Download UvA-DARE (Digital Academic Repository) C

UvA-DARE (Digital Academic Repository)
C-type lectin signaling in dendritic cells: molecular control of antifungal inflammation
Wevers, B.A.
Link to publication
Citation for published version (APA):
Wevers, B. A. (2014). C-type lectin signaling in dendritic cells: molecular control of antifungal inflammation
General rights
It is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s),
other than for strictly personal, individual use, unless the work is under an open content license (like Creative Commons).
Disclaimer/Complaints regulations
If you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating
your reasons. In case of a legitimate complaint, the Library will make the material inaccessible and/or remove it from the website. Please Ask
the Library: http://uba.uva.nl/en/contact, or a letter to: Library of the University of Amsterdam, Secretariat, Singel 425, 1012 WP Amsterdam,
The Netherlands. You will be contacted as soon as possible.
UvA-DARE is a service provided by the library of the University of Amsterdam (http://dare.uva.nl)
Download date: 16 Jun 2017
Pa
g
hin
s
a
cr
rty
one.
16
Chapter one
one.
C-type lectin receptors orchestrate
antifungal immunity
- an introduction
Future Microbiology, 8; 839-854 (2013) – published in modified form
Brigitte A. Wevers, Teunis B.H.
Department of Experimental Immunology, Academic Medical
Geijtenbeek and Sonja I.
Center, University of Amsterdam, Amsterdam, NL.
Gringhuis
Fungal infections are an emerging threat for human
health. A coordinated host immune response is
fundamental to successful elimination of an invading
fungal microbe. A panel of C-type lectin receptors
expressed on dendritic cells enables innate recognition
of fungal cell wall carbohydrates and tailors adaptive
responses by presenting antigen as well as instruction of
CD4+ T helper cell fates. Well-balanced T helper cell type
1 and interleukin-17-producing T helper cell responses
are crucial in antifungal immunity and facilitate
phagocyte clearance of fungal encounters. Strikingly,
different classes of fungi trigger distinct sets of C-type
lectin receptors to evoke a pathogen-specific T helper
response. In this chapter we have outlined the key roles
of several C-type lectin receptors during the generation
of protective antifungal immunity, with special emphasis
on the distinct signaling pathways and transcriptional
programs triggered by these receptors, which collaborate
to orchestrate polarization of the T helper response.
C-type lectin receptors orchestrate antifungal immunity
17
FUNGAL PATHOGENS: FAR BEYOND COMMENSALISM & OPPORTUNISTIC INFECTION
Fungi are ubiquitous in the environment. Some fungi, including Malassezia species (spp.),
Candida spp., and Pneumocystis jirovecii (formerly Pneumocystis carinii) have successfully
established life-long commensal relationships with the human host, and colonize cutaneous
and mucosal surfaces without necessarily causing disease1. Even, fungal microbes are being
recognized as intestinal commensals (referred to as the mycobiome) that strongly interact
with the gut immune system2. Pathogenic fungi take advantage of an altered state of host
immunity to cause (lethal) opportunistic infections, with a rapidly growing population of
immunosuppressed patients at risk3. Commensal fungal-derived ligands (i.e. β-1,6-glucans)
can also drive chronic clonal expansion of mature B cells, and, in doing so, might contribute to the pathogenesis of B cell chronic lymphocytic leukemia (B-CLL)4. Although largely
unrecognized, this view of fungal epidemiology is dramatically challenged by the growing incidence of fungal diseases in seemingly healthy individuals. Emerging pathogenic
fungi, such as Coccidioides immitis, Histoplasma capsulatum, and Blastomyces dermatitidis,
have developed many elaborate mechanisms to overcome host immune surveillance
and establish primary and life-threatening infections1. Highly virulent Cryptococcus gattii
genotypes have gained recent prominence following a major and ongoing outbreak of fatal
cryptococcal meningitis in western North America5.
Resisting fungal infection. The human immune system is equipped with effective defense
mechanisms to mediate protection to fungal infection, yet activation of these responses
requires the coordinated activation and complex interplay of specialized types of immune
cells. Hence, host immunity has to accomplish a challenging task: maintaining tissue
homeostasis by eradicating invading fungi that can cause harm, while preventing immunopathology and tolerating the commensal fungal strains being important for our health6,7.
