Elsevier

Molecular Immunology

Volume 42, Issue 4, February 2005, Pages 501-510
Molecular Immunology

Review
Activation of NK cell cytotoxicity

https://doi.org/10.1016/j.molimm.2004.07.034Get rights and content

Abstract

Natural killer (NK) cells are innate effector lymphocytes necessary for defence against stressed, microbe-infected, or malignant cells. NK cells kill target cells by either of two major mechanisms that require direct contact between NK cells and target cells. In the first pathway, cytoplasmic granule toxins, predominantly a membrane-disrupting protein known as perforin, and a family of structurally related serine proteases (granzymes) with various substrate specificities, are secreted by exocytosis and together induce apoptosis of the target cell. The granule-exocytosis pathway potently activates cell-death mechanisms that operate through the activation of apoptotic cysteine proteases (caspases), but can also cause cell death in the absence of activated caspases. The second pathway involves the engagement of death receptors (e.g. Fas/CD95) on target cells by their cognate ligands (e.g. FasL) on NK cells, resulting in classical caspase-dependent apoptosis. The comparative role of these pathways in the pathophysiology of many diseases is being dissected by analyses of gene-targeted mice that lack these molecules, and humans who have genetic mutations affecting these pathways. We are also now learning that the effector function of NK cells is controlled by interactions involving specific NK cell receptors and their cognate ligands, either on target cells, or other cells of the immune system. This review will discuss the functional importance of NK cell cytotoxicity and the receptor/ligand interactions that control these processes.

Introduction

Natural killer (NK) cells are specialized innate lymphocytes capable of responding to virus-infected and tumor cells. Unlike cytotoxic T lymphocytes (CTL), NK cells do not require antigen-specific recognition to kill target cells, and as such are capable of limiting viral infection prior to the induction of adaptive immune responses. Compelling evidence for a critical role for NK cells in limiting viral infection has been provided by studies with herpesviruses, such as cytomegalovirus (CMV), herpes simplex virus (HSV) and Epstein-Barr virus (EBV), as well as HIV (Biron et al., 1999). NK cells also exhibit spontaneous cytotoxicity against major histocompatibility complex (MHC) class I-deficient target cells, and in particular they participate in the innate immune responses against transformed cells and tumor metastases in vivo (Smyth et al., 1999b, Smyth et al., 2002). The effector functions of NK cells, including cytotoxicity and the capacity to produce a variety of cytokines (including IFN-γ) following activation, are of pathophysiological importance. Granule exocytosis is the major mechanism of killing used by NK cells, however, these cells also express members of the tumor necrosis factor (TNF) superfamily, the role of which has recently begun to be elucidated. NK cell function is tightly regulated by a balance between positive and negative signals provided by a diverse array of cell surface receptors (Cerwenka and Lanier, 2001). Activation requires the action of pro-inflammatory cytokines in combination with differential engagement of cell surface receptors. In particular, cytokines such as IL-12, IL-15, IL-18, IL-21 and IFN-αβ can induce NK cell proliferation, as well as promoting NK cell cytotoxicity and/or production of IFN-γ (Biron et al., 1999). NK cells are inhibited by receptors that recognize MHC class I molecules, and thus healthy cells expressing normal levels of MHC class I are generally protected from NK cell-mediated lysis. Virus-infected and malignant cells may express reduced levels of MHC class I molecules and thus, provided that they also express appropriate ligands, they become vulnerable to NK cell attack. Thus triggering of activating receptors, accompanied by reduced signaling through inhibitory receptors, leads to NK cell activation. The activation of NK cells also results from the concerted action of co-stimulatory molecules already well characterized for their function in T cells. In this review we focus on the role and regulation of NK cell cytotoxicity.

Section snippets

NK cell effector functions

The direct killing of cancer or pathogen-infected cells is just one component of the total NK cell response. NK cell production of cytokines, such as IFN-γ, also restricts tumor angiogenesis and stimulates adaptive immunity. IFN-γ, secreted by NK cells, plays a critical role in suppressing pathogen challenge, both to contain the initial infection, and to promote an appropriate adaptive response that may take several days to mature. NK cells mediate cell killing though a variety of mechanisms,

Granule exocytosis – an introduction

The cytotoxic granules of NK cells are complex organelles that combine specialized storage and secretory functions with the general degradative functions of typical lysosomes. Granules package the mediators of a diverse range of cell-death pathways that have evolved to rapidly kill cells harbouring intracellular pathogens. Although pathogens have devised ways to prolong the life of an infected cell by blocking apoptotic cell death, the death mediated by an activated NK cell remains rapid,

Granulysin

Granulysin is a member of the saposin-like protein family that includes amoebapores and NK lysin. It is a cytolytic protein present in the granules of human CTLs and NK cells (Clayberger and Krensky, 2003) and is lytic against both microbes and tumors (Gamen et al., 1998, Stenger et al., 1998). Its structure suggests a potential mechanism of action whereby granulysin functions as a lytic molecule; the positive charges of granulysin appear to orient the molecule towards the negatively charged

Biological relevance of NK cell cytotoxic mechanisms

The relative importance of each of the granule proteins in NK cell cytotoxicity has been evaluated in vivo using gene-targeted mice, and deduced from patients carrying specific mutations in relevant genes. Granule proteins are expressed by most cytotoxic lymphocytes, including NK cells, CTLs, NKT cells and γδ+T cells. For the purpose of this review, we will limit the discussion of granule proteins to their role in NK cell function.

