Although NK cells form a relatively small subset of lymphocytes, they play a very important role in immuno-surveillance of cancer and virus-infected cells and contribute to antigen-specific immune response through production of cytokines (e.g., IFNγ, TNFα), as well as activation of dendritic cells.
Activation of NK cells, mediated by recognition of their ligands by activation receptors, triggers a complex and highly regulated response leading to death of a target cell. Killing of target cell is achieved by localized secretion of lytic granules at the immunological synapse area. A variety of activating receptors and various signaling pathways are involved in NK-cell cytotoxicity; nonetheless, virtually all cytolytic responses require reorganization of the actin cytoskeleton for proper cell adhesion, immune synapse formation, sustained signaling, and delivery of lytic granules to the target cell. However, the orchestration of cytoskeletal elements required for cytolysis is still unclear, and identification of components critical for cytotoxic cell function is of great importance. By comparing physiologic activating and inhibitory signals, we have identified a novel multiprotein complex composed of proteins involved in actin cytoskeleton rearrangements: Wiskott-Aldrich Syndrome protein (WASp), WASp interacting protein (WIP), actin, and myosin IIA. The complex was formed during NK-cell activation, and its formation was disrupted by NK-cell inhibitory signaling, indicating its importance for NK-cell cytotoxic activity and demonstrating that inhibitory signaling in NK cells affects cytoskeletal function.
Subsequently, WIP, an important regulator of WASp activity and actin polymerization, was found to play a critical role in formation of the complex and NK-cell cytotoxicity and was a specific target of KIR inhibitory signaling. Furthermore, WIP over-expression enhances NK-cell cytotoxicity, whereas the disruption of WIP expression abrogates lytic activity of NK cells. Importantly, we demonstrated that WIP is associated with lytic granules in NK cells and has a critical role in their transport to the cell-cell interface (immunological synapse) in response to NK-cell activation. We hypothesize that WIP, a converging point of several signaling pathways, is indispensable for NK-cell function, and WIP activity is controlled by phosphorylation at multiple sites that is regulated alternatively by NK-cell activation and inhibitory signaling.
These studies provided a strong inspiration to expand our interest to the process of lytic granule exocytosis, the last step of NK-cell cytotoxic activity. Membrane fusion events are mediated directly by soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins, often referred to as the “membrane fusion machinery.” Given the critical role of exocytosis in NK-cell function, it is surprising that from almost 40 SNARE proteins in humans only the role of syntaxin 11 (STX11) is well established in NK-cell degranulation, as evidenced by studies of an autosomal recessive disorder, familial hemophagocytic lymphohistiocytosis 4 (FHL4).
Interestingly, although the disease is characterized by an over-activation of the immune system, an impaired function of NK and cytotoxic T cells is a hallmark feature of FHL, due to a deletion or a nonsense mutation in the STX11 gene, resulting in defects in the exocytic pathway. However, the protein machinery involved in lytic granule exocytosis is still poorly understood in NK cells, and one of the fundamental problems is to define and understand the processes regulating exocytosis in NK-cell function.
We hypothesize that NK cells use a specific set of SNARE proteins for the exocytosis of lytic granules, distinct from SNARE proteins utilized during endocytosis, and possibly partially overlapping with cytokine exocytosis machinery.
Dr. Krzewski received his Ph.D. from the University of Gdansk, Poland, in 2002. He did his postdoctoral training in the laboratory of Jack Strominger, M.D., at Harvard University, where he developed his interest in natural killer (NK) cells and their biology and investigated regulation of cytolytic activity of NK cells. In 2009, he joined NIAID as a staff scientist to work with John E. Coligan, Ph.D., in the Receptor Cell Biology Section.
Peruzzi G, Femnou L, Gil-Krzewska A, Borrego F, Weck J, Krzewski K, Coligan JE. Membrane-type 6 matrix metalloproteinase regulates the activation-induced downmodulation of CD16 in human primary NK cells. J Immunol. 2013 Jul 12. Epub ahead of print.
Krzewski K, Cullinane AR. Evidence for defective Rab GTPase-dependent cargo traffic in immune disorders. Exp Cell Res. 2013 Jun 26. Epub ahead of print.
Krzewski K, Gil-Krzewska A, Nguyen V, Peruzzi G, Coligan JE. LAMP1/CD107a is required for efficient perforin delivery to lytic granules and NK-cell cytotoxicity. Blood. 2013 Jun 6;121(23):4672-83.
Krzewski K, Coligan JE. Human NK cell lytic granules and regulation of their exocytosis. Front Immunol. 2012;3:335.
Park YP, Choi SC, Kiesler P, Gil-Krzewska A, Borrego F, Weck J, Krzewski K, Coligan JE. Complex regulation of human NKG2D-DAP10 cell surface expression: opposing roles of the γc cytokines and TGF-β1. Blood. 2011 Sep 15;118(11):3019-27.
Krzewski K, Gil-Krzewska A, Watts J, Stern JN, Strominger JL. VAMP4- and VAMP7-expressing vesicles are both required for cytotoxic granule exocytosis in NK cells. Eur J Immunol. 2011 Nov;41(11):3323-9.
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Last Updated August 21, 2013