IL-2 induces a WAVE2-dependent pathway for actin reorganization that enables WASp-independent human NK cell function
Jordan S. Orange, Sumita Roy-Ghanta, Emily M. Mace, Saumya Maru, Gregory D. Rak, Keri B. Sanborn, Anders Fasth, Rushani Saltzman, Allison Paisley, Linda Monaco-Shawver, Pinaki P. Banerjee, Rahul Pandey
Jordan S. Orange, Sumita Roy-Ghanta, Emily M. Mace, Saumya Maru, Gregory D. Rak, Keri B. Sanborn, Anders Fasth, Rushani Saltzman, Allison Paisley, Linda Monaco-Shawver, Pinaki P. Banerjee, Rahul Pandey
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Research Article Immunology

IL-2 induces a WAVE2-dependent pathway for actin reorganization that enables WASp-independent human NK cell function

Abstract

Wiskott-Aldrich syndrome (WAS) is a primary immunodeficiency associated with an increased susceptibility to herpesvirus infection and hematologic malignancy as well as a deficiency of NK cell function. It is caused by defective WAS protein (WASp). WASp facilitates filamentous actin (F-actin) branching and is required for F-actin accumulation at the NK cell immunological synapse and NK cell cytotoxicity ex vivo. Importantly, the function of WASp-deficient NK cells can be restored in vitro after exposure to IL-2, but the mechanisms underlying this remain unknown. Using a WASp inhibitor as well as cells from patients with WAS, we have defined a direct effect of IL-2 signaling upon F-actin that is independent of WASp function. We found that IL-2 treatment of a patient with WAS enhanced the cytotoxicity of their NK cells and the F-actin content at the immunological synapses formed by their NK cells. IL-2 stimulation of NK cells in vitro activated the WASp homolog WAVE2, which was required for inducing WASp-independent NK cell function, but not for baseline activity. Thus, WAVE2 and WASp define parallel pathways to F-actin reorganization and function in human NK cells; although WAVE2 was not required for NK cell innate function, it was accessible through adaptive immunity via IL-2. These results demonstrate how overlapping cytoskeletal activities can utilize immunologically distinct pathways to achieve synonymous immune function.

Authors

Jordan S. Orange, Sumita Roy-Ghanta, Emily M. Mace, Saumya Maru, Gregory D. Rak, Keri B. Sanborn, Anders Fasth, Rushani Saltzman, Allison Paisley, Linda Monaco-Shawver, Pinaki P. Banerjee, Rahul Pandey

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Figure 3

IL-2 increases cytotoxicity and F-actin reorganization in wiskostatin-treated NK cells.

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IL-2 increases cytotoxicity and F-actin reorganization in wiskostatin-tr...
(A) Cytotoxic activity of YTS cells (left), NK92 cells (middle), or PBMCs (right) pretreated with vehicle control (c, black solid line), IL-2 (gray solid line), wiskostatin (Wisk, black dashed line), and wiskostatin followed by IL-2 (Wisk+IL-2, gray dashed line). Cells were incubated with IL-2 where specified for 30 minutes prior to the assay. K562 target cells were used for NK92, and 721.221 target cells for YTS and PBMCs. Cytotoxicity was measured by 4-hour 51Cr-release assays. Each point represents the mean of 5 individual assays, each performed in triplicate + SD. For each graph, decreases mediated by wiskostatin and increases in wiskostatin-treated cells mediated by IL-2 were significant (*P < 0.05, 2-tailed Mann-Whitney U test). (B) Change in F-actin content measured as percentage increase in phalloidin MFI via flow cytometry in YTS cells (left), NK92 cells (middle), or PBMCs (right). Vehicle control-treated cells were compared with IL-2–treated (black), wiskostatin-treated (gray), and wiskostatin- and IL-2–treated (white). Each bar represents the mean of 3 or more experiments ± SD (*P < 0.05, significant changes from control-treated cells). 30-minute wiskostatin pretreatment was followed by 30-minute IL-2 treatment. (C) Representative TEM images of platinum rotary-shadowed, sonicated, and extracted YTS cells that had been adhered to glass coated with CD18 and anti-CD28. Prior to adherence to the glass, cells were treated with vehicle (left), wiskostatin (center), or wiskostatin followed by IL-2 (right). Cortical F-actin network detail is shown. Original magnification, ×6000 (scale bar: 5 μm [top]) and ×30,000 (scale bar: 1 μm [bottom]).