The human immune system comprises two arms that complement one another: innate
immunity (‘natural’ immunity) and adaptive immunity (‘acquired’ immunity). The innate
infection8. Skin and epithelial surfaces act as physical barriers of protection, and at mucosal
tissues, mucus layers and immunoglobulin A (IgA) secreted by plasma B cells work in concert
to prevent tissue invasion by fungal pathogens9-11. Innate effector cells residing in skin and
mucosa, such as interleukin (IL)-17-producing innate lymphoid cells (ILCs) and epithelial
cells, further contribute by producing antimicrobial peptides12. When fungal pathogens
successfully breach host barriers, rapidly recruited phagocytic cells, including neutrophils
and macrophages, facilitate immune protection during the earliest stages of infection and
mediate local fungal elimination13,14. Despite these effective immune mechanisms, the innate
system lacks specificity or the ability to generate immunological memory and life-long protection. These tasks are accomplished by the effector B and T lymphocyte populations from
one.
system facilitates immediate but non-specific host defense mechanisms against microbial
18
Chapter one
Box 1. Pathogen-specific lympocyte populations.
1. Pattern recognition
2. Maturation
DAMPs
PAMPs
Memory cells
3. Migration to central
lymphoid organs
CD8
IL-2
IL-12
Naive
CD8+
T cell
Dendritic cell
Mature DC
Effector
CTL
Granzyme
Perforin
IL-2
(TH1)
MHC I
Cytokines
IFN-I
CD8
Costimulation
Memory cells
B cell
IL-2
IFN-γ
MHC II
B cell
TH cell
(T-bet)
IL-12
Naive
B cell
IL-21
IL-27
TFH cell
(Bcl-6)
Naive
CD4+
T cell
IL-23
IL-6
TH17 cell
(RoRγT)
IL-4
Plasma
cell
IgG2a
IFN-γ
(TH1)
IL-4
IL-13
Phagocyte activation
IL-22
IL-17
IL-1β
B cell help
Immunity to fungi,
intracellular pathogens
Immunity to fungi
and bacteria
Mucosal homeostasis
TGF-β
TH2 cell
(GATA3)
Immunity to extracellular parasites
T reg
(Foxp3)
IL-2
TGF-β
TGF-β
IL-10
Regulation,
tolerance
IgE
Introduction
IgG1
IL-4
(TH2)
Upon delivery of their cognate antigen by DCs,
of the effector or ‘helper’ CD4+ T cell subset
in addition to receiving co-stimulatory and
family, discovered more than 25 years ago150.
cytokine signals, naive T cells in secondary
Ever since, numerous other heterogeneous
lymphoid organs become activated and
Th subsets have been characterized – the
differentiate into effector lymphocyte
three most prevalent being Th17 cells that
populations -each with specific functions and
produce IL-17151, T regulatory (Treg) cells33
gene expression programs for appropriate
and T follicular helper (Tfh) cells152. Through
elimination of different types of microbes.
actions of lineage-specific transcription
Th1 and Th2 cells are the founding members
factors, CD4+ T cells differentiate into effector
C-type lectin receptors orchestrate antifungal immunity
19
the adaptive immune system, characterized
subsets that secrete restricted patterns of
by lineage-specific effector molecules and
cytokines and express different chemokine
regulate host immunity in a pathogen-specif-
type of threat encountered153,154. Th1 cells are
dedicated to efficiently combat intracellular
bacteria and viruses, by producing IFN-γ,
ic manner (Box 1). Not surprisingly therefore,
B and T cells play a central role in providing
optimal protection to insults by fungal patho-
while Th2 cells produce IL-4 and IL-13 for
gens15,16, with the fundamental importance
defense to extracellular parasites155. The Th17
of effector CD4+ T cells dramatically exem-
cell subset, on the other hand, selectively
plified by the exceptionally high frequency
produces IL-17, providing protection to fungal
of life-threatening cryptococcal infections
and bacterial threats21. The population of
in HIV-1/AIDS patients with declined CD4+ T
Treg cells, producing cytokines TGF-β and
cell numbers17.
IL-10, plays a crucial role in the maintenance
of immune homeostasis156. Tfh cells represent
another effector CD4+ T cell population
with a specialized function: Tfh cells help
B cells generate antibody responses to T
Two functionally distinct CD4+ T cell
subsets are considered key to effective fungal microbe elimination: T helper cell type
1 (Th1) and IL-17-producing T helper (Th17)
cell-dependent antigens for clearance of
cells. Th1 and Th17 responses can be induced
pathogens by phagocytes or the complement
in parallel, yet, although still considerably
system . Effector CD8 T cells, cytotoxic T
uncertain, their degree of contribution is pre-
cells -in contrast to helper CD4+ T cells not
sumably context dependent, e.g. pathogen-
further subdivided- produce multiple effector
and/or tissue-specific18: Th1 cells take part
molecules, such as perforin and granzyme,
in the cellular defense, important during dis-
and are specialized in destroying virally
seminated disease, and orchestrate optimal
157
+
infected cells and/or tumor cells158. About
10% of the effector CD4+ and CD8+ T cells
acquire a memory phenotype: quiescent
long-lived central memory T cell (Tcm) and
macrophage activation, whereas Th17 cells
predominantly maintain barrier immunity
at mucosal surfaces and act on neutrophils.
