PMID- 27682583 OWN - NLM STAT- MEDLINE DCOM- 20170905 LR - 20181113 IS - 1939-4586 (Electronic) IS - 1059-1524 (Print) IS - 1059-1524 (Linking) VI - 27 IP - 22 DP - 2016 Nov 7 TI - Functional nanoscale coupling of Lyn kinase with IgE-FcepsilonRI is restricted by the actin cytoskeleton in early antigen-stimulated signaling. PG - 3645-3658 AB - The allergic response is initiated on the plasma membrane of mast cells by phosphorylation of the receptor for immunoglobulin E (IgE), FcepsilonRI, by Lyn kinase after IgE-FcepsilonRI complexes are cross-linked by multivalent antigen. Signal transduction requires reorganization of receptors and membrane signaling proteins, but this spatial regulation is not well defined. We used fluorescence localization microscopy (FLM) and pair-correlation analysis to measure the codistribution of IgE-FcepsilonRI and Lyn on the plasma membrane of fixed cells with 20- to 25-nm resolution. We directly visualized Lyn recruitment to IgE-FcepsilonRI within 1 min of antigen stimulation. Parallel FLM experiments captured stimulation-induced FcepsilonRI phosphorylation and colocalization of a saturated lipid-anchor probe derived from Lyn's membrane anchorage. We used cytochalasin and latrunculin to investigate participation of the actin cytoskeleton in regulating functional interactions of FcepsilonRI. Inhibition of actin polymerization by these agents enhanced colocalization of IgE-FcepsilonRI with Lyn and its saturated lipid anchor at early stimulation times, accompanied by augmented phosphorylation within FcepsilonRI clusters. Ising model simulations provide a simplified model consistent with our results. These findings extend previous evidence that IgE-FcepsilonRI signaling is initiated by colocalization with Lyn in ordered lipid regions and that the actin cytoskeleton regulates this functional interaction by influencing the organization of membrane lipids. CI - (c) 2016 Shelby et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution-Noncommercial-Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0). FAU - Shelby, Sarah A AU - Shelby SA AD - Department of Chemistry and Chemical Biology and Field of Biophysics, Cornell University, Ithaca, NY 14853. FAU - Veatch, Sarah L AU - Veatch SL AD - Department of Biophysics, University of Michigan, Ann Arbor, MI 48109. FAU - Holowka, David A AU - Holowka DA AD - Department of Chemistry and Chemical Biology and Field of Biophysics, Cornell University, Ithaca, NY 14853. FAU - Baird, Barbara A AU - Baird BA AD - Department of Chemistry and Chemical Biology and Field of Biophysics, Cornell University, Ithaca, NY 14853 bab13@cornell.edu. LA - eng GR - R01 GM110052/GM/NIGMS NIH HHS/United States GR - R01 GM117552/GM/NIGMS NIH HHS/United States GR - T32 GM008267/GM/NIGMS NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural DEP - 20160928 PL - United States TA - Mol Biol Cell JT - Molecular biology of the cell JID - 9201390 RN - 0 (Receptors, IgE) RN - 37341-29-0 (Immunoglobulin E) RN - EC 2.7.10.2 (lyn protein-tyrosine kinase) RN - EC 2.7.10.2 (src-Family Kinases) SB - IM MH - Actin Cytoskeleton/metabolism MH - Animals MH - Cell Culture Techniques MH - Cell Membrane/metabolism MH - Computer Simulation MH - Immunoglobulin E/metabolism MH - Mast Cells/metabolism MH - Mice MH - Microscopy, Fluorescence MH - Phosphorylation MH - Receptors, IgE/*metabolism MH - Signal Transduction/physiology MH - src-Family Kinases/*metabolism PMC - PMC5221596 EDAT- 2016/11/05 06:00 MHDA- 2017/09/07 06:00 PMCR- 2017/01/22 CRDT- 2016/09/30 06:00 PHST- 2016/06/16 00:00 [received] PHST- 2016/09/20 00:00 [accepted] PHST- 2016/11/05 06:00 [pubmed] PHST- 2017/09/07 06:00 [medline] PHST- 2016/09/30 06:00 [entrez] PHST- 2017/01/22 00:00 [pmc-release] AID - mbc.E16-06-0425 [pii] AID - E16-06-0425 [pii] AID - 10.1091/mbc.E16-06-0425 [doi] PST - ppublish SO - Mol Biol Cell. 2016 Nov 7;27(22):3645-3658. doi: 10.1091/mbc.E16-06-0425. Epub 2016 Sep 28.