Modulation of human neutrophil responses to CD32 cross-linking by serine/threonine phosphatase inhibitors: cross-talk between serine/threonine and tyrosine phosphorylation.
The interplay between serine/threonine and tyrosine phosphorylation was studied in human neutrophils. The direct effects of calyculin and okadaic acid, potent inhibitors of PP1 and PP2A serine/threonine phosphatases, on the patterns of neutrophil phosphorylation, and their effects on the responses of neutrophils to CD32 cross-linking were monitored. After a 2-min incubation with 10-6 M calyculin, a transient tyrosine phosphorylation of a subset of proteins, among which Cbl and Syk, was observed. After a longer incubation (>5 min) with calyculin, concomitant with an accumulation of serine and threonine phosphorylation, neutrophil responses to CD32 cross-linking were selectively altered. Tyrosine phosphorylation of Cbl in response to CD32 cross-linking was inhibited by calyculin, and this inhibition was linked with a slower electrophoretic mobility of Cbl as a consequence of its phosphorylation on serine/threonine residues. However, tyrosine phosphorylation of Syk and of the receptor itself were not affected. Furthermore, the mobilization of intracellular calcium stimulated by CD32 cross-linking was totally abrogated by calyculin. Finally, the stimulation of superoxide production observed in response to CD32 cross-linking was enhanced in calyculin-treated cells. These results suggest that serine/threonine phosphorylation events regulate the signaling pathways activated by CD32 cross-linking in neutrophils and identify a novel mechanism of modulation of the functional responsiveness of human neutrophils to CD32 cross-linking.
Rollet-Labelle
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E
Centre de Recherche en Rhumatologie et Immunologie, Centre de recherche du Centre Hospitalier Universitaire de Quebec (CHUQ), Quebec, Canada.
Gilbert
C
C
Naccache
P H
PH
D000328:Adult; D002118:Calcium / Q000037:antagonists & inhibitors / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D008387:Marine Toxins; D018375:Neutrophil Activation / Q000187:drug effects; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology* / Q000276:immunology* / Q000378:metabolism; D050722:Oncogene Protein v-cbl; D010080:Oxazoles / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D010766:Phosphorylation / Q000187:drug effects; D017452:Receptors, IgG / Q000276:immunology* / Q000378:metabolism* / Q000502:physiology; D015682:Retroviridae Proteins, Oncogenic / Q000302:isolation & purification / Q000378:metabolism; D012694:Serine / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism; D013912:Threonine / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*
D004791:Enzyme Inhibitors; D008387:Marine Toxins; D050722:Oncogene Protein v-cbl; D010080:Oxazoles; D017452:Receptors, IgG; D015682:Retroviridae Proteins, Oncogenic; D013481:Superoxides; D013912:Threonine; D014443:Tyrosine; D012694:Serine; C059041:calyculin A; D010749:Phosphoprotein Phosphatases; D002118:Calcium