The existence of a macrofphage β-glucan-binding receptor had been recognized for about three decades before the identification of Dectin-1 in 2001 as the receptor for these carbohydrates (Brown and Gordon 2001). Many of the immunomodulatory properties attributed to β-glucans were previously ascribed to the Complement Receptor (CR-3, Mac-1, CD11b/CD18), which was thought to directly recognize β-glucan, among many other ligands. Although additional receptors recognizing β-glucan have been identified, such as lactosylceramide, Langerin, and the scavenger receptors CD5, CL-P1, SCARF1, and CD36, the generation of Dectin-1 knockout (KO) mice has demonstrated that β-glucan recognition by this receptor is central to the immune response to many fungal pathogens. From data generated over the past decade, it is now known that Dectin-1 has a pivotal role in the innate immune protective response to fungi, and that this receptor can induce the adaptive immune system driving the development of T helper (Th-1) and Th-17 responses. Dectin-1 has also been implicated in the recognition of Mycobacterium tuberculosis (Mtb), and recognizes an unidentified endogenous ligand. Indeed, Dectin-1 was originally identified in 2000 by Ariizumi et al. as a dendritic cell (DC)-specific molecule which was able to recognize and co-stimulate T cells, via this endogenous ligand. (See Brown 2006; Goodridge et al. 2009; Marakalala et al. 2011b for reviews.)
Structure: Dectin-1 is a type II receptor which contains an extracellular carbohydrate recognition domain (CRD), a stalk region, a transmembrane domain, and an intracellular tail containing a nonclassical immunoreceptor tyrosine activatory motif (ITAM), known as either an ITAM-like or a hemITAM. Based on CRD sequence similarity, Dectin-1 is part of the Group V C-type lectin-like family with high homology to the natural killer (NK) C-type lectin-like receptors (CLRs), and although it contains the conserved structural characteristics of these proteins, it lacks the residues normally thought to be responsible for carbohydrate binding. In fact, unlike these NK CLRs which have proteinaceous ligands, Dectin-1 binds soluble and insoluble β-(1,3)-linked glucans in a calcium-independent manner. Furthermore, Dectin-1 does not form homo- or heterodimers, and it lacks the cysteine residues within its stalk region; features often found in other NK CLRs (Brown and Gordon 2001; Ariizumi et al. 2000). Dectin-1 does, however, form higher molecular mass structures in the presence of its ligands, such as laminarin (Brown et al. 2007), and two Dectin-1 molecules are thought to be necessary for signaling via spleen tyrosine kinase (Syk) (discussed more in detail below).
Similar to the other closely related C-type lectin-like receptors in the NK gene complex, Dectin-1 is encoded by six exons. Two functional β-glucan-binding isoforms are expressed: one full-length version translated from all six exons (Dectin-1a) and one stalkless version (Dectin-1b) lacking exon 3. The human Dectin-1 has a further six alternatively spliced variants, some lacking intact exons and others with small insertions resulting in premature stop codons. Within the CRD is a shallow surface ligand-binding groove thought to be the ligand-binding site, and Trp221 and His223, which flank this groove, have been shown to be essential for binding to β-glucans. Although the endogenous ligand is still unidentified, binding of Dectin-1 to T cells was not inhibitable by β-glucans suggesting a second ligand-binding site (See Brown 2006; Marakalala et al. 2011b; Brown 2010 for reviews).
Function and Ligands
The binding efficiency of Dectin-1 depends on both the backbone chain length and degree of branching of the β-glucan polysaccharide. A variety of β-glucan and β-glucan–rich polymers are used in vitro as Dectin-1 ligands, the most frequent being the antagonists, laminarin and glucan-phosphate, the agonist curdlan, and zymosan, a complex Saccharomyces cerevisiae – derived β-glucan-rich particle that also contains multiple ligands for receptors other than Dectin-1. The purity, degree of polymerization, branching, structure, and solubility can vary extensively between these ligands and all of these factors influence the cellular responses triggered through Dectin-1. β-glucans are present in the cell wall of most fungal species, but are often only exposed during specific morphological states. Through recognition of these carbohydrates, Dectin-1 binds to a variety of fungal species including Candida, Aspergillus, Coccidioides, Pneumocystis, Saccharomyces, Trichophyton, Microsporum, and Penicillium. In addition to fungi, β-glucans are found in plants, and in some bacteria (but not Mycobacteria). As β-glucans are not produced by mammals, they act as pathogen-associated molecular patterns (PAMPs) (Brown and Gordon 2001; Adams et al. 2008; Rosas et al. 2008; Brown 2010; Kerrigan and Brown 2010).
