Kinetics of Tyrosine Phosphorylation When IgE Dimers Bind to FC∊ Receptors on Rat Basophilic Leukemia Cells

Wofsy, Carla; Kent, Ute M.; Mao, Su-Yau; Metzger, Henry; Goldstein, Byron

doi:10.1074/jbc.270.35.20264

Cited by 30 publications

(24 citation statements)

References 54 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We have previously proposed that in the cell line we are studying, the failure of phosphorylation to correlate with aggregation under some experimental conditions could be explained if the kinase responsible for k 12 is limiting (43). Recent experiments have provided experimental support for this proposal (44).…”

Section: Kinetics Of Phosphorylation and Dephosphorylation Of Fc⑀ri Amentioning

confidence: 87%

See 1 more Smart Citation

Characterization of Protein-tyrosine Phosphatases That Dephosphorylate the High Affinity IgE Receptor

Mao

Metzger

1997

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

An early event that follows aggregation of the high affinity receptor for IgE (Fc⑀RI) is the phosphorylation of protein tyrosines, especially those on the ␤-and ␥-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 Fc⑀RI. Upon disaggregation, the in vivo dephosphorylation of the Fc⑀RI 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 Fc⑀RI 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 Fc⑀RI was comparable in intact and permeabilized cells. Thus, much of the activity utilized by the cell to dephosphorylate the Fc⑀RI is likely to be associated with the plasma membrane.The high affinity IgE receptor on mast cells and basophils (Fc⑀RI) has a central role in mediating allergic responses (1, 2). Aggregation of the receptors results in phosphorylation of protein tyrosines as an early event that leads to a variety of later cellular phenomena (3, 4). The receptor itself lacks sequences typical for intrinsic protein-tyrosine kinase activity (5), and experimentally, receptors purified by affinity columns or well washed immunoprecipitates show virtually no kinase activity in vitro (6, 7). However, a variety of studies have implicated a kinase weakly associated with the receptor (6,8,9). In rats, the critical initial kinase appears to be Lyn (9). The receptorassociated Lyn from resting cells displays tyrosine kinase activity toward exogenous substrates, but little or no phosphorylation of the receptor itself is observed unless the receptors are aggregated (10 -13). These observations and those made on the effect of inhibitors of phosphatases (12,14) imply that proteintyrosine phosphatases (PTPs) 1 are continuously modulating the resting system. The studies with inhibitors and other experimental approaches (15) show that the PTPs also continuously act on aggregated receptors. Finally, when individual receptors dissociate from the aggregate, e.g. by addition of monomeric hapten after stimulation by multivalent antigen, rapid dephosphorylation of the receptor and of other cellular proteins is observed (7, 10 -13).The PTPs involved in these processes remain undefined, and the purpose of the current study was to investigate some of their characteristics. We first developed an in vitro assay to test the PTP activity in total cell lysate toward receptors that had been phosp...

show abstract

Section: Kinetics Of Phosphorylation and Dephosphorylation Of Fc⑀ri Amentioning

confidence: 87%

“…A simple scheme was proposed as a model on which to base future experiments (Fig. 8) (43). Earlier data showed that k Ϫ22 is substantial and effectively overwhelms k 22 .…”

Section: Kinetics Of Phosphorylation and Dephosphorylation Of Fc⑀ri Amentioning

confidence: 99%

Characterization of Protein-tyrosine Phosphatases That Dephosphorylate the High Affinity IgE Receptor

Mao

Metzger

1997

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

show abstract

“…As in peripheral blood basophils [unpublished results] and RBL-2H3 mast cells [49,50], overall anti-phosphotyrosine reactivity increases with anti-IgE concentrations up to a maximum, and remains elevated at very high anti-IgE concentrations that no longer support degranulation. The dissociation of the extent of secretion and tyrosine phosphorylation at high antigen concentrations is unexplained, but may reflect differences in phosphoprotein species or in phosphorylation sites within individual proteins under different stimulation conditions.…”

Section: Discussionmentioning

confidence: 99%

The identification and characterization of umbilical cord blood-derived human basophils

Kepley¹,

Pfeiffer²,

Schwartz

et al. 1998

Journal of Leukocyte Biology

View full text Add to dashboard Cite

Cross-linking allergen-specific immunoglobin E on human peripheral blood basophils results in the release of histamine and other inflammatory mediators that initiate allergy and asthma. The signaling pathways leading from IgE binding to mediator release have not been well established, mainly due to the difficulty in obtaining adequate numbers of highly purified basophils. It was the goal of this study to easily obtain Fc⑀RI-positive human basophils in high yield and purity for studies of signal transduction pathways. We describe an in vitro culture system in which pulsing normal human cord blood leukocytes with interleukin-3 (IL-3) for 3-4 h followed by incubation in medium with fetal bovine serum generates a cell population that is predominately Fc⑀RI positive between 14 and 28 days of culture. These cells resemble peripheral blood basophils when examined by light and electron microscopy. Like normal blood basophils, they express the integrins, CD11b, CD18, CD29, and CD49d. A majority of the IL-3-pulsed cells also express a marker recognized by the basophilspecific antibody, 2D7. Fc⑀RI cross-linking results in a time and dose-dependent release of histamine. Fc⑀RI cross-linking also stimulates protein-tyrosine phosphorylation, thought to be the first event leading to the IgE-mediated activation of peripheral blood basophils. These studies establish cord blood as an accessible source from which basophillike cells can be developed to examine Fc⑀RI-mediated signal transduction. J. Leukoc. Biol. 64: 474-483; 1998.

show abstract

“…When this receptor is triggered degranulation of the mast cell/basophil occurs and mediators important in allergic reactions are released. The kinetics and degree of aggregation required for signaling has been extensively studied by several groups and several models have been developed of this system (59)(60)(61)(62). Results that have emerged from these studies have revealed that simple cyclic dimers of bound FcεRI are not sufficient to induce activation (60), and this suggested that complex structures were required in order to trigger internal signaling events.…”

Section: Signal Transduction Modelsmentioning

confidence: 99%

Mathematical modeling of immunological reactions

Morel

1998

Front Biosci

View full text Add to dashboard Cite

Kinetics of Tyrosine Phosphorylation When IgE Dimers Bind to FC∊ Receptors on Rat Basophilic Leukemia Cells

Cited by 30 publications

References 54 publications

Characterization of Protein-tyrosine Phosphatases That Dephosphorylate the High Affinity IgE Receptor

Characterization of Protein-tyrosine Phosphatases That Dephosphorylate the High Affinity IgE Receptor

The identification and characterization of umbilical cord blood-derived human basophils

Mathematical modeling of immunological reactions

Contact Info

Product

Resources

About