmed that activation and repression of hHSS transcription induced by Sp1 and HNF4alpha resulted from binding of these factors to these two cis-elements respectively. Overexpression of HNF4alpha led to a dramatic repression of the promoter activity and, in contrast, the activity was markedly elevated by overexpression of Sp1. Furthermore, overexpression of HNF4alpha1, one of the HNF4alpha isoforms, resulted in a dramatic suppression of the promoter activity. Moreover, repression of HNF4alpha expression by siRNA (small interfering RNA) remarkably enhanced the hHSS mRNA level. It has been reported previously that expression of HNF4alpha is functionally regulated by dexamethasone. To further confirm the transcriptional control of HNF4alpha on hHSS, we tested the effect of dexamethasone on hHSS transcription in HepG2 cells. In the present study we have demonstrated that the expression of the hHSS gene was down-regulated at the transcriptional level by dexamethasone in HepG2 cells. A deletion and decoy assay revealed that binding of HNF4alpha to nucleotides -209/-204 was responsible for the suppression of hHSS promoter activity by dexamethasone. Increases in the HNF4alpha-binding activity and expression were simultaneously observed in an electrophoretic mobility-shift assay and Western blot analysis. These results suggested that Sp1 activates hHSS basal expression, but HNF4alpha inhibits hHSS gene expression.
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