Binding of these carbohydrates to Dectin-1 induces the production of various cytokines and chemokines, such as interleukin (IL)-1beta, IL-1alpha, IL-2, IL-6, IL-10, IL-23, macrophage inflammatory protein (MIP)-1alpha and MIP-2, tumor necrosis factor ( TNFalpha), and the production of arachidonate metabolites. Dectin-1 can also induce phagocytosis and the respiratory burst, in both neutrophils and macrophages. Stimulation of Dectin-1 is able to drive maturation of DCs and promote CD4+ and CD8+ T-cell responses to exogenous antigens, leading to the development of Th1 and Th17 adaptive immunity (see Willment and Brown 2010; Kerrigan and Brown 2010 for reviews).
Engagement of Dectin-1 with its unidentified, but non-carbohydrate, endogenous ligand induces the expression of activation markers on T cells, production of IFN-γ, and cellular proliferation. Evidence suggests that Dectin-1 may also have a role in phagocytosis of apoptotic cells and cross presentation of cellular antigens. A further endogenous ligand, the intracellular Ran-binding protein RanBPM, has been identified for the cytosolic human Dectin-1E isoform, which lacks both the transmembrane and stalk regions. RanBPM interacts with the GTPase Ran and may act as a scaffolding protein to coordinate signaling from cell surface receptors (Willment and Brown 2010).
Upon engagement of Dectin-1, the cytoplasmic ITAM-like motif becomes phosphorylated on the membrane-proximal tyrosine (Tyr 15) by Src family kinases, leading to recruitment of Syk. The exact mechanism of Syk binding is unclear, as binding normally requires two phosphorylated tyrosine residues in traditional ITAMs, although it has been suggested that Syk links two Dectin-1 molecules. Dectin-1 is able to signal through various pathways, the most well characterized to date involves Syk, caspase recruitment domain (CARD)-9, Bcl-10, and Malt1,which results in the activation of the transcription factor NFkB canonical c-Rel and p65 subunits and the non-canonical RelB subunit. The more recently identified pathway involves the kinase Raf1 which integrates with the Syk pathway by sequestering the RelB and p65 into inactive dimers and ultimately biases the cytokine production toward a Th1 and Th17 profile. Both Syk and Raf1 pathways are required for directing Dectin-1-mediated adaptive responses. The following signaling molecules are also involved in Dectin-1-mediated signaling: the Src family kinases (Lck and Src), phosphoinositide 3-kinase, mitogen-activated protein kinases (ERK and p38), and protein kinases (Akt and Jnk). The SLP76-BLNK adapter proteins are partially required, and the Tec kinase Btk is differentially used depending on the cell type examined. Dectin-1 signaling also leads to the activation of the transcription factor NFAT through PLC-γ2, acting downstream of Syk, resulting in the induction of transcription factor early growth response (Egr)2 and Egr3. NFAT activation by Dectin-1 induces cyclooxygenase (cox)-2 and regulates the production of IL-2, IL-10, and IL-12 (See Goodridge et al. 2009; Kerrigan and Brown 2010; Willment and Brown 2010; Mocsai et al. 2010; den Dunnen et al. 2010; and Kerrigan and Brown 2011 for recent reviews).
Pathogens present a diverse set of ligands or PAMPS to the immune system and therefore it is not surprising that a number of pattern recognition receptors (PPRs) are engaged simultaneously. Depending on the combination of receptors activated, the resultant response can vary significantly from a synergistic enhancement to repression of downstream responses. Triggering via Dectin-1 and MyD88-coupled Toll-like receptors (TLRs), for example, gives rise to a synergistic upregulation of MIP-2, MIP-1α, IL-10, IL-23, IL-6, and TNFalpha, but simultaneously downregulates the production of IL-12. Both Syk and Raf1 are required for the collaboration between Dectin-1 and the TLRs, and cross talk between the pathways functions to regulate the balance of Th17 and Th1 responses (Gringhuis et al. 2009).
In addition to collaborating with the TLRs, interactions with other non-TLR PPRs, have been shown to modulate responses to pathogens. For example, triggering the mannose receptor or DC-SIGN, modulates the DC responses to Mtb, and SIGN-R1 and Dectin-1 are both required for the production of reactive oxygen species (ROS) in response to Candida in macrophages (Zenaro et al. 2009; Takahara et al. 2011). It was also recently demonstrated that expression of IL-1beta in response to β-glucans in human macrophages occurs via Dectin-1/Syk and required the activation of the intracellular NRLP-3 inflammasome, the production of ROS, and a potassium efflux (Gross et al. 2009; Hise et al. 2009; Kankkunen et al. 2010).