effector memory T cell (Tem) populations
These effector T cells secrete cytokines to
that are able to quickly respond to antigen
mediate their influence on other immune
re-exposure . Memory B cell generation,
cells during an antifungal immune response.
in addition to isotype switching to IgG, IgA
Th1 cells secrete interferon-γ (IFN-γ), which
and IgE subtypes, depends on cytokine
triggers a plethora of systemic effector mech-
present in the local environment (i.e. Th2-
anisms, such as antibody class switching to
derived IL-4 induces IgE class-switching)160.
opsonizing subtypes, upregulation of MHC
159
Although Th cells have been considered
terminally differentiated immune cells, this
view is considerably challenged in recent
molecules for enhanced antigen presentation, and stimulation of macrophage effector
years, with many examples of Th cells
functions (e.g. production reactive oxygen
flexible in their cytokine production profile,
intermediates)19. IFN-γ might also directly
and hence their effector phenotype153.
affect fungal growth, as it inhibits the yeastto-hyphal transition in C. albicans20. The Th17
one.
receptors, ensuring tailored responses to the
20
Chapter one
effector molecule is an important mediator of tissue inflammation: IL-17 acts on a broad range
of immune and non-immune cells and is key to the recruitment, migration and activation
of neutrophils21. Th17 cells are also an important source for IL-22, which promotes, together
with IL-17, production of protective antimicrobial peptides to mediate mucosal microbial
resistance22,23 (Box 1). The protective role of Th17 responses during fungal infection is underscored by the severe recurrent and chronic Candida infections in patients with genetic defects
in the Th17 axis, including individuals suffering from chronic granulomatous disease (CGD)
and hyper IgE syndrome24-30. Similarly, Th17 cells and IL-17 have been shown to mediate
protection in numerous experimental mouse models of fungal infection31,32. Paradoxically,
exaggerated antimicrobial Th17 responses are often associated with tissue damage. The
magnitude of pathogenic Th17 cell activity as well as unwanted Th17 responses directed
against commensal fungi can be kept in check by regulatory T (Treg) cells, the natural
gatekeepers of immune homeostasis33. In any case, finely tuned Th1 and Th17 responses
probably maximize fungal elimination and, at the same time, minimize host tissue damage
during inflammation (recent reviews on this topic have been published elsewhere:16,34,35).
Instruction of the adaptive (antifungal) response is coordinated through the actions of
specialized antigen-presenting cells (APCs), principally dendritic cells (DCs), which provide
all signals necessary for naive T cells to acquire an effector phenotype: T cell receptor (TCR)
stimulation, co-stimulation and cytokines36, thereby contributing to T cell-dependent B cell
help and generation of antibody responses.
DENDRITIC CELLS AND THE GENERATION OF ANTIFUNGAL IMMUNITY
DCs reside in the periphery or circulate through blood and monitor for signs of microbial
attack, but also host derived danger signals released in response to stress, tissue damage
and necrotic cell death. Being specialized in sensing conserved microbial structures termed
pathogen-associated molecular patterns (PAMPs) and damage-associated molecular
patterns (DAMPs), through pattern recognition receptors (PRRs), DCs can discriminate
between different classes of potential danger37,38. PAMPs refer to molecules associated
with pathogenic and non-pathogenic microbes, such as cell wall components and nucleic
acids of fungal, bacterial and viral origin, while DAMPs are endogenous molecules released
upon stress or damage to the host: amongst others high-mobility group protein 1 (HMGB1),
heat-shock proteins (HSPs), extracellular ATP and uric acid crystals. Immature DCs capture
Introduction
and internalize pathogens or self-molecules, simultaneous encounter with PAMPs/DAMPs
induces a cascade of phenotypical changes. This so-called maturation process entails
upregulation of costimulatory molecules, lymphoid tissue homing receptor CCR7, and major
histocompatibility complex (MHC)-antigen complexes, allowing DCs to activate naive T cells
in central lymphoid organs. Responding T cells start to proliferate and differentiate, and, as
C-type lectin receptors orchestrate antifungal immunity
21
distinct effector populations, rapidly enter sites of local inflammation were they perform
their effector function to aid in pathogen-specific clearance (Box 1).
