Super-resolution imaging of remodeled synaptic actin reveals different synergies between NK cell receptors and integrins

Abstract

Natural killer (NK) cells secrete lytic granules to directly kill virus-infected or transformed cells and secrete cytokines to communicate with other cells. Threedimensional super-resolved images of F-actin, lytic granules, and IFN-␥ in primary human NK cells stimulated through different activating receptors reveal that both IFN-␥ and lytic granules accumulated in domains where the periodicity of the cortical actin mesh at the synapse opened up to be penetrable. Ligation of some activating receptors alone (eg, CD16 or NKG2D) was sufficient to increase the periodicity of the actin mesh, but surprisingly, ligation of others (eg, NKp46 or CD2) was not sufficient to induce cortical actin remodeling unless LFA-1 was coligated. Importantly, influenza virus particles that can be recognized by NK cells similarly did not open the actin mesh but could if LFA-1 was coligated. This leads us to propose that immune cells using germline-encoded receptors to directly recognize foreign proteins can use integrin recognition to differentiate between free pathogens and pathogen-infected cells that will both be present in blood. This distinction would not be required for NK cell receptors, such as NKG2D, which recognize host cell-encoded proteins that can only be found on diseased cells and not pathogens. (Blood. 2012;120(18):3729-3740) IntroductionNatural killer (NK) cells are innate lymphocytes whose responses are controlled through the balance of signals from germlineencoded activating and inhibitory receptors. 1 Integration of signals occurs across a structured interface, termed the immune synapse, between NK cells and target cells. 2,3 The spatial and temporal organization of the synapse is important for coordinating interactions between immune cell receptors, kinases, phosphatases, and adaptors as well as for directing the interaction between cells. [4][5][6] When signals downstream of activating receptors dominate, a cytolytic NK cell synapse can be assembled across which lytic molecules are secreted toward the target cell. [7][8][9] One of the best characterized NK cell activating receptors is NKG2D, which recognizes stress-induced ligands, such as MHC class I chain-related protein A (MICA), and triggers NK cells to stop migrating, spread symmetrically, and activate cytoskeletal reorganization. 8,10,11 Super-resolution microscopy revealed that remodeling of cortical actin occurs in domains within the central region of the synapse establishing secretory domains where lytic granules dock. 12,13 It is also established that cytokines can be secreted directionally across immune synapses. 14 In particular, IFN-␥ has been demonstrated to be directionally secreted in vitro from T cells to antigen-presenting cells and in vivo between T cells and brain cells. [15][16][17][18] NK cell-mediated IFN-␥ secretion is important for shaping the Th1 immune response, modulating dendritic cell and macrophage activation and stimulating antiproliferative effects in virus-infected or transformed cells. [19][20][21][22] NK cell expression of IFN-␥ c...

Lytic immune synapse function requires filamentous actin deconstruction by Coronin 1A

“…With increased density of synaptic F-actin, it was possible that degranulation is obstructed in the absence of Coro1A, because lytic granules are secreted through hypodense regions of the F-actin network at the NK cell synapse (12)(13)(14). To determine if Coro1A is required for the formation of granule-permissive openings in actin meshworks, we measured activation-induced clearances in the synaptic F-actin network using STED imaging.…”

“…Although cortical F-actin has long been considered a barrier to exocytosis of granule-like organelles (11) in some cell types, ligation of NK cell activating and adhesion receptors results in the formation of conduits in F-actin that permit and actually facilitate NK cell degranulation (12)(13)(14). This finding suggests that fine regulation and deconstruction of the synaptic F-actin meshwork is required for the formation of granule-permissivesized clearances (12).…”

Lenalidomide augments actin remodeling and lowers NK-cell activation thresholds

“…[3][4][5][6][7][8] Superresolution microscopy revealed that activating receptor ligation triggers remodeling of cortical actin in specific domains within the NK-cell immune synapse where lytic granules and vesicles containing IFN-g accumulate. [9][10][11][12][13] Multiple myeloma (MM) is a hematologic malignancy characterized by a clonal proliferation of plasma cells in bone marrow and is associated with progressive dysregulation of the immune system. 14 NK cells may initially contribute to the control of malignant cells [15][16][17] and evidence suggests that NKG2D is involved in NK-cell recognition of MM cells.…”

Ras-related C3 Botulinum Toxin Substrate (Rac) and Src Family Kinases (SFK) Are Proximal and Essential for Phosphatidylinositol 3-Kinase (PI3K) Activation in Natural Killer (NK) Cell-mediated Direct Cytotoxicity against Cryptococcus neoformans

“…2). As mentioned above, LFA-1, which has also been implicated in conjugate formation of NK cells, was not involved in cryptococcal killing (45,46). Although conjugate formation can be achieved by other signaling pathways, the lack of Rac and LFA-1 involvement may suggest an unidentified actin remodeling process (45).…”