It should be noted that the downstream responses and signaling pathways utilized by Dectin-1 are controlled by the environment within which the receptor is expressed, both in terms of the type of cell and its state of activation, and the nature of the ligand being recognized by the receptor. For example, Dectin-1 does not induce cytokine production in some macrophage populations, but it can induce cytokine production in DCs. This has been linked to differential utilization of CARD9 and effects of various cytokines, such as granulocyte macrophage-colony-stimulating factor (GM-CSF), used to generate the DCs. In the generation of Fms-related tyrosine kinase 3 ligand (Flt3L), matured DCs or bone marrow derived macrophages, both cell types, despite expressing Dectin-1, display Dectin-1–nonresponsive phenotypes. Conversely, use of GM-CSF or IFNgamma in vitro induces a Dectin-1 responsive cell type. Levels of Dectin-1 surface expression are also dependent on posttranslational modifications such as glycosylation, the formation of complexes with other PPRs, such as the TLRs and integral membrane proteins such as CD37 and CD63, and its ability to translocate to lipid rafts where it can co-localize with Syk and PLC-gamma2. The co-localization of Dectin-1 with TLR-2 requires osteopontin, an intracellular protein, which also acts as an adapter molecule facilitating interactions with Syk, increasing Dectin-1-mediated responses, such as cytokine production and facilitating the respiratory burst (Marakalala et al. 2011b; Willment and Brown 2010; Inoue et al. 2011).
Expression and Regulation
Murine Dectin-1 is present in immune cell-rich tissues such as the spleen, lung, thymus, kidney, and liver as well as other organs such as the stomach and small intestine. It is a predominantly myeloid expressed molecule with macrophages, neutrophils, CD11clow and CR-3+ splenocyte, and macrophage subpopulation of splenic red and white pulp all expressing high levels of Dectin-1. Peripheral blood monocytes, inflammatory and alveolar macrophages, Kupffer cells, microglia, lamina propria macrophages, and inflammatory-recruited neutrophils also express this receptor. Dectin-1 positive DC subsets in the peripheral blood, lymph nodes, spleen, dermis, medullary and corticomedullary regions of the thymus, and lamina propria have been identified. Low levels of Dectin-1 expression have been observed on resident peritoneal macrophages, while eosinophils, B cells, and DCs and macrophages of the kidney, heart, eye, and brain are all Dectin-1 negative. The levels of murine Dectin-1 expression can be upregulated by the presence of cytokines that bias the immune response toward a Th-2 profile, such as IL-4 and IL-13, and which gives rise to alternatively activated macrophages. While exogenous GM-CSF will increase Dectin-1 levels, the addition of IL-10, lipopolysaccharide (LPS), dexamethasone, and β-glucans will downregulate surface expression. On T-cell populations, Dectin-1 has been detected on a subset of splenic T cells and on an IL-17–producing subset of γδ T-cell receptor and chemokine receptor 6–positive T cells (See Brown 2006; Willment and Brown 2010; Reid et al. 2009 for reviews).
Human Dectin-1 is expressed on monocytes, immature and mature human monocyte-derived DCs, B cells, eosinophils, neutrophils, CD1a+ Langerhans cells, and peripheral blood DCs (CD1c+CD19−), but not on plasmacytoid DCs (although there is mRNA present in these cells) or NK cells. However, some unidentified CD4+ T-cell subsets do express this receptor. Cell-specific expression of human Dectin-1 mRNA has been reported in monocytes, macrophages, neutrophils, mast cells, eosinophils, and monocytic, B- and T-cell lines. Regulation of the transcription has been observed in vitro with the addition of various cytokines, TLR ligands, and pathogens influencing levels (Willment and Brown 2010).
Dectin-1 is a type II transmembrane receptor, however, when overexpressed in cell lines Dectin-1 localizes to both the perinuclear compartments and the cell surface. There is relatively less Dectin-1b expressed on the cell surface compared with Dectin-1a, possibly due to the lack of glycosylation signals within the former. In human neutrophils, Dectin-1 is located on the plasma membrane (γ-fraction) and within the azurophilic granules (α-fraction) inside the cytoplasm, and in murine macrophages the amounts of intracellular Dectin-1 versus the surface expressed Dectin-1 is affected by the levels of the tetraspanin CD37. The intracellular route of Dectin-1 after binding to soluble ligands, such as laminarin, allows it to be recycled to the surface, whereas larger ligands retain Dectin-1 within the phagosomal compartment leading to de novo–synthesized Dectin-1 expressed on the cell surface. After uptake of ligand, Dectin-1 co-localizes with the lysosomal membrane glycoprotein-1 (LAMP-1) and the tetraspanin CD63. Depending on the nature of the ligand, Dectin-1 can be observed in phagolysosomal compartments, during zymosan uptake, or only in early phagosomes during Candida uptake (Brown 2006; Willment and Brown 2010).
