Characterization of Protein-tyrosine Phosphatases That Dephosphorylate the High Affinity IgE Receptor*
- From the Arthritis and Rheumatism Branch, NIAMS, National Institutes of Health, Bethesda, Maryland 20892
Abstract
An early event that follows aggregation of the high affinity receptor for IgE (FcεRI) is the phosphorylation of protein tyrosines, especially those on the β- and γ-subunits of the receptor. Disaggregation of the receptors leads to their rapid dephosphorylation, but even stably aggregated receptors undergo continual rounds of phosphorylation and dephosphorylation. We developed assays to study dephosphorylation of the receptors and other cellular proteins. Whole cell extracts dephosphorylated both subunits of the receptors rapidly and were as active against aggregated as against disaggregated FcεRI. Upon disaggregation, the in vivodephosphorylation of the FcεRI and several other proteins followed first-order kinetics with closely similar rate constants despite substantial differences in the extent of phosphorylation. These results suggest that the level of phosphorylation of FcεRI is largely controlled by the aggregation-induced action of kinase(s) and not from changes in susceptibility to or activity of the phosphatases. Much of the total phosphatase is lost when the cells are permeabilized, but the rate of dephosphorylation of disaggregated FcεRI was comparable in intact and permeabilized cells. Thus, much of the activity utilized by the cell to dephosphorylate the FcεRI is likely to be associated with the plasma membrane.
Footnotes
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↵* The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ To whom correspondence should be addressed: NIAMS, National Institutes of Health, Bldg. 10, Rm. 9N-228, 10 Center Dr., MSC 1820, Bethesda, MD 20892-1820. Tel.: 301-496-2612; Fax: 301-402-0012.
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↵1 The abbreviations used are: PTP, protein-tyrosine phosphatase; DNP, 2,4-dinitrophenyl; DNP25-BSA, dinitrophenylated bovine serum albumin; RBL, rat basophilic leukemia; PAGE, polyacrylamide gel electrophoresis; PIPES, 1,4-piperazinediethanesulfonic acid; ITAM, immune recognition tyrosine activation motif.
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↵2 Rodgers, W., and Rose, J. K. (1996) J. Cell Biol 135, 1515–1523.
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- Received November 8, 1996.
- Revision received February 24, 1997.
