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Published on: December 1, 2014
Regulation of mast cell signaling through high-affinity IgE receptor by CD45 protein tyrosine phosphatase
K Murakami1, S Sato, S Nagasawa
1Department of Hygienic Chemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita-ku, Sapporo 060-0812, Japan.
Abstract:
The transmembrane tyrosine phosphatase CD45 regulates the activity of src family protein tyrosine kinases (PTK) and thereby influences the signaling via such receptors as T and B cell antigen receptors associated with these PTK. However, its implication in signaling through the mast cell receptor with high affinity for IgE (FcepsilonRI) is less clear, although Lyn, a member of the src family, plays an important role in FcepsilonRI-mediated signaling. To define a role for CD45 in FcepsilonRI signal transduction, we established CD45 high expressing rat basophilic leukemia cell lines (RBL-CD45H) and cell lines expressing trace amounts of CD45 (RBL-CD45L). We demonstrate that although all RBL-CD45L cell lines degranulate following IgE- and antigen-induced FcepsilonRI aggregation, the response is significantly reduced at a low dose of antigen. The cells show a delayed and slowed Ca(2+) mobilization even though at a higher dose where the cells degranulate to a similar extent as RBL-CD45H. This diminished Ca(2+) response is restored by reconstitution of RBL-CD45L with a chimeric molecule containing the cytoplasmic phosphatase domains of rat CD45. Furthermore, tyrosine phosphorylation of FcepsilonRI, association of FcepsilonRI with Lyn and PTK activity associated with FcepsilonRI, all of which are enhanced upon FcepsilonRI aggregation in RBL-CD45H, are impaired in RBL-CD45L. Finally, we show that FcepsilonRI is physically associated with CD45 in RBL-CD45H prior to receptor aggregation. Thus, we propose that, although not indispensable in mast cell degranulation, CD45 positively regulates the signaling through FcepsilonRI by promoting the activation of FcepsilonRI-associated Lyn.
Insights
Transmembrane tyrosine phosphatase CD45 positively regulates signaling through the mast cell receptor (FcepsilonRI) by enhancing Lyn kinase activity. While not essential for degranulation, CD45 is crucial for robust calcium mobilization and FcepsilonRI signaling.
Area of Science:
- Immunology
- Cell Signaling
- Molecular Biology
Background:
- CD45, a transmembrane tyrosine phosphatase, regulates src family kinases (PTKs) and influences signaling through T and B cell receptors.
- The role of CD45 in signaling via the high-affinity IgE receptor (FcepsilonRI) on mast cells is less understood, despite Lyn, an src family kinase, being critical for FcepsilonRI signaling.
Purpose of the Study:
- To elucidate the specific role of CD45 in FcepsilonRI signal transduction in mast cells.
- To investigate how CD45 expression levels affect FcepsilonRI-mediated signaling pathways.
Main Methods:
- Generation of rat basophilic leukemia cell lines with high (RBL-CD45H) and low (RBL-CD45L) CD45 expression.
- Analysis of FcepsilonRI-mediated degranulation, calcium mobilization, and Lyn kinase activity in response to antigen stimulation.
- Assessment of FcepsilonRI and Lyn association and tyrosine phosphorylation.
Main Results:
- RBL-CD45L cells showed reduced degranulation and delayed calcium mobilization at low antigen doses compared to RBL-CD45H cells.
- Diminished calcium response in RBL-CD45L cells was restored by reconstituting with CD45 phosphatase domains.
- Tyrosine phosphorylation of FcepsilonRI, Lyn association, and PTK activity were impaired in RBL-CD45L cells.
- FcepsilonRI was found to be physically associated with CD45 in RBL-CD45H cells prior to receptor aggregation.
Conclusions:
- CD45 positively regulates FcepsilonRI signaling by enhancing the activation of associated Lyn kinase.
- CD45 is not indispensable for mast cell degranulation but plays a significant role in modulating FcepsilonRI signal transduction.
- Physical association between CD45 and FcepsilonRI is important for efficient signaling.
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