title: Protein glutathionylation in cellular compartments: a constitutive redox signal.
abstract: Glutathione provides means of regulating protein function by the process of glutathionylation. Despite the role of oxidative stress biomarkers assumed recently by glutathionylated proteins in human diseases, so far no information is available on the intracellular distribution of glutathionylated proteins in human cell lines. In this study, we combined the specificity of monoclonal antibody labeling for protein-bound glutathione (GS-Pro) with the ability of confocal microscopy to localize molecules with high spatial resolution. We performed immunofluorescence analysis on dermal fibroblasts, both in steady state than in proliferative conditions, and on in situ extracted matrix samples. For the first time, we report the compartmentalization of constitutively glutathionylated proteins in different subcellular districts and we found a tight association between glutathione, nuclear lamina, and cytoskeleton. In proliferating cells, total GS-Pro fluorescence increases in the early phases of growth and significantly drops when cells reach confluence. Interestingly, a nuclear shift of GS-Pro was observed between 6 and 48 hours after plating, becoming homogeneous with the cytoplasm when growth slows. The ability to visualize a detailed intracellular distribution of this critical marker of protein oxidation may provide an additional tool to highlight pathways in turns 'redox-activated' and to identify new pathogenic pathways in human diseases.
authors.affiliation: Laboratories of Research, Bambino Gesù Children’s Hospital, IRCCS, Rome, Italy. stefania.petrini@opbg.net
authors.forename: Stefania
authors.initials: S
authors.lastname: Petrini
authors.forename: Chiara
authors.initials: C
authors.lastname: Passarelli
authors.forename: Anna
authors.initials: A
authors.lastname: Pastore
authors.forename: Giulia
authors.initials: G
authors.lastname: Tozzi
authors.forename: Marianna
authors.initials: M
authors.lastname: Coccetti
authors.forename: Manuela
authors.initials: M
authors.lastname: Colucci
authors.forename: Marzia
authors.initials: M
authors.lastname: Bianchi
authors.forename: Rosalba
authors.initials: R
authors.lastname: Carrozzo
authors.forename: Enrico
authors.initials: E
authors.lastname: Bertini
authors.forename: Fiorella
authors.initials: F
authors.lastname: Piemonte
mesh_terms: D000911:Antibodies, Monoclonal; D002451:Cell Compartmentation / Q000502:physiology*; D049109:Cell Proliferation; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D005978:Glutathione / Q000378:metabolism*; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D018613:Microscopy, Confocal; D034881:Nuclear Lamina / Q000378:metabolism; D010084:Oxidation-Reduction; D017382:Reactive Oxygen Species / Q000494:pharmacology; D012867:Skin / Q000648:ultrastructure
chemical_list: D000911:Antibodies, Monoclonal; D003598:Cytoskeletal Proteins; D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide; D005978:Glutathione