Role in Immunity and Homeostatis
The role of Dectin-1 has been extensively examined using murine models but there are now human studies, discussed more in detail below, which highlight the importance of Dectin-1 in antifungal immunity. The Dectin-1 KO mice are phenotypically normal in the absence of infection, however, they show increased susceptibility to infections with Candida (with some mouse and Candida strain variation), Aspergillus, and Pneumocystis (Drummond et al. 2011). Macrophages derived from KO animals were still able to recognize Pneumocystis and Candida albicans (strain 18804, but not strain SC5314), despite the demonstrated in vitro role of Dectin-1 in the recognition of zymosan and live fungi. Pneumocystis and Candida albicans 18804 elicited similar cytokine profiles in KO and wild-type (WT) macrophages; however, Candida albicans SC5314 recognition was impaired by the absence of Dectin-1 and the observed cytokine profile was altered. Both Pneumocystis and Aspergillus do not induce a ROS response in the KO mice, highlighting Dectin-1’s role in killing of fungi (Drummond et al. 2011). In addition to these earlier KO studies, more recent reports have reinforced Dectin-1’s role in cell-specific antifungal immunity. Mice with a macrophage-specific Dectin-1 KO, examined in the context of gastrointestinal models of candidiasis, show a more severe phenotype, and mice with a neutrophil-specific calcineurin deficiency are more susceptible to disseminated candidiasis. Dectin-1, and not the TLRs, was shown to be the activator of calcineurin in neutrophil responses to Candida (Greenblatt et al. 2010; Gales et al. 2010).
Classically the induction of a Th-1 immune response was thought to be required for antifungal immunity, but the role of Th-17 and T-regulatory cells has recently become prominent with mice lacking a Th-17 response being more susceptible to Candida infection (see Vautier et al. 2010 for review). The induction of the adaptive response, in particular the Th-17 response, is completely dependent on the presence of CARD9, but was not altered in the Dectin-1 KO mice during Candida infection (Kerrigan and Brown 2011). This induction was recently demonstrated to be mediated by Dectin-2, signaling via Fc-gamma receptor, Syk, and CARD9, which interacts with the alpha-mannans present in Candida (Saijo et al. 2010). In contrast to Candida infections, murine Dectin-1 is essential during Aspergillus infections for both the innate immune response and the induction of a Th-17 response, modulating the balance between Th-1 and Th-17 responses by inhibiting Th-1 CD4 T-cell differentiation (Werner et al. 2009; Rivera et al. 2011).
There are a few examples where a role for Dectin-1 has been demonstrated in vitro but analysis of the KO response to infectious challenge has not shown any significant role for Dectin-1 in vivo. In vitro experiments had demonstrated roles for Dectin-1 in binding and developing a Th1/Th17 profile for Mtb, whereas the in vivo experiments using the Dectin-1 KO mouse provided evidence that alternative receptors are able to compensate for the lack of Dectin-1 (Kerrigan and Brown 2011; Zenaro et al. 2009; Marakalala et al. 2011a). In the case of Cryptococcus, spores were demonstrated to bind Dectin-1 in vitro, but the KO mice are not more susceptible than WT to infection (Giles et al. 2009; Nakamura et al. 2007). Although not yet examined using Dectin-1 KO mice, susceptibility to Coccidioides infection was found to be dependent on the strain of mice, with C57BL/6 mice more susceptible than DBA/2. The differences in susceptibility may be due to the relative levels of Dectin-1a and Dectin-1b mRNA transcripts, with some mouse strains either expressing both transcripts equally or predominantly the Dectin-1b transcript. In vitro, these two isoforms have been shown to have slightly different activities, for example, cells expressing Dectin-1b produce more TNFalpha than Dectin-1a under the same conditions. There are also three murine Dectin-1 single nucleotide polymorphisms (SNPs) (R37Q, S73P, and V165A) differing between C57BL/6 and BALB/c mice that have been identified, but these SNPs have not been linked to any functional effects (Willment and Brown 2010).