Pattern recognition receptors. Instruction of the adaptive response by DCs is subject to
tight regulation, and this process is dependent on a large panel of germ-line encoded
PRRs39. DCs express a large variety of membrane- and cytoplasmic-localized PRRs,
including the archetypical Toll-like receptors (TLRs) and C-type lectin receptors (CLRs)
and NOD-like receptors (NLRs) (further discussed in Box 2). To coordinate DC-induced
inflammatory responses, PRRs control four crucial processes. First, PRRs allow DCs
to discriminate between different classes of PAMPs and DAMPs, and as such ‘license’
them to drive pathogen-specific responses37. Moreover, DC-expressed PRRs facilitate
internalization and processing of pathogen-derived antigens for subsequent antigen
presentation in the context of MHC molecules. In addition, a selective set of PRRs induces intracellular signaling for activation of two additional processes: transcriptional
activation of a core set of innate response genes, leading to expression of co-stimulatory molecules, chemokines and cytokines40,41; and the assembly of cytosolic protein
complexes, inflammasomes, for posttranslational processing of IL-1β family members42
(Figure 1). Since the local cytokine milieu created by dendritic cells is instrumental to
the fate lineage decision of differentiating CD4+ T helper cells36,43, PRR-induced signaling is crucial for clonal expansion and differentiation of a responding antigen-primed
T cell population.
Fundamental to the expression of many inflammatory cytokine and chemokine genes
is the activation of the nuclear factor-κ B (NF-κB) family of transcription factors, which are
designated as central coordinators of the innate immune response. NF-κB homo- and
heterodimers are retained inactive within the cytoplasm by inhibitory proteins of the IκB
family. Upon a PRR-mediated signal, the IκB inhibitory complex is degraded, and subsequently initiates release and nuclear translocation of NF-κB dimers44. In addition, PRRs also
activate other transcription factors, such as transcription factor activator protein-1 (AP-1)
factors (IRFs) for induction of type I interferon (IFN-I) responses.
Tailoring T helper responses to fungal infection. The local cytokine milieu created by DCs
is instrumental to the fate lineage decision of differentiating CD4+ T helper cells36,43; cytokine actions involve direct induction or repression of a lineage-specific transcription
factor or essential growth factor(s). Regarding activation and maintenance of human
antifungal T h1 and T h17 effector subsets, several cytokines are considered of crucial
importance. T h1 cells differentiate from naive T cells in response to DC-derived IL-1245.
IL-12 binding to its cognate receptor (IL12R) on activated CD4+ T cells triggers, via STAT4,
transcription of the lineage-specific transcription factor T-bet46, which mediates the
one.
for expression of cytokines and chemokines, as well as numerous interferon regulatory
22
Chapter one
i Pattern recognition
Flagellin
LPS
TLR5
TLR4
ii Antigen presentation
Mannose
CLRs
RLRs
ds/ssRNA
Phagosytosis
MHC I
Phagosome
NLRs
MDP
RIG-I MDA5
Surface
NOD2 NLRP3
DAMPs
Endosome
PAMPs
Viruses
Host cell death
Necrosis
Sap130
MHC II
Dectin-1
Dectin-2
TLRs
dsRNA
Lysosome
Endogenous
proteins
TLR9 TLR7/TLR8
CpG
DNA
ssRNA
Endosome
dsDNA
IFI16
MHC II
loading
Fungi
Bacteria
TLR3
Mincle
AIM2
β-glucan
PYHINs
MHC I MHC II
Proteasome
MHC I
loading
Surface
ER
Golgi
Figure 1. Four principal roles of pattern recognition receptors (PRRs). (i) Innate immune cell-associated
PRRs recognize distinct types of pathogen associated molecular patterns (PAMPs) or damage-associated
molecular patterns (DAMPs), allowing instruction of tailored adaptive immune responses. Numerous
PRR families have been characterized; while some are stationed at cell membranes, such as Toll-like
receptors (TLRs) and C-type lectin receptors (CLRs), others are located within the cytosol. RIG-I-like
receptors (RLRs), NOD-like receptors (NLRs) and PYHIN sensors. Prominent PRR family members and
their cognate ligands are depicted in the figure. (ii) PRRs facilitate internalization and/or processing of
peptide-derived antigens for presentation in the context of major histocompatibility (MHC) class I and II I
secretion of IFN-γ. Human T h17 cell fate determination involves multiple cytokines: IL-6,
IL-23, IL-1β, IL-21 and TGF-β21, although debate continues regarding requirement and
primary source of the latter. T h17 development is dependent on transcription factors
Introduction
STAT3 and RORγt. Signaling by DC-derived IL-6 and IL-23 directly activates STAT3
