title: Microclusters of inhibitory killer immunoglobulin-like receptor signaling at natural killer cell immunological synapses.
abstract: We report the supramolecular organization of killer Ig-like receptor (KIR) phosphorylation using a technique applicable to imaging phosphorylation of any green fluorescent protein-tagged receptor at an intercellular contact or immune synapse. Specifically, we use fluorescence lifetime imaging (FLIM) to report Förster resonance energy transfer (FRET) between GFP-tagged KIR2DL1 and a Cy3-tagged generic anti-phosphotyrosine monoclonal antibody. Visualization of KIR phosphorylation in natural killer (NK) cells contacting target cells expressing cognate major histocompatibility complex class I proteins revealed that inhibitory signaling is spatially restricted to the immune synapse. This explains how NK cells respond appropriately when simultaneously surveying susceptible and resistant target cells. More surprising, phosphorylated KIR was confined to microclusters within the aggregate of KIR, contrary to an expected homogeneous distribution of KIR signaling across the immune synapse. Also, yellow fluorescent protein-tagged Lck, a kinase important for KIR phosphorylation, accumulated in a multifocal distribution at inhibitory synapses. Spatial confinement of receptor phosphorylation within the immune synapse may be critical to how activating and inhibitory signals are integrated in NK cells.
authors.affiliation: Division of Cell and Molecular Biology, Imperial College London, London SW7 2AZ, England, UK.
authors.forename: Bebhinn
authors.initials: B
authors.lastname: Treanor
authors.forename: Peter M P
authors.initials: PM
authors.lastname: Lanigan
authors.forename: Sunil
authors.initials: S
authors.lastname: Kumar
authors.forename: Chris
authors.initials: C
authors.lastname: Dunsby
authors.forename: Ian
authors.initials: I
authors.lastname: Munro
authors.forename: Egidijus
authors.initials: E
authors.lastname: Auksorius
authors.forename: Fiona J
authors.initials: FJ
authors.lastname: Culley
authors.forename: Marco A
authors.initials: MA
authors.lastname: Purbhoo
authors.forename: David
authors.initials: D
authors.lastname: Phillips
authors.forename: Mark A A
authors.initials: MA
authors.lastname: Neil
authors.forename: Deborah N
authors.initials: DN
authors.lastname: Burshtyn
authors.forename: Paul M W
authors.initials: PM
authors.lastname: French
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Davis
mesh_terms: D002460:Cell Line; D021961:Cell Membrane Structures / Q000378:metabolism* / Q000648:ultrastructure; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D007365:Intercellular Junctions / Q000276:immunology* / Q000648:ultrastructure; D007694:Killer Cells, Natural / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D010766:Phosphorylation; D011743:Pyrimidines / Q000494:pharmacology; D011940:Receptor Aggregation / Q000276:immunology; D011971:Receptors, Immunologic / Q000037:antagonists & inhibitors / Q000378:metabolism*; D054340:Receptors, KIR; D054342:Receptors, KIR2DL1; D015398:Signal Transduction / Q000187:drug effects / Q000276:immunology*; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D011743:Pyrimidines; D011971:Receptors, Immunologic; D054340:Receptors, KIR; D054342:Receptors, KIR2DL1; D019061:src-Family Kinases