Gene expression profiling reveals a signaling role of glutathione in redox regulation.

Proteins can form reversible mixed disulfides with glutathione (GSH). It has been hypothesized that protein glutathionylation may represent a mechanism of redox regulation, in a fashion similar to that mediated by protein phosphorylation. We investigated whether GSH has a signaling role in the response of HL60 cells to hydrogen peroxide (H2O2), in addition to its obvious antioxidant role. We identified early changes in gene expression induced at different times by H2O2 treatment, under conditions that increase protein glutathionylation and minimal toxicity. We then investigated the effect of prior GSH depletion by buthionine sulfoximine and diethylmaleate on this response. The analysis revealed 2,016 genes regulated by H2O2. Of these, 215 genes showed GSH-dependent expression changes, classifiable into four clusters displaying down- or up-regulation by H2O2, either potentiated or inhibited by GSH depletion. The modulation of 20 selected genes was validated by real-time RT-PCR. The biological process categories overrepresented in the largest cluster (genes whose up-regulation was inhibited by GSH depletion) were NF-kappaB activation, transcription, and DNA methylation. This cluster also included several cytokine and chemokine ligands and receptors, the redox regulator thioredoxin interacting protein, and the histone deacetylase sirtuin. The cluster of genes whose up-regulation was potentiated by GSH depletion included two HSPs (HSP40 and HSP70) and the AP-1 transcription factor components Fos and FosB. This work demonstrates that GSH, in addition to its antioxidant and protective function against oxidative stress, has a specific signaling role in redox regulation.

[1]  P. Ghezzi.  ReviewRegulation of protein function by glutathionylation , 2005, Free radical research.

[2]  J. Mieyal,et al.  Glutaredoxin: role in reversible protein s-glutathionylation and regulation of redox signal transduction and protein translocation. , 2005, Antioxidants & redox signaling.

[3]  Jean YH Yang,et al.  Bioconductor: open software development for computational biology and bioinformatics , 2004, Genome Biology.

[4]  Myriam Gorospe,et al.  Calorie Restriction Promotes Mammalian Cell Survival by Inducing the SIRT1 Deacetylase , 2004, Science.

[5]  H. Gaskins,et al.  A combined in vitro/bioinformatic investigation of redox regulatory mechanisms governing cell cycle progression. , 2004, Physiological genomics.

[6]  Kazuo T. Suzuki,et al.  Synchronized generation of reactive oxygen species with the cell cycle. , 2004, Life sciences.

[7]  R. Brigelius-Flohé,et al.  Redox events in interleukin-1 signaling. , 2004, Archives of biochemistry and biophysics.

[8]  Bing Zhang,et al.  GOTree Machine (GOTM): a web-based platform for interpreting sets of interesting genes using Gene Ontology hierarchies , 2004, BMC Bioinformatics.

[9]  Ana Rute Neves,et al.  Yeast Life-Span Extension by Calorie Restriction Is Independent of NAD Fluctuation , 2003, Science.

[10]  P. Pavlidis.  Using ANOVA for gene selection from microarray studies of the nervous system. , 2003, Methods.

[11]  J. Vandekerckhove,et al.  Identification of proteins undergoing glutathionylation in oxidatively stressed hepatocytes and hepatoma cells , 2003, Proteomics.

[12]  C. Nathan.  Specificity of a third kind: reactive oxygen and nitrogen intermediates in cell signaling. , 2003, The Journal of clinical investigation.

[13]  A. Garg,et al.  Reactive oxygen intermediates in TNF signaling. , 2002, Molecular immunology.

[14]  Z. Werb,et al.  Integrins engage mitochondrial function for signal transduction by a mechanism dependent on Rho GTPases , 2002, The Journal of cell biology.

[15]  A. Sica,et al.  Glutathione protects mice from lethal sepsis by limiting inflammation and potentiating host defense. , 2002, The Journal of infectious diseases.

[16]  P. Ghezzi,et al.  Identification by redox proteomics of glutathionylated proteins in oxidatively stressed human T lymphocytes , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[17]  T. Galeotti,et al.  Reactive Oxygen Species as Downstream Mediators of Angiogenic Signaling by Vascular Endothelial Growth Factor Receptor-2/KDR* , 2002, The Journal of Biological Chemistry.

[18]  P. Ghezzi,et al.  Protein glutathionylation: coupling and uncoupling of glutathione to protein thiol groups in lymphocytes under oxidative stress and HIV infection. , 2002, Molecular immunology.

[19]  L. Oberley,et al.  Discrete Generation of Superoxide and Hydrogen Peroxide by T Cell Receptor Stimulation , 2002, The Journal of experimental medicine.

[20]  John Quackenbush,et al.  Genesis: cluster analysis of microarray data , 2002, Bioinform..

[21]  Xiangdong Wu,et al.  Hydrogen Peroxide Generated during Cellular Insulin Stimulation Is Integral to Activation of the Distal Insulin Signaling Cascade in 3T3-L1 Adipocytes* , 2001, The Journal of Biological Chemistry.

[22]  R. Weinberg,et al.  hSIR2SIRT1 Functions as an NAD-Dependent p53 Deacetylase , 2001, Cell.

[23]  S. Ghosh,et al.  Toll-like receptor-mediated NF-kappaB activation: a phylogenetically conserved paradigm in innate immunity. , 2001, The Journal of clinical investigation.

[24]  M. Pangalos,et al.  The use of TaqMan RT-PCR assays for semiquantitative analysis of gene expression in CNS tissues and disease models , 2000, Journal of Neuroscience Methods.

[25]  V. Velarde,et al.  Role of reactive oxygen species in bradykinin-induced mitogen-activated protein kinase and c-fos induction in vascular cells. , 2000, Hypertension.

[26]  J. Raymond,et al.  5-HT2A receptors stimulate mitogen-activated protein kinase via H2O2 generation in rat renal mesangial cells , 2000 .

[27]  R. Alexander,et al.  Reactive Oxygen Species Mediate the Activation of Akt/Protein Kinase B by Angiotensin II in Vascular Smooth Muscle Cells* , 1999, The Journal of Biological Chemistry.

[28]  M. Matsui,et al.  Identification of Thioredoxin-binding Protein-2/Vitamin D3 Up-regulated Protein 1 as a Negative Regulator of Thioredoxin Function and Expression* , 1999, The Journal of Biological Chemistry.

[29]  C. Volbracht,et al.  CD95‐mediated murine hepatic apoptosis requires an intact glutathione status , 1999, Hepatology.

[30]  A. Zibert,et al.  Immunodominant B-cell domains of hepatitis C virus envelope proteins E1 and E2 identified during early and late time points of infection. , 1999, Journal of hepatology.

[31]  K. Vasquez,et al.  Glutathione levels in antigen-presenting cells modulate Th1 versus Th2 response patterns. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[32]  E. Tekle,et al.  Epidermal Growth Factor (EGF)-induced Generation of Hydrogen Peroxide , 1997, The Journal of Biological Chemistry.

[33]  V. Ferrans,et al.  Requirement for Generation of H2O2 for Platelet-Derived Growth Factor Signal Transduction , 1995, Science.

[34]  K. Schulze-Osthoff,et al.  Functions of glutathione and glutathione disulfide in immunology and immunopathology , 1994, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[35]  K. Nose,et al.  Production of hydrogen peroxide by transforming growth factor-beta 1 and its involvement in induction of egr-1 in mouse osteoblastic cells , 1994, The Journal of cell biology.

[36]  J. Mcgregor,et al.  Use of N-acetyl cysteine to increase intracellular glutathione during the induction of antitumor responses by IL-2. , 1994, Journal of immunology.

[37]  I. Slukvin,et al.  Increased susceptibility of experimental animals to infectious organisms as a consequence of ethanol consumption. , 1994, Alcohol and alcoholism (Oxford, Oxfordshire). Supplement.

[38]  S. Moncada,et al.  The L-arginine-nitric oxide pathway. , 1993, The New England journal of medicine.

[39]  J. Wang,et al.  Regulation by glutathione of the activation and differentiation of IL-4-dependent activated killer cells. , 1993, Cellular immunology.

[40]  M. Roederer,et al.  Glutathione deficiency and human immunodeficiency virus infection , 1992, The Lancet.

[41]  W. Dröge,et al.  HIV-induced cysteine deficiency and T-cell dysfunction--a rationale for treatment with N-acetylcysteine. , 1992, Immunology today.

[42]  G. Vendemiale,et al.  Hepatic glutathione content in patients with alcoholic and non alcoholic liver diseases. , 1988, Life sciences.

[43]  S. D. Murphy,et al.  Effect of diethylmaleate and other glutathione depletors on protein synthesis. , 1986, Biochemical pharmacology.

[44]  J. Miners,et al.  The effects of buthionine sulphoximine (BSO) on glutathione depletion and xenobiotic biotransformation. , 1984, Biochemical pharmacology.

[45]  H. Gilbert.  Redox control of enzyme activities by thiol/disulfide exchange. , 1984, Methods in enzymology.

[46]  H. Sies,et al.  Identification and quantitation of glutathione in hepatic protein mixed disulfides and its relationship to glutathione disulfide. , 1983, Biochemical pharmacology.

[47]  A. Holmgren,et al.  Relative contributions of thioltransferase-and thioredoxin-dependent systems in reduction of low-molecular-mass and protein disulphides. , 1983, The Biochemical journal.

[48]  H. Sies,et al.  Increase in hepatic mixed disulphide and glutathione disulphide levels elicited by paraquat. , 1982, Biochemical pharmacology.

[49]  F. Tietze.  Enzymic method for quantitative determination of nanogram amounts of total and oxidized glutathione: applications to mammalian blood and other tissues. , 1969, Analytical biochemistry.