and subsequently RORγt. IL-21, another STAT3 activator, is expressed by T h17 cell and
promotes maintenance of T h17 differentiation via an autocrine route. IL-1β functions
during early and late stages of T h17 cell commitment, possibly by counteracting the
inhibitory effects of IL-12 and IL-10 on T h17 differentiation47. T h17 cells produce the
23
C-type lectin receptors orchestrate antifungal immunity
iii Gene transcription
iv Inflammasome activation
TLR4
Dectin-1
Dectin-1
CD14
TRIF
K+ efflux
Myd88
ROS
Stress
signals
IRAK
Syk
TLR3
Syk
PAMPs
DAMPs
C/B/M
scaffold
Mytochondria
Canonical
TRIF
NLRP3
Non-cannonical
ASC
ASC
Pro-caspase-8
RIG-I
MAVS
RIP2
NOD1/2
Pro-caspase-1
MDA5
TBK1
IRFs
Type I IFNs
TRAFs
TAK1
NF-kB AP1
Chemokines
Cytokines
Antimicrobial peptides
Costimulatory molecules
Caspase-1
IL-1β
Processing
IL-18
Caspase-8
Cytokine
release
Pro-IL-1β
Pro-IL-18
 molecules to naive CD4+ and CD8+ T cells, respectively. (iii) Several PRRs transduce intracellular
signaling upon their activation, leading to transcriptional activation of numerous innate response
genes. TRAF adaptor proteins account for integration and diversification of PRR signaling for activation
of different transcription factors. (iv) Furthermore, PRRs can mediate the assembly and activation of
cytosolic protein complexes -caspase-1-containing canonical or caspase-8-containing non-canonical
inflammasomes- for posttranslational processing and maturation of cytokines from the IL-1β family.
C/B/M, CARD9-Bcl-10-MALT1; dsDNA/RNA, double-stranded DNA/RNA; MDP, muramyl dipeptide; IFNs,
signature cytokines IL-17A (referred to as IL-17), IL-17F and IL-2221 (Box 1, with Figure
1 in Chapter 6 providing a more comprehensive overview). Foxp3-expressing Tregs
restrain uncontrolled chronic T h1 and T h17 effector responses deleterious to the host
and exist as a mature T cell subpopulation in the periphery (natural (n)Tregs), but can
also be induced from naive CD4+ T cells by IL-2. Induced regulatory T cells (iTregs)
acquire suppressive activity in response to transforming growth factor-β (TGF-β)48.
Thus, DCs are masters in command of an army of lymphocytes and hence shape the
adaptive arm of an ensuing antifungal inflammatory response. Depending on the
one.
interferons; ROS, reactive oxygen species; ss-RNA, single-stranded RNA.
24
Chapter one
fungal species encountered as well as the host cell type, specific PRRs will be activated,
which elicit distinct downstream signaling events that collectively determine the overall
adaptive response tailored to the encountered microbes.
SENSING FUNGAL INVASION: C-TYPE LECTIN RECEPTORS TAKE CENTER STAGE
The fungal cell wall composition is dynamic and highly variable, yet consists of a multitude
of putative and unique PRR ligands49. The core structure is dominated by polysaccharides,
comprising mainly β-1,3-glucan, β-1,6-glucan and chitin polymers, surrounded by a layer
enriched in mannosylated glycoproteins50. Phospholipomannan, α-glucans, and galactomannan constitute cell wall components found in only a minority of fungi51. Fungal
components can be recognized by more than one receptor (e.g. β-glucan recognition by
both langerin and dectin-1), resulting in differential responses, and certain PRRs transduce
divergent intracellular signaling pathways upon binding distinct ligands; exemplified by
the mannose- and fucose-based signaling induced by DC-specific ICAM-3-grabbing non-integrin (DC-SIGN) with pro- or anti-inflammatory outcomes, respectively (further discussed
below). Notably, as we demonstrate in Chapter 2 of this thesis, even closely related strains
within one taxonomic group can differentially trigger innate receptors on dendritic cells52.
In sum, expression of a plethora of innate PRRs permits the host immune system to mount
an effective, and above-all, tailored antifungal adaptive response.
TLRs, among the most well characterized PRRs, have been assigned function in the anti-
fungal immune response. Fatal Aspergillus fumigatus infections observed in Toll-deficient
Drosophila provided an initial link between TLR components and antifungal immunity53. In
mammalian studies with murine infection models, TLRs were found to have critical roles in
both innate recognition and driving protective responses54,55. Strikingly, however, the control
of antifungal defense in men is not dominated by any of the TLR members. Humans with
genetic defects in the universal TLR adaptor molecule -shared as well by the IL-1 receptor
(IL-1R) and IL-18R- MyD88 are highly susceptible to bacterial, but not fungal infections56,57.