Functional consequences of perforin deficiency

Studies in gene-disrupted mice indicate that perforin is vital for NK cell cytotoxicity (Kagi et al., 1994) and it has an indispensable, but undefined, role in Grz-mediated apoptosis. The mapping of perforin mutations in a lethal, inherited human disorder of immune dysregulation known as familial haemophagocytic lymphohistiocytosis (FHL), also highlights the important role for perforin in human NK cell cytotoxicity (Stepp et al., 1999). A thorough examination of individual mutations in FHL

Functional consequences of Grz and granulysin deficiency

The role of Grz in NK cell cytotoxic responses remains comparatively poorly defined in vivo. Individual Grz show considerable functional redundancy, so that mice that are deficient in one, or even several Grz, have more focal immune deficits than perforin-deficient mice on the same genetic background. The NK cells of mice that lack GrzA (Ebnet et al., 1995) induce morphologically normal apoptosis in target cells in vitro, and those of mice that are deficient in the GrzB cluster (Heusel et al.,

TRAIL

Members of the TNF family of cytokines are expressed by NK cells, and are important mediators of apoptosis that both shape and regulate the immune system. TNF-related apoptosis-inducing ligand (TRAIL), also known as Apo2 ligand, is a type II transmembrane protein belonging to the TNF superfamily. Five receptors for TRAIL have been identified in humans, and two of them, TRAIL-R1 (DR4) and TRAIL-R2 (DR5), are capable of transducing an apoptotic signal (Degli-Esposti, 1999). The other three

Receptor classes – an introduction

When NK cell inhibitory receptors bind to MHC class I molecules, their effector functions (cytotoxicity and cytokine production) are blocked. Inhibitory receptors specific for MHC class I or MHC class I-related molecules can provide protection for target cells that express normal levels of class I molecules on their surface. Three such inhibitory receptor families have been discovered and characterized: the killer cell Ig-like receptors (KIR) in humans, the Ly49 lectin-like receptors in mice

Conclusions

There is still a great deal to learn about the biological importance of various NK cell cytotoxic pathways. In particular, a good understanding of what role Grz play in NK cell function remains to be established. It is still not clear whether Grz have important pro-apoptotic functions in vivo and at present their physiological importance appears restricted to particular viral infections. Perforin and TRAIL have emerged as important effectors in NK cell cytotoxicity against both tumors and

Acknowledgements

We thank Joe Trapani for helpful discussions. Work in our laboratories was supported by a Cancer Research Institute Post-Doctoral Fellowship to YH, an Melbourne Research Scholarship to EC, a Human Frontiers Science Program research grant to MJS and HY, National Health and Medical Research Council of Australia Project Grants to MJS and MAD-E, a Program Grant to MJS, a Dora Lush Postgraduate Award to SEAS and a Research Fellowship to MJS. MAD-E is supported by a Wellcome Trust Overseas Senior

References (95)

  • B.J. Rukamp et al.

    Subsite specificities of granzyme M: a study of inhibitors and newly synthesized thiobenzyl ester substrates

    Arch. Biochem. Biophys.

    (2004)
  • M.J. Smyth et al.

    Nature's TRAIL – on a path to cancer immunotherapy

    Immunity

    (2003)
  • M.J. Smyth et al.

    Granzymes: exogenous proteinases that induce target cell apoptosis

    Immunol. Today

    (1995)
  • M.J. Smyth et al.

    Purification and cloning of a novel serine protease, RNK-Met-1, from the granules of a rat natural killer cell leukemia

    J. Biol. Chem.

    (1992)
  • S.E. Street et al.

    Perforin and interferon-gamma activities independently control tumor initiation, growth, and metastasis

    Blood

    (2001)
  • K. Takeda et al.

    Involvement of tumor necrosis factor-related apoptosis-inducing ligand in NK cell-mediated and IFN-gamma-dependent suppression of subcutaneous tumor growth

    Cell. Immunol.

    (2001)
  • E. Tomasello et al.

    Gene structure, expression pattern, and biological activity of mouse killer cell activating receptor-associated protein (KARAP)/DAP-12

    J. Biol. Chem.

    (1998)
  • J.A. Trapani et al.

    Localization of granzyme B in the nucleus. A putative role in the mechanism of cytotoxic lymphocyte-mediated apoptosis

    J. Biol. Chem.

    (1996)
  • J.A. Trapani et al.

    Proapoptotic functions of cytotoxic lymphocyte granule constituents in vitro and in vivo

    Curr. Opin. Immunol.

    (2000)
  • W.M. Yokoyama et al.

