Stimulation of the alveolar macrophage respiratory burst by ADP causes selective glutathionylation of protein tyrosine phosphatase 1B.
H(2)O(2) produced by stimulation of the macrophage NADPH oxidase is involved both in bacterial killing and as a second messenger in these cells. Protein tyrosine phosphatases (PTPs) are targets for H(2)O(2) signaling through oxidation of their catalytic cysteine, resulting in inhibition of their activity. Here, we show that, in the rat alveolar macrophage NR8383 cell line, H(2)O(2) produced through the ADP-stimulated respiratory burst induces the formation of a disulfide bond between PTP1B and GSH that was detectable with an antibody to glutathione-protein complexes and was reversed by DTT addition. PTP1B glutathionylation was dependent on H(2)O(2) as the presence of catalase at the time of ADP stimulation inhibited the formation of the conjugate. Interestingly, other PTPs, i.e., SHP-1 and SHP-2, did not undergo glutathionylation in response to ADP stimulation of the respiratory burst, although glutathionylation of these proteins could be shown by reaction with 25 mM glutathione disulfide in vitro. While previous studies have suggested the reversible oxidation of PTP1B during signaling or showed PTP1B glutathionylation in vitro, the present study directly demonstrates that physiological stimulation of H(2)O(2) production results in PTP1B glutathionylation in intact cells, which may affect downstream signaling.
Rinna
Alessandra
A
School of Natural Sciences, University of California Merced, P.O. Box 2039, Merced, CA 95344, USA.
Torres
Martine
M
Forman
Henry Jay
HJ
D000244:Adenosine Diphosphate / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D002330:Carmustine / Q000494:pharmacology; D002374:Catalase / Q000378:metabolism / Q000494:pharmacology; D002460:Cell Line; D005978:Glutathione / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D016676:Macrophages, Alveolar / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D051381:Rats; D016897:Respiratory Burst / Q000187:drug effects*
D047908:Intracellular Signaling Peptides and Proteins; D000244:Adenosine Diphosphate; D006861:Hydrogen Peroxide; D002374:Catalase; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; C516610:Ptpn1 protein, rat; C516727:Ptpn11 protein, rat; C509250:Ptpn6 protein, rat; D005978:Glutathione; D002330:Carmustine