Autoantibodies against the multicatalytic proteinase in patients with systemic lupus erythematosus.

Arribas, Joaquı́n; Rodríguez, M.; Forno, R Alvarez-Do; Castaño, José G.

doi:10.1084/jem.173.2.423

Cited by 34 publications

(20 citation statements)

References 25 publications

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“…After electrophoresis the gels were either stained with Coomassive blue or transferred to nitrocellulose as de scribed [7], Nitrocellulose filters were reversibly stained with 0.1 % Ponceau red in 1 % acetic acid and blocked overnight at 4°C with blocking buffer: 50 mmol/l Tris-Cl (pH 7.5), 0.5 mol/1 NaCl and 3% (w/ v) bovine serum albumin (BSA) for immunoblot analy sis. The filters for immunoblot were incubated with anti-C2 antibody affinity-purified against: the fulllength recombinant C2 polypeptide; the C2 protein construct mC2.26 selecting antibodies directed against the C2 COOH (DEPAEKADEPMEH), or the con struct mC2.24 for antibodies directed to a different epitope of the C2 subunit located in the N-terminal in respect to the C2 COOH as described [8], Immunoblot developing was done either by the chemiluminescence method (ECL, Amersham) or with 4-chloro-l-naphthol.…”

Section: Protein Analysis By Sds-page and Immunoblottingmentioning

confidence: 99%

“…After the cells had grown to 50-70% confluency, the coverslips were fixed and permeabilized with absolute methanol at -20 ° C for 15 min, rinsed 3 times with TBS (50 mmol Tris-Cl, pH 7.5; 0.15 mol/1 NaCl) and used for indirect immunofluorescence. Anti-MCP antibody [7] was added at a 1/400 dilution in TBS con taining 3% BSA and incubated with the cells for 1 h at room temperature. After three washes (10 min each) with TBS, the second antibody, a fluorescein-conju gated goat anti-rabbit antibody (Boehringer), was add ed at a 1/300 dilution in TBS containing 3% BSA, together with 2 gmol/1 of 4,6-diamidino-2-phenylindole) for DNA staining.…”

Section: Cell Immunofluorescencementioning

confidence: 99%

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Modulation of the Multicatalytic Proteinase Complex by Lipids, Interconversion and Proteolytic Processing

Arizti

Arribas²,

Castaño³

1993

Enzyme Protein

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In studying the modulation of multicatalytic proteinase (MCP), we have focused on three main aspects; (1) modulation of the activity of the MCP complex by lipids, showing that cardiolipin, sulfatides and gangliosides are potent activators of the enzymatic activity of the complex; (2) modulation by interconversion of MCP subunits, showing that casein kinase II is able to phosphorylate the C8 (this subunit is also be main in vivo phosphorylated subunit) and C9 subunits of the complex in vitro and that a 26-kD subunit is phosphorylated in vitro by protein kinase C, and (3) modulation by proteolytic processing, extending our previous observation of proteolytic processing of the C2 COOH terminus, the presence of enzymatic activities in different subcellular fractions able to convert the intact C2 (32 kD) subunit to a 28-kD polypeptide by removal of at least the last 9-13 amino acids of the C2 polypeptide. The data presented illustrate that the MCP complex is probably under tight and multifactorial control in vivo.

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Section: Protein Analysis By Sds-page and Immunoblottingmentioning

confidence: 99%

Section: Cell Immunofluorescencementioning

confidence: 99%

Modulation of the Multicatalytic Proteinase Complex by Lipids, Interconversion and Proteolytic Processing

Arizti

Arribas²,

Castaño³

1993

Enzyme Protein

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“…We have also detected a gene coding for the proteasome a7/C8 subunit protein on the protein filters and have confirmed its specificity on protein microarrays and by Western immunoblotting analysis. In humans, proteins of the 20S proteasome, such as the subunit C9a, have been frequently recognized in different autoimmune diseases, including SLE [25,26], suggesting these proteins may have potential as marker proteins for general autoimmune inflammation and cell damage [27]. In this context, we do not know if the proteasome a7/C8 subunit is specific for SLE or is detected in a more general way, in other mouse models of autoimmune diseases.…”

Section: Discussionmentioning

confidence: 91%

Mouse protein arrays from a TH1 cell cDNA library for antibody screening and serum profiling

et al. 2005

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The mouse is the premier genetic model organism for the study of disease and development. We describe the establishment of a mouse T helper cell type 1 (T H 1) protein expression library that provides direct access to thousands of recombinant mouse proteins, in particular those associated with immune responses. The advantage of a system based on the combination of large cDNA expression libraries with microarray technology is the direct connection of the DNA sequence information from a particular clone to its recombinant, expressed protein. We have generated a mouse T H 1 expression cDNA library and used protein arrays of this library to characterize the specificity and cross-reactivity of antibodies. Additionally, we have profiled the autoantibody repertoire in serum of a mouse model for systemic lupus erythematosus on these protein arrays and validated the putative autoantigens on highly sensitive protein microarrays. D 2004 Elsevier Inc. All rights reserved.Keywords: T H 1; cDNA expression library; Automation; Mouse; High-density protein arrays; Microarrays; Protein chips; Serum profiling; SLE; Autoantibody The mouse is the premier genetic model organism for the study of disease and development [1]. The final sequencing of the entire mouse genome [2] will greatly increase the advantages associated with mouse models. One of the next steps in expanding the usefulness of mouse models will be to make available tens of thousands of mouse proteins for large-scale high-throughput analyses, e.g., the study of antibody-protein, protein-protein, and peptide-protein interactions and enzyme activities and the characterization of antibody specificity. Such high-throughput studies are greatly simplified by protein microarray technology, whereby thousands of biomolecules are immobilized at high density onto chemically modified surfaces. The advantages of a system based on the combination of large cDNA expression libraries with microarray technology are the direct accessibility of the expressed recombinant proteins and of the DNA sequence information from a particular clone and the ability to re-array the recombinant proteins from selected clones to generate a new microarray for the creation of high-density protein arrays on glass chips [3]. For the generation of protein biochips high-throughput subcloning of open reading frames from the genome of humans, Saccharomyces cerevisiae, Arabidopsis thaliana, and Caenorhabditis elegans have been described [34][35][36][37][38][39]. Such recombination-based cloning approaches are strongly dependent on the progress in genome sequencing projects and the annotation of those sequences [40,37]. This means that previously uncharacterized proteins will be absent,

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“…Originally, 35% of studied SLE patients were shown to have antiproteasome antibodies [210]. A more recent report has shown that 58% of patients with SLE and 62% of patients with myositis have antiproteasome antibodies which were not detected in healthy control individuals or in patients with mixed connective tissue disease and rheumatoid arthritis.…”

Section: Various Immunological Disordersmentioning

confidence: 98%

Ubiquitin- and proteasome-dependent pathway of protein degradation as an emerging therapeutic target

Wójcik¹

2000

Emerging Therapeutic Targets

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Autoantibodies against the multicatalytic proteinase in patients with systemic lupus erythematosus.

Cited by 34 publications

References 25 publications

Modulation of the Multicatalytic Proteinase Complex by Lipids, Interconversion and Proteolytic Processing

Modulation of the Multicatalytic Proteinase Complex by Lipids, Interconversion and Proteolytic Processing

Mouse protein arrays from a TH1 cell cDNA library for antibody screening and serum profiling

Ubiquitin- and proteasome-dependent pathway of protein degradation as an emerging therapeutic target

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