    A family of murine NK cell receptors specific for target cell MHC class I molecules

    Semin. Immunol.

    (1995)
  • H. Arase et al.

    Direct recognition of cytomegalovirus by activating and inhibitory NK cell receptors

    Science

    (2002)
  • M. Azuma et al.

    Involvement of CD28 in MHC-unrestricted cytotoxicity mediated by a human natural killer leukemia cell line

    J. Immunol.

    (1992)
  • E. Baker et al.

    The genes encoding NK cell granule serine proteases, human tryptase-2 (TRYP2) and human granzyme A (HFSP), both map to chromosome 5q11–q12 and define a new locus for cytotoxic lymphocyte granule tryptases

    Immunogenetics

    (1994)
  • P.J. Beresford et al.

    Granzyme A loading induces rapid cytolysis and a novel form of DNA damage independently of caspase activation

    Immunity

    (1999)
  • C.A. Biron et al.

    Natural killer cells in antiviral defense: function and regulation by innate cytokines

    Annu. Rev. Immunol.

    (1999)
  • J. Brady et al.

    IL-21 induces the functional maturation of murine NK cells

    J. Immunol.

    (2004)
  • M.G. Brown et al.

    Vital involvement of a natural killer cell activation receptor in resistance to viral infection

    Science

    (2001)
  • K.A. Browne et al.

    Cytosolic delivery of granzyme B by bacterial toxins: evidence that endosomal disruption, in addition to transmembrane pore formation, is an important function of perforin

    Mol. Cell. Biol.

    (1999)
  • E. Carbone et al.

    A new mechanism of NK cell cytotoxicity activation: the CD40–CD40 ligand interaction

    J. Exp. Med.

    (1997)
  • A. Cerwenka et al.

    Ectopic expression of retinoic acid early inducible-1 gene (RAE-1) permits natural killer cell-mediated rejection of a MHC class I-bearing tumor in vivo

    Proc. Natl. Acad. Sci. U.S.A.

    (2001)
  • A. Cerwenka et al.

    Natural killer cells, viruses and cancer

    Nat. Rev. Immunol.

    (2001)
  • E. Cretney et al.

    Increased susceptibility to tumor initiation and metastasis in TNF-related apoptosis-inducing ligand-deficient mice

    J. Immunol.

    (2002)
  • J.E. Davis et al.

    Granzyme A and B-deficient killer lymphocytes are defective in eliciting DNA fragmentation but retain potent in vivo anti-tumor capacity

    Eur. J. Immunol.

    (2001)
  • M. Degli-Esposti

    To die or not to die – the quest of the TRAIL receptors

    J. Leukoc. Biol.

    (1999)
  • A. Diefenbach et al.

    Ligands for the murine NKG2D receptor: expression by tumor cells and activation of NK cells and macrophages

    Nat. Immunol.

    (2000)
  • A. Diefenbach et al.

    Rae1 and H60 ligands of the NKG2D receptor stimulate tumour immunity

    Nature

    (2001)
  • A. Diefenbach et al.

    Selective associations with signaling proteins determine stimulatory versus costimulatory activity of NKG2D

    Nat. Immunol.

    (2002)
  • A.O. Dokun et al.

    Specific and nonspecific NK cell activation during virus infection

    Nat. Immunol.

    (2001)
  • K. Ebnet et al.

    Granzyme A-deficient mice retain potent cell-mediated cytotoxicity

    EMBO J.

    (1995)
  • Z. Fan et al.

    Cleaving the oxidative repair protein Ape1 enhances cell death mediated by granzyme A

    Nat. Immunol.

    (2003)
  • S. Gamen et al.

    Granulysin-induced apoptosis. I. Involvement of at least two distinct pathways

    J. Immunol.

    (1998)
  • A.B. Geldhof et al.

    Expression of B7-1 by highly metastatic mouse T lymphomas induces optimal natural killer cell-mediated cytotoxicity

    Cancer Res.

    (1995)
  • T.A. Gruber et al.

    Requirement for NK cells in CD40 ligand-mediated rejection of Philadelphia chromosome-positive acute lymphoblastic leukemia cells

    J. Immunol.

    (2002)
  • W. Hashimoto et al.

    Differential antitumor effects of administration of recombinant IL-18 or recombinant IL-12 are mediated primarily by Fas–Fas ligand- and perforin-induced tumor apoptosis, respectively

    J. Immunol.

    (1999)
  • Y. Hayakawa et al.

    Cutting edge: tumor rejection mediated by NKG2D receptor–ligand interaction is dependent upon perforin

    J. Immunol.

    (2002)
  • Y. Hayakawa et al.

    NK cell TRAIL eliminates immature dendritic cells in vivo and limits dendritic cell vaccination efficacy

    J. Immunol.

    (2004)
  • A. Hutloff et al.

    ICOS is an inducible T-cell co-stimulator structurally and functionally related to CD28

    Nature

    (1999)
  • Cited by (534)

    View all citing articles on Scopus
    View full text