Protein glutathionylation: coupling and uncoupling of glutathione to protein thiol groups in lymphocytes under oxidative stress and HIV infection.
We show here that exposure to oxidative stress induces glutathione (GSH) modification of protein cysteinyl residues (glutathionylation) in T cell blasts. Treating the cells with the oxidant diamide induces thiolation of a series of proteins that can be detected by 2D electrophoresis when 35S-cysteine is used to label the intracellular GSH pool. This thiolation is reversible, proteins are rapidly dethiolated and GSH is released from proteins once the oxidants are washed and the cells are allowed to recover. Dethiolation is dependent on the availability of GSH and thiols, since it is inhibited by GSH-depleting agents and improved by N-acetyl-L-cysteine (NAC). The capacity of these agents to reverse glutathionylation is diminished in T cell blasts infected in vitro with HIV, which is known to cause oxidative stress. Consistent with these findings, the activity of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), an enzyme known to be inhibited by glutathionylation, is inhibited in diamide-treated cells and recovers rapidly when cells are allowed to dethiolate. Further, GAPDH activity is diminished by GSH-depleting agents and augmented by NAC. Thus, reversible glutathionylation of proteins can rapidly shift the activity of a key metabolic enzyme and thereby result in dramatic, reversible changes in cellular metabolism.
Ghezzi
Pietro
P
Department of Genetics, Stanford University School of Medicine, Stanford, CA, USA. ghezzi@marionegri.it
Romines
Brie
B
Fratelli
Maddalena
M
Eberini
Ivano
I
Gianazza
Elisabetta
E
Casagrande
Simona
S
Laragione
Teresa
T
Mengozzi
Manuela
M
Herzenberg
Leonore A
LA
Herzenberg
Leonard A
LA
D000111:Acetylcysteine; D003958:Diamide / Q000737:chemistry / Q000378:metabolism; D005978:Glutathione / Q000737:chemistry / Q000378:metabolism*; D015658:HIV Infections / Q000097:blood*; D006801:Humans; D018384:Oxidative Stress; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D013438:Sulfhydryl Compounds; D013601:T-Lymphocytes / Q000737:chemistry / Q000378:metabolism*
D011506:Proteins; D013438:Sulfhydryl Compounds; D003958:Diamide; D005978:Glutathione; D000111:Acetylcysteine