Abstract
Cell surface receptors that bind the Fc segment of antibodies to initiate signaling play fundamental roles in immune responses. Multiple, diverse Fc receptors (e.g., Fc gamma, Fc-alpha, and Fc-epsilon) are expressed on different immune cells, including natural killer cells, macrophages, mast cells, and neutrophils. Fc receptors bind particular antibody isotypes (e.g., IgG, IgA, IgE, respectively) thereby sensitizing the cells to their specific antigens. Receptor clustering by antigen or other multivalent ligands induces a signaling cascade that leads to targeted secretion of chemical mediators (e.g., histamine, cytokines, and chemokines) and other cell-specific responses. Spatial targeting and compartmentalization are common mechanisms for regulating Fc receptor signaling. However, the tools for studying these dynamic interactions at cellular levels have been limited due to the nanoscale dimensions of the signaling complexes and their dispersal across the cell surface. To overcome these limitations in our model system, we use microfabricated surfaces containing spatially defined ligands to cluster and activate IgE receptors (FcεRI), which initiate allergic responses by mast cells. Micron-scale control of receptor assemblies allows investigation with conventional fluorescence microscopy of spatially regulated redistributions of intracellular signaling components. This approach in conjunction with biochemical techniques has proven valuable for investigating immune receptor signaling.
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Acknowledgments
This work was supported by NIH grant R01-AI18306 followed by R01-GM117552. We appreciate experimental resources provided by the Cornell NanoScale Science and Technology Facility (NSF ECCS-1542081) and the Cornell Biotechnology Resource Center Imaging Facility, including the Zeiss LSM 710 confocal microscope (NIH S10RR025502). This chapter is significantly updated from an original edition [16]. Recent advances in the methodology include those made by Devin Wakefield [8].
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Mohr, J.D., Ramezani, M., Holowka, D., Baird, B.A. (2022). Micropatterned Ligand Arrays to Investigate Spatial Regulation of Cellular Signaling Initiated by Clustered Fc Receptors. In: Rast, J., Buckley, K. (eds) Immune Receptors. Methods in Molecular Biology, vol 2421. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1944-5_1
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DOI: https://doi.org/10.1007/978-1-0716-1944-5_1
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