A role for Dectin-1 in human antifungal immunity was recently demonstrated with the characterization of two different SNPs, which affect the levels of human Dectin-1 surface expression, and their ability to induce cytokine responses to Candida. These data demonstrate that patients with a homozygous Dectin-1 Y238X SNP, resulting in a prematurely truncated protein, are more susceptible to mucocutaneous fungal infections, but not systemic infections. Cells from these patients display defective cytokine production, in particular IL-17, TNFalpha and IL-6, but are still able to bind and phagocytose live Candida. This same SNP also has been implicated in increased susceptibility to both invasive Aspergillosis and Candida colonization during transplantation, and has clinical relevance if either the donor and/or transplant recipient of hematopoietic stem cells carries the Y238X SNP. The second characterized SNP I223S, which renders cells transduced with I223S Dectin-1 unable to bind fungal particles due to lack of surface expression, may play a role in the immunity to fungi (Marakalala et al. 2011b; Cunha et al. 2010; Plantinga et al. 2010).
The Y238X SNP patient studies have demonstrated a role for human Dectin-1 in mucosal immunity and not in systemic Candida fungal infections, due to an impaired Th-17 response, which is now thought to be pivotal during mucosal infections. This is contrary to the role in the mouse where the induction of IL-17 in response to Candida is not Dectin-1 dependent (see above). A further difference in host response is observed during Aspergillus infections; human Dectin-1 induces a strong Th-1 and a weak Th-17 response, while murine Dectin-1 inhibits the Th-1 response and induces a protective Th-17 response. These differences in immune responses may be due to site-specific immune reactions, such as mucosal versus systemic, and species diversity in the role of Th17 cells. The disparity between the human and murine immune responses to fungal infections needs further investigation, before significant progress can be made using β-glucan adjuvants targeting Dectin-1 for clinical applications (Vautier et al. 2010; Rivera et al. 2011; Kerrigan and Brown 2011; Chai et al. 2010).
Using a mouse model of zymosan-induced arthritis, a role for Dectin-1 in autoimmune disease has been suggested. The clinical relevance of Dectin-1 in human autoimmune disease is less clear as in studies of chronic inflammation, the Y238X polymorphism did not appear to be a susceptibility factor in irritable bowel diseases or rheumatoid arthritis (RA). This is despite an observed influx of Dectin-1 expressing macrophages in bowel sections obtained from both Crohn’s disease and diverticulitis patients, and synovial tissue sections from RA patients, but not those from osteoarthritis patients. It has been suggested, although not proven, that human Dectin-1 may play a role in psoriasis, firstly because Dectin-1 mRNA and those of a number of other PPRs, is upregulated in skin epidermis biopsies obtained from psoriasis patients relative to healthy controls, and secondly, the larger number of infiltrating Langerhans cells in the psoriasis samples are Dectin-1 positive (Willment and Brown 2010; Marakalala et al. 2011b).
Dectin-1 has a pivotal role in the immune response to fungal pathogens, as evidenced both from human and mouse studies. It is a predominantly myeloid restricted receptor, and its expression on these cell types is crucial for the recognition and killing of fungal pathogens and induction of the subsequent antifungal immune responses. Depending on the cell type and the nature of the ligand, it triggers intracellular signaling via its ITAM-like motif through various pathways, including Syk/CARD9 and Raf1, resulting in the activation of nuclear transcription factors. Collaborating together with multiple other PPRs, a variety of other cellular responses, such as phagocytosis, the respiratory burst, and arachidonate metabolism are initiated and the innate and adaptive immune responses are modulated through the regulation of cytokine and chemokine production. Although Dectin-1 recognizes β-glucans, it also has an uncharacterized ligand on Mycobacteria and an endogenous, non-carbohydrate ligand, on T cells and apoptotic cells. In addition to a clearly demonstrated role in antifungal immunity, Dectin-1 has been implicated in autoimmunity and may play a role in other inflammatory diseases. The study of Dectin-1, and its collaboration with other receptors, has given significant new insights into the underlying mechanisms involved in the development of the innate and adaptive immunity to fungi.
- Adams EL, Rice PJ, Graves B, Ensley HE, Yu H, Brown GD, et al. Differential high affinity interaction of Dectin-1 with natural or synthetic glucans is dependent upon primary structure and is influenced by polymer chain length and side chain branching. J Pharmacol Exp Ther. 2008;325(1):115–23.CrossRefPubMedGoogle Scholar
- Cunha C, Di Ianni M, Bozza S, Giovannini G, Zagarella S, Zelante T, et al. Dectin-1 Y238X polymorphism associates with susceptibility to invasive aspergillosis in hematopoietic transplantation through impairment of both recipient- and donor-dependent mechanisms of antifungal immunity. Blood. 2010;116(24):5394–402.CrossRefPubMedGoogle Scholar
- Takahara K, Tokieda S, Nagaoka K, Takeda T, Kimura Y, Inaba K. C-type lectin SIGNR1 enhances cellular oxidative burst response against C. albicans in cooperation with dectin-1. Eur J Immunol. 2011. doi: 10.1002/eji.200940188.