Strictly under conditions of severe immunosuppression, single nucleotide polymorphisms
(SNPs) in human TLR1 and TLR4 genes predispose to infection with fungi57,58. Also, human
TLRs are considered incapable to autonomously elicit robust Th17 skewing52. During fungal
infection, engagement of most TLRs potentiates strong IL-12p70 production, and thus favors
Th1 polarization16,59, although Treg activation by TLR2 is considered an exception60. Murine
Introduction
TLRs exhibit some potential to augment Th17 responses61, possibly reflected by their ability
to activate transcription factor NF-κB subunit c-Rel (discussed below) in some instances, in
marked contrast to their counterparts in men52,62. Thus, at least in the human setting, TLRs
cannot be held solely responsible for the control of fungal elimination. Presumably, TLRs
contribute to fungal binding and operate as co-stimulators that promote or repress signals
C-type lectin receptors orchestrate antifungal immunity
25
by other PRRs to shape the overall antifungal response35.
Emerging evidence indicates that the control of human antifungal defense is instead
dominated by C-type lectin receptors (CLRs; Box 2). Langerin and mannose receptor (MR) are
CLR sensors for fungi with endocytic activity and facilitate or, in the case of MR, contribute
to phagocytosis of fungal particles63-66. These receptors subsequently direct delivery of the
fungal cargo into the appropriate phagosomal route; internalized fungi are either processed
for antigen presentation, intracellular NLRP3/caspase-1 inflammasome activation, or, alternatively, for degradation in an attempt to clear the fungal threat. It is becoming evident that
several myeloid CLRs act as PRRs that exhibit potential to transduce intracellular signaling
to direct transcription of innate response genes40. Among these signaling CLRs, several have
been implicated in antifungal immunity, all with distinct mechanisms of action: dectin-1,
dectin-2, mincle, and DC-SIGN67-70. Whether langerin and MR, besides promoting fungal
uptake and processing, transduce intracellular signaling has not been formally proven.
It is, however, likely that both CLRs modulate intracellular signaling indirectly, simply by
influencing recruitment of signaling receptors to the phagocytic synapse71.
It is of particular interest that CLR signaling through downstream assembly of a complex
containing CARD9 is indispensable for the generation of a Th17-dominated response52,72,73,
and additionally induces Th1 polarization72,74. Dectin-1 represents the prototype antifungal
CLR, which renders DCs fully competent to direct Th1 and Th17 immunity after exposure
to fungal β-glucan72,74. Recent genetic studies in humans signify the importance of the
dectin-1/CARD9 axis, as signaling defects have been connected to susceptibility to fungal
infection. A SNP in dectin-1 (Y238X) that introduces a premature stop codon and prevents
functional dectin-1 expression, predisposes to chronic mucocutaneous candidiasis (CMC),
due to limited production of IL-17 and low numbers of Th17 cells in peripheral blood75.
Even, in patients receiving immunosuppressive medication, the dectin-1 Y238X SNP has
been associated with enhanced susceptibility to invasive aspergillosis76-78. Patients with a
loss-of-function CARD9 mutation similarly display greatly reduced numbers of circulating
Th17 cells and CMC, but with far more severe clinical symptoms and manifestations of lethal
models: deficiency of CARD9 or its upstream effector PKC-δ, results in invasive infection
and rapid lethality, rather than the lack of dectin-1 alone72,81. In sum, these studies strongly
suggest redundancy between CARD9-coupled receptors, whereas CARD9 dysfunction is
detrimental for host control of fungal infection.
Apart from dectin-1, CLRs dectin-2 and mincle also transmit signals via the CARD9
module73,82. Indeed, dectin-1 is thought to be dispensable for protection of mice against
infection with certain subtypes of fungi, with induction of Th17 responses to systemic C.
albicans predominated by dectin-2, not dectin-173,83, and likewise, several studies have suggested contribution of mincle to establishment of protective immunity in mice69,84,85. The
studies described in this thesis aimed at the functional characterization of both dectin-2
one.
systemic disease79,80. This discrepancy has been observed likewise in murine infection
26
Chapter one
Box 2. Pattern Recognition Receptors.
The innate immune system senses presence of
and expression of genes encoding cytokines,
potential danger via recognition of molecular
chemokines and antimicrobial peptides162.
structures unique to different types of microbes
Expression of TLRs is cell-type specific.
