Src-Dependent Syk Activation Controls CD69-Mediated Signaling and Function on Human NK Cells

NK cells are defined as those cells that lyse tumor cells without priming. In this study, we show that the preincubation of resting human NK cells with the leukemia cell CTV-1 primes NK cells to lyse NK-resistant cell lines, primary leukemias, and solid tumors even when HLA-matched, allogeneic or autologous. The primed NK cells remained nonresponsive to HLA-C matched and mismatched normal mononuclear cells from multiple donors. CD69, a known NK trigger receptor, was shown to be the predominant trigger on the tumor-primed NK cells because lysis was blocked with the rCD69 protein. The lack of lytic activity against normal hemopoietic cells implied that the ligand for CD69 is tumor restricted, and this was confirmed by experiments using fluorochrome labeled rCD69. It has been recently shown that resting NK cells require prior stimulation with IL-2 before triggering by all known NK-triggering ligands. In this study, we show that a tumor cell can provide the NK priming signal independently of IL-2. These data provide evidence for two NK evasion strategies for tumor cells, namely the prevention of priming (type1 evasion) and failure to trigger (type 2 evasion). Most NK-resistant cell lines are type 1 and fail to prime resting NK cells but are lysed by IL-2-primed NK cells. In contrast, CTV-1 cells prime resting NK cells but fail to trigger (type 2), and coincubation with CTV-1 primes for triggering by type 1 NK-resistant tumor cells. These tumor-activated NK cells lyse a broad spectrum of tumor cells with a degree of specificity never previously reported.

Section: Discussionsupporting

confidence: 57%

Tumor-Primed Human Natural Killer Cells Lyse NK-Resistant Tumor Targets: Evidence of a Two-Stage Process in Resting NK Cell Activation

North

Bakhsh

Marden

et al. 2007

“…In addition, our studies showed that anti-FasL mAb only partially blocked DNA fragmentation in Jurkat cells, suggesting the involvement of other apoptosis-inducing elements, such as TNF-related apoptosis-inducing ligand (2) or other TNF family members (53)(54)(55). It should also be noted that 2DL4-induced expression of CD25 (IL-2R␣ chain) and CD69 (an activation receptor) (56,57) indicates that, in addition to promoting strong IFN-␥ production and Fas-mediated target cell cytotoxicity, engagement of the receptor may prime NK cells to respond subsequently to IL-2 and target cells in vivo.…”

Section: Discussionmentioning

confidence: 74%

KIR2DL4 Is an IL-2-Regulated NK Cell Receptor That Exhibits Limited Expression in Humans but Triggers Strong IFN-γ Production

Kikuchi-Maki

Yusa

Catina

et al. 2003

156

148

Killer cell Ig-like receptor (KIR)2DL4 (2DL4, CD158d) was previously described as the only KIR expressed by every human NK cell. It is also structurally atypical among KIRs because it possesses a basic transmembrane residue, which is characteristic of many activating receptors, but also contains a cytoplasmic immunoreceptor tyrosine-based inhibitory motif (ITIM). We expressed epitope-tagged 2DL4 in an NK-like cell line to study receptor function. Three distinct 2DL4 cDNA clones were analyzed: one encoding the “conventional” 2DL4 with the cytoplasmic ITIM (2DL4.1) and two encoding different cytoplasmic truncated forms lacking the ITIM (2DL4.2 and 2DL4∗). Surprisingly, one truncated receptor (2DL4.2), which is the product of a prevalent human 2DL4 allele, was not expressed on the cell surface, indicating that some individuals may lack functional 2DL4 protein expression. Conversely, both 2DL4.1 and 2DL4∗ were expressed on the cell surface and up-regulated by IL-2. Analysis of primary NK cells with anti-2DL4 mAb confirmed the lack of surface expression in a donor with the 2DL4.2 genotype. Donors with the 2DL4.1 genotype occasionally expressed receptor only on CD56high NK cells, although their expression was up-regulated by IL-2. Interestingly, Ab engagement of epitope-tagged 2DL4 triggered rapid and robust IFN-γ production, but weak redirected cytotoxicity in an NK-like cell line, which was the opposite pattern to that observed upon engagement of another NK cell activating receptor, NKp44. Importantly, both 2DL4.1 and 2DL4∗ exhibited similar activation potential, indicating that the ITIM does not influence 2DL4.1 activating function. The unique activation properties of 2DL4 suggest linkage to a distinct signaling pathway.

“…CNF1, which in our instance reinforced the cytotoxic activity of NK cells, also augmented the expression of certain surface molecules, e.g., cell adhesion molecules such as CD2, or activation-associated molecules such as CD69. In particular, the cross-linking of the triggering receptor CD69 is known to induce a cytotoxic activity in activated NK cells by triggering tyrosine phosphorylation and consequently activation of an array of molecules comprising the Rho family-specific exchange factor Vav1 (40,41). Of interest, CD69 expression was significantly augmented in NK cells challenged with CNF1.…”

Section: Discussionmentioning

confidence: 99%

The Rac-Activating Toxin Cytotoxic Necrotizing Factor 1 Oversees NK Cell-Mediated Activity by Regulating the Actin/Microtubule Interplay

Malorni

Quaranta

Straface

et al. 2003

The cell cytoskeleton is widely acknowledged as a master for NK cell function. Specifically, actin filaments guide the NK cell binding to target cells, engendering the formation of the so-called immunological synapse, while microtubules direct the killer behavior. All these cytoskeleton-dependent activities are competently governed by the Rho GTPases, a family of regulatory molecules encompassing the three different subfamilies, Rho, Rac, and Cdc42. By using a Rac GTPase-activating bacterial protein toxin from Escherichia coli named cytotoxic necrotizing factor 1 (CNF1), we obtained results supporting the activation of Rac GTPase as a booster for effector cell-binding efficiency, recruitment ability, and, consequently, cytotoxicity. In particular, the augmented killer capacity of CNF1-treated NK cells was associated with the increased expression of certain cell adhesion or activation-associated molecules and the reshaping of the actin and microtubule networks. Importantly, CNF1 counteracted the activity exerted by toxins disrupting the cytoskeletal architecture. Hence, the activation of Rho GTPases, particularly Rac, induced by CNF1, appears to orchestrate a dynamic cross talk between microtubules and actin filaments, leading to a fruitful NK cell activity and polarization state. Our findings suggest that protein toxins might be viewed as modulators of NK cell cytotoxic activity and could possibly be regarded as useful pharmacological tools for certain Rho-linked immune diseases in the near future.