Differential susceptibilities of serine/threonine phosphatases to oxidative and nitrosative stress.
Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are signal-transducing molecules that regulate the activities of a variety of proteins. In the present investigation, we have compared the effects of superoxide (O2-), nitric oxide (NO), and hydrogen peroxide (H2O2) on the activities of three highly homologous serine/threonine phosphatases, protein phosphatase type 1 (PP1), protein phosphatase type 2A (PP2A), and calcineurin (protein phosphatase type 2B). Although superoxide, generated from xanthine/xanthine oxidase or paraquat, and NO, generated from (+/-)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide or sodium nitroprusside, potently inhibited the phosphatase activity of calcineurin in neuroblastoma cell lysates, they had relatively little effect on the activities of PP1 or PP2A. In contrast, H2O2 inhibited the activities of all three phosphatases in lysates but was not a potent inhibitor for any of the enzymes. Calcineurin inactivated by O2-, NO, and H2O2 could be partially reactivated by the reducing agent ascorbate or by the thiol-specific reagent dithiothreitol (DTT). Maximal reactivation was achieved by the addition of both reagents, which suggests that ROS and RNS inhibit calcineurin by oxidizing both a catalytic metal(s) and a critical thiol(s). Reactivation of H2O2-treated PP1 also required the combination of both ascorbate and DTT, whereas PP2A required only DTT for reactivation. These results suggest that, despite their highly homologous structures, calcineurin is the only major Ser/Thr phosphatase that is a sensitive target for inhibition by superoxide and nitric oxide and that none of the phosphatases are sensitive to inhibition by hydrogen peroxide.
Sommer
Debbie
D
Department of Pediatrics, College of Medicine, University of Nebraska Medical Center, Omaha, NE 68198-6255, USA.
Coleman
Stacy
S
Swanson
Stanley A
SA
Stemmer
Paul M
PM
D000818:Animals; D019703:Calcineurin / Q000187:drug effects / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D020536:Enzyme Activators / Q000494:pharmacology; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D009447:Neuroblastoma / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism* / Q000494:pharmacology; D020030:Nitric Oxide Donors / Q000494:pharmacology; D016877:Oxidants / Q000378:metabolism / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D010269:Paraquat / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000187:drug effects / Q000378:metabolism*; D011817:Rabbits; D019163:Reducing Agents / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism* / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D014969:Xanthine Oxidase / Q000378:metabolism / Q000494:pharmacology
D020536:Enzyme Activators; D020030:Nitric Oxide Donors; D016877:Oxidants; D019163:Reducing Agents; D013481:Superoxides; D009569:Nitric Oxide; D006861:Hydrogen Peroxide; D014969:Xanthine Oxidase; D019703:Calcineurin; D010749:Phosphoprotein Phosphatases; D010269:Paraquat