(PAMPs) or damaged-self (DAMPs), through
pattern recognition receptors (PRRs)39. Different
The C-type lectin receptor (CLR) superfamily
PRRs recognize different types of PAMPs,
is a large group of proteins characterized
while a given pathogen or self-molecule can be
by the presence of one or more C-type
recognized by multiple PRRs simultaneously.
lectin-like domain(s) (CTLDs)163. CLRs
This allows PRRs -in specific combinations- to
primarily sense carbohydrate moieties
instruct the adaptive immune system how to
such as mannose and fucose (e.g. by DC-
respond best: providing information about
SIGN) as well as β-glucan (i.e. by dectin-1) on
the initiation, type, duration and magnitude
pathogens and host-derived glycoproteins,
of the response . Hence, PRRs dictate the
but some recognize F-actin filaments (i.e.
outcome of an ensuing immune response.
DNGR-1)164,165, and ribonucleoproteins82
Numerous classes of PRRs have been identified,
released by necrotic host cells (i.e. mincle).
functioning in distinct extra- and intracellular
CLRs are implicated in cell adhesion and
compartments but also cell types. Most
communication processes, detection of cell
prominent PRR families are listed below, along
death, and uptake of (altered-) self and non-
with some of their best-studied members:
self-antigens145,166. In addition, several CLRs are
161
able to transduce Syk-dependent signaling,
Membrane bound receptors.
thereby predominating the antifungal immune
Toll-like receptors (TLRs), amongst the best-
response. This occurs directly through an
studied PRRs, are glycosylated type I membrane
intracellular ITAM-like domain (i.e. dectin-1),
proteins, comprising a leucine-rich repeat
or indirectly via association with an ITAM-
(LRR) ectodomain for recognition of well-
containing adaptor molecule (e.g. dectin-2
defined PAMPs. The 10 known human TLRs
and mincle)40, resulting in induction and/or
are stationed at the cell surface for sensing
modulation of cytokine and type I interferon
(myco)bacterial, fungal and parasitic PAMPs
(IFN) responses via transcription factors
(e.g. lipopolysaccharide from gram-negative
NF-κB, IRF1 and IRF5 (findings described
bacteria by TLR4 and bacterial lipoproteins
in Chapters 2, 3, 4 and 5 of this thesis).
Introduction
by TLR1/TLR2 and TLR2/TLR6 complexes),
or signal from intracellular vesicles upon
Cytoplasmic sensors.
detection of double-stranded (ds)RNA (TLR3),
NOD-like receptors (NLRs) constitutes the
single-stranded microbial (ss)RNA (TLR7) or
largest family of the cytoplasmic localized
CpG-rich methylated microbial DNA (TLR9).
PRRs. Similar to the TLR family, NLRs contain
TLRs recruit a single or distinct set of Toll-IL-1
LRR motifs for detection of wide variety of
receptor TIR-domain-containing adaptor
PAMPs and DAMPs, while, in contrast to TLRs,
molecules, such as Myd88, TRIF, TIRAP
they are expressed by a wide variety of cell
and TRAM, to their cytosolic domains for
types. A central nucleotide-binding domain
induction of downstream signaling events
controls NLR oligomerization, while their
C-type lectin receptors orchestrate antifungal immunity
27
caspase-recruitment (CARD), pyrin (PYD) and
dependent NF-κB activation, but also mediate
baculovirus-inhibitory repeat (BIR) domains
NLRP3 inflammasome assembly (RIG-I). The
facilitate intracellular signal transduction.
third RLR family member, LGP2, lacks a CARD
The NLR family comprises more than 20
motif for type I IFN signaling and is thought to
members, which can be further divided into
function as a modulator of RIG-I and MDA5168.
NLRPs (previously NALPs) and NODs (also
RLR family members are well known for their
known as NLRCs). NLRPs are well-known
ability to crosstalk with TLRs and other PRRs
for their ability to sense viral infection, upon
for modulation of adaptive immune responses;
which they assemble into cytoplasmic
aberrant or dysregulated RLR signaling has
protein complexes (inflammasomes) together
been linked to development of autoimmunity.
with ASC and caspase-1 for maturation
of IL-1β cytokines. NLRP3 assembles into
The PYHIN protein family is a group of IFN-
inflammasome complexes upon indirect
inducible proteins, sensing cytosolic dsDNA
sensing of a wide variety of microbial and
from viruses and bacteria through their PYD
endogenous stress signals (i.e. RNA viruses,
and/or HIN200 domain(s). AIM2 is a PYHIN
bacteria and mitochondria-derived stress
member with established PRR function,
signals: mROS, mtDNA and cadiolipin),
mediating inflammasome assembly but
presumably via detecting a commonly induced
not gene transcription, while numerous
K+ efflux. NLRC4 assembles inflammasomes
PYHINs, including IFI16, are putative dsDNA
upon recognition of bacterial flagellin and a
sensors signaling for STING-dependent
type III and IV secretion system components .
167
induction of cytokines and type I IFNs169.
Despite containing a CARD-domain, NOD1 and
NOD2 are non-inflammasome forming NLRs;
NOD1 and NOD2 sense bacterial peptidoglycan
motifs and as oligomers associate with
adaptor RIP2 for induction of signaling leading
to expression of cytokines, chemokines
and reactive oxygen species (ROS)42.
and MDA5, are a family of DExD/H box RNA
helicases, sensing ssRNA and dsRNA from viral
origin or processed-self within the cytoplasmic
compartment. RIG-I is also capable of sensing
dsDNA indirectly, after it has been processed
into ssRNA structures via RNA polymerase
III (RNAP III). RIG-I and MDA5 signal via an
IPS-1 signalosome for induction of type I IFN
responses via transcription factors IRF1, IRF3
and IRF7 or cytokines via CARD9/Bcl-10-
one.
RIG-I-like receptors (RLRs), including RIG-I
28
Chapter one
and mincle in human antifungal immunity, and in accordance with the above notions, we
provide evidence for key roles for both dectin-2 (Chapter 2) and mincle (Chapters 3, 4 and 5).
DC-SIGN signals independent of the CARD9 axis, yet binds several fungi and dynamically
regulates assembly of its signalosome to amplify or inhibit T helper polarization86. Thus,
multiple CLRs (i.e. dectin-1, DC-SIGN, dectin-2 and mincle) signal collaboratively to yield
the most optimal antifungal response; the mechanisms of which are now beginning to be
elucidated at the molecular level.
The remainder of this chapter focuses on the distinct signaling pathways and transcriptional
programs by which CLRs dectin-1, DC-SIGN, dectin-2 and mincle can influence the adaptive
outcome of an antifungal response. We explore the hypothesis that, in terms of effector
mechanisms, signaling CLRs implicated in human antifungal immunity can be classified
into two distinct groups: (i) receptors that act individually and provide all transcriptional
signals required to bridge innate and adaptive responses -that is by instructing CD4+ T
helper cell polarization, and (ii) receptors that respond cooperatively and modulate signals
from other PRRs to fine-tune a particular adaptive response. Dectin-1 is the prototype of
an inducer, specialized in directing both Th1 and Th17 cell responses, whereas DC-SIGN
is a prominent member of the second class that distinctively modulates Th1 responses. In
fact, our current studies (described in Chapters 2, 3, 4 and 5 of this thesis) reveal that CLRs
dectin-2 and mincle can also be classified as modulating CLRs.
DECTIN-1: THE CORNERSTONE OF HUMAN ANTIFUNGAL IMMUNITY
Dectin-1, the central paradigm for a signaling CLR, is expressed primarily in cells from
myeloid origin, including DCs, macrophages, monocytes, neutrophils, langerhans cells
(LCs) and eosinophils, yet also found on B cells and mouse innate γδ T cells63,87,88. By means
of its β-1,3-glucan and β-1,6-glucan carbohydrate specificity, dectin-1 is capable of binding
most if not all fungi, due to the abundance of β-glucans (polymers of D-glucose linked by
β-glucosidic bonds) in nearly all fungal cell walls50. Ligand binding by dectin-1 occurs in a
Ca2+-independent manner, which is divergent from most other CLRs. Studies in both human
and mice have documented recognition of numerous pathogenic species by dectin-1,
including Aspergillus spp., Candida spp., Coccidiodides spp., capsule-deficient C. neoformans,
Fonsecaea pedrosoi, H. capsulatum, and P. jirovecii85,89-95. Also, dectin-1 interacts with myco-
Introduction
bacteria, albeit via recognition of a yet unknown ligand96,97.
Not surprisingly, fungal cell wall β-glucan abundance influences initial innate detec-
tion by dectin-1. The opportunistic pathogen C. albicans has a dimorphic appearance, and
β-glucans become more accessible during transition from the commensal yeast form into
the invasive filamentous form98. Although the ability to undergo phase transition is strong-
C-type lectin receptors orchestrate antifungal immunity
29
ly associated with fungal pathogenicity98, the host might use this event to discriminate
invasion from colonization and provoke an antifungal inflammatory response99. Dectin-1
presumably is an important player in local tissue immunosurveillance, with commensal
fungi being important constituents of the host skin, oral, and gut microbiota2. Interestingly, a
dectