Short-term IL-15 priming leaves a long-lasting signalling imprint in mouse NK cells independently of a metabolic switch

Abstract

IL-15 priming of NK cells is a broadly accepted concept, but the dynamics and underlying molecular mechanisms remain poorly understood. We show that as little as 5 min of IL-15 treatment in vitro, followed by removal of excess cytokines, results in a long-lasting, but reversible, augmentation of NK cell responsiveness upon activating receptor cross-linking. In contrast to long-term stimulation, improved NK cell function after short-term IL-15 priming was not associated with enhanced metabolism but was based on the increased steady-state phosphorylation level of signalling molecules downstream of activating receptors. Inhibition of JAK3 eliminated this priming effect, suggesting a cross talk between the IL-15 receptor and ITAM-dependent activating receptors. Increased signalling molecule phosphorylation levels, calcium flux, and IFN-γ secretion lasted for up to 3 h after IL-15 stimulation before returning to baseline. We conclude that IL-15 rapidly and reversibly primes NK cell function by modulating activating receptor signalling. Our findings suggest a mechanism by which NK cell reactivity can potentially be maintained in vivo based on only brief encounters with IL-15 trans-presenting cells.

piggyBac system to co-express NKG2D CAR and IL-15 to augment the in vivo persistence and anti-AML activity of human peripheral blood NK cells

“…Since short-term IL-15 priming could significantly increase the killing capacity of mouse NK cells and human NK cells, expressing membrane-bound IL-15 displayed a significantly higher cytolytic activity against a set of cancer cell lines in a short-term cytotoxicity assay, 44 , 45 we mixed CAR-NK cells with target cancer cells at low effector to target (E:T) cell ratios from 0.5:1 to 4:1 and performed 3-h in vitro cytotoxicity assays. The killing efficiencies of NKG2D CAR/IL15-NK cells against three tested cancer lines significantly increased over those offered by NKG2D CAR-NK cells ( Figure 4 D).…”

Control of NK cell tolerance in MHC class I-deficiency by regulated SHP-1 localization to the activating immune synapse

“…A cellular barcoding protocol was again used to minimize experimental variability (Fig. S4a) 45 . Besides signaling molecules implicated as direct SHP-1 targets, we also examined activation of ERK1/2, a down-stream read-out of NK cell activation 46 .…”

“…For phosphoflow experiments, enriched NK cells were stained with surface antibody and biotin-rat-anti mouse NKp46 (29A1.4, Biolegend) as described above. The stimulation, barcoding and staining were performed as described in our previous publications 45, 80 . The barcoding dyes which were used in this study were Pacific Orange and Alexa Fluor 700 NHS esters (Invitrogen).…”

“…Mouse NK cells were enriched from a single cell suspension of total splenocytes using a customized negative selection protocol as described previously 45 . Human NK cells were enriched from peripheral blood mononuclear cells (PBMC) using the human NK cell isolation kit (Miltenyi).…”

Dynamics of IL‐15/IL‐15R‐α expression in response to HSV‐1 infection reveal a novel mode of viral immune evasion counteracted by iNKT cells

“…The latter mechanism accounts for most of the effects of IL-15 on immune responses [17]. In the context of barrier immunology, IL-15 is critical for a variety of local tissue immune functions [18], and acts to activate multiple immune cell types, including NK cells [19,20], CD8 + T cells [21], γδ T cells [22], and mucosa-associated invariant T cells [23]. IL-15 is expressed by many cell types, and in the context of skin, HSV-1-infected DCs stimulate CD8 + T-cell priming through IL-15 production [24].…”