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Your search keyword '"Gianazza, Elisabetta"' showing total 5 results
Start Over Author "Gianazza, Elisabetta" Journal proceedings of the national academy of sciences of the united states of america
5 results on '"Gianazza, Elisabetta"'

4. Redox regulation of surface protein thiols: identification of integrin alpha-4 as a molecular target by using redox proteomics

Author

Filippo Casoni, Giancarlo Bianchi, Pietro Ghezzi, Tania Massignan, Teresa Laragione, Valentina Bonetto, Elisabetta Gianazza, Laragione, Teresa, Bonetto, Valentina, Casoni, Filippo, Massignan, Tania, Bianchi, Giancarlo, Gianazza, Elisabetta, and Ghezzi, Pietro

Subjects
Electrophoresis, Sulfhydryl Compound, Myosin light-chain kinase, Myosin Light Chains, Time Factors, Free Radicals, Proteome, Integrin alpha4, Integrin, Jurkat Cell, Mass Spectrometry, Cell Line, Cell membrane, chemistry.chemical_compound, Jurkat Cells, Cytosol, Electrophoresi, medicine, Cell Adhesion, Humans, Electrophoresis, Gel, Two-Dimensional, Sulfhydryl Compounds, Cell adhesion, Actin, Multidisciplinary, biology, Dose-Response Relationship, Drug, Myosin Heavy Chains, Chemistry, Cell Membrane, Myosin Heavy Chain, Glutathione, Biological Sciences, Actins, Acetylcysteine, Myosin Light Chain, medicine.anatomical_structure, Membrane protein, Biochemistry, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization, biology.protein, Biophysics, Leukocytes, Mononuclear, Free Radical, Oxidation-Reduction, Intracellular, Human
Abstract

Thiols affect a variety of cell functions, an effect known as redox regulation. We show here that treatment (1–2 h) of cells with 0.1–5 mMN-acetyl-l-cysteine (NAC) increases surface protein thiol expression in human peripheral blood mononuclear cells. This effect is not associated with changes in cellular glutathione (GSH) and is also observed with a non-GSH precursor thiolN-acetyl-d-cysteine or with GSH itself, which is not cell-permeable, suggesting a direct reducing action. NAC did not augment protein SH in the cytosol, indicating that they are already maximally reduced under normal, nonstressed, conditions. By using labeling with a non permeable, biotinylated SH reagent followed by two-dimensional gel electrophoresis and analysis by MS, we identified some of the proteins associated with the membrane that are reduced by NAC. These proteins include the following: integrin α-4, myosin heavy chain (nonmuscle type A), myosin light-chain alkali (nonmuscle isoform), and β-actin. NAC pretreatment augmented integrin α-4-dependent fibronectin adhesion and aggregation of Jurkat cells without changing its expression by fluorescence-activated cell sorter, suggesting that reduction of surface disulfides can affect proteins function. We postulate that some of the activities of NAC or other thiol antioxidants may not only be due to free radical scavenging or increase of intracellular GSH and subsequent effects on transcription factors, but could modify the redox state of functional membrane proteins with exofacial SH critical for their activity.

Published
2003

5. Redox regulation of surface protein thiols: identification of integrin alpha-4 as a molecular target by using redox proteomics.

Author

Laragione T, Bonetto V, Casoni F, Massignan T, Bianchi G, Gianazza E, and Ghezzi P

Subjects
Acetylcysteine chemistry, Actins metabolism, Cell Adhesion, Cell Line, Cell Membrane metabolism, Cytosol metabolism, Dose-Response Relationship, Drug, Electrophoresis, Electrophoresis, Gel, Two-Dimensional, Free Radicals, Glutathione metabolism, Humans, Integrin alpha4 metabolism, Jurkat Cells, Leukocytes, Mononuclear metabolism, Mass Spectrometry, Myosin Heavy Chains chemistry, Myosin Light Chains chemistry, Oxidation-Reduction, Proteome, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization, Sulfhydryl Compounds, Time Factors, Integrin alpha4 physiology
Abstract

Thiols affect a variety of cell functions, an effect known as redox regulation. We show here that treatment (1-2 h) of cells with 0.1-5 mM N-acetyl-L-cysteine (NAC) increases surface protein thiol expression in human peripheral blood mononuclear cells. This effect is not associated with changes in cellular glutathione (GSH) and is also observed with a non-GSH precursor thiol N-acetyl-D-cysteine or with GSH itself, which is not cell-permeable, suggesting a direct reducing action. NAC did not augment protein SH in the cytosol, indicating that they are already maximally reduced under normal, nonstressed, conditions. By using labeling with a non permeable, biotinylated SH reagent followed by two-dimensional gel electrophoresis and analysis by MS, we identified some of the proteins associated with the membrane that are reduced by NAC. These proteins include the following: integrin alpha-4, myosin heavy chain (nonmuscle type A), myosin light-chain alkali (nonmuscle isoform), and beta-actin. NAC pretreatment augmented integrin alpha-4-dependent fibronectin adhesion and aggregation of Jurkat cells without changing its expression by fluorescence-activated cell sorter, suggesting that reduction of surface disulfides can affect proteins function. We postulate that some of the activities of NAC or other thiol antioxidants may not only be due to free radical scavenging or increase of intracellular GSH and subsequent effects on transcription factors, but could modify the redox state of functional membrane proteins with exofacial SH critical for their activity.

Published
2003
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