Abstract
The adhesion and degranulation-promoting adaptor protein (ADAP) serves as a multifunctional scaffold and is involved in the formation of immune signaling complexes. To date only limited and moreover conflicting data exist regarding the role of ADAP in NK cells. To extend existing knowledge we investigated ADAP-dependency of NK cells in the context of in vivo infection with the intracellular pathogen Listeria monocytogenes (Lm). Ex vivo analysis of infection-primed NK cells revealed impaired cytotoxic capacity in NK cells lacking ADAP as indicated by reduced CD107a surface expression and inefficient perforin production. However, ADAP-deficiency had no global effect on NK cell morphology or intracellular distribution of CD107a-containing vesicles. Proteomic definition of ADAPko and wild type NK cells did not uncover obvious differences in protein composition during the steady state and moreover, similar early response patterns were induced in NK cells upon infection independent of the genotype. In line with protein network analyses that suggested an altered migration phenotype in naïve ADAPko NK cells, in vitro migration assays uncovered significantly reduced migration of both naïve as well as infection-primed ADAPko NK cells compared to wild type NK cells. Notably, this migration defect was associated with a significantly reduced expression of the integrin CD11a on the surface of splenic ADAP-deficient NK cells 1 day post-Lm infection. We propose that ADAP-dependent alterations in integrin expression might account at least in part for the fact that during in vivo infection significantly lower numbers of ADAPko NK cells accumulate in the spleen i.e., the site of infection. In conclusion, we show here that during systemic Lm infection in mice ADAP is essential for efficient cytotoxic capacity and migration of NK cells.
Highlights
The coupling of transmembrane receptors to intracellular signaling pathways is mediated by adapter proteins that are made up of various protein domains without enzymatic or transcriptional activity
To first confirm the role of adaptor protein (ADAP) in cytokine production by naïve NK cells, they were isolated from the spleen of naïve wild type and conventional ADAP wild type and knock out (ADAPko) mice and were stimulated in vitro with anti-NK1.1 alone or in combination with IL-2/IL-12 or with PMA/ionomycin followed by flow cytometric analysis of IFN-γ production
We extended our analyses to an experimental setting allowing NK cell priming within their natural environment, i.e., during in vivo infection, and to a more physiological NK cell in vitro stimulation set-up using YAC-1 target cells
Summary
The coupling of transmembrane receptors to intracellular signaling pathways is mediated by adapter proteins that are made up of various protein domains without enzymatic or transcriptional activity. The Adhesion and degranulation-promoting adapter protein ADAP, known as Fyn-binding protein (Fyb) or SLAP-130 serves, amongst others, as a scaffold adapter protein specific for the hematopoietic lineage that so far has been mainly studied in the context of the activation of and effector functions in T cells [2]. ADAP consists of several domains that can associate with proteins involved in cell migration, cellular adhesion and re-arrangement of the cytoskeleton in T cells [3]. ADAP-deficient T cells show reduced migration toward chemokines [6], impaired formation of the immunological synapse [4, 5] and impaired activation, differentiation and resident memory formation during acute infections [6, 7]
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