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Quantitative Nano-Scale Analysis of IgE-FcεRI Clustering and Coupling to Early Signaling Proteins
- Publication Year :
- 2012
-
Abstract
- Antigen-mediated crosslinking of IgE bound to its receptor, FcεRI, initiates a transmembrane signaling cascade that results in mast cell activation in the allergic response. Using immunogold labeling of intact RBL mast cells and scanning electron microscopy (SEM), we visualize molecular reorganization of IgE-FcεRI and early signaling proteins on both leaflets of the plasma membrane, without the need for ripped off membrane sheets. As quantified by pair correlation analysis, we observe dramatic changes in the nano-scale distribution of IgE-FcεRI after binding of multivalent antigen to stimulate transmembrane signaling, and this is accompanied by similar clustering of Lyn and Syk tyrosine kinases, and adaptor protein LAT. We find that Lyn co-redistributes with IgE-FcεRI into clusters that cross-correlate throughout 20 min of stimulation. Inhibition of tyrosine kinase activity reduces the numbers of both IgE-FcεRI and Lyn in stimulated clusters. Coupling of these proteins is also decreased when membrane cholesterol is reduced either before or after antigen addition. These results provide evidence for involvement of FcεRI phosphorylation and cholesterol dependent membrane structure in the interactions that accompany IgE-mediated activation of RBL mast cells. More generally, this SEM view of intact cell surfaces provides new insights into the nano-scale organization of receptor-mediated signaling complexes in the plasma membrane.
- Subjects :
- biology
Chemistry
Receptors, IgE
Syk
Signal transducing adaptor protein
Immunogold labelling
Immunoglobulin E
Article
Surfaces, Coatings and Films
Cell biology
Rats
Biochemistry
LYN
Materials Chemistry
biology.protein
Microscopy, Electron, Scanning
Tumor Cells, Cultured
Phosphorylation
Animals
Nanotechnology
Physical and Theoretical Chemistry
Signal transduction
Tyrosine kinase
Signal Transduction
Subjects
Details
- Language :
- English
- Database :
- OpenAIRE
- Accession number :
- edsair.doi.dedup.....acf86689e34bf8fd5c4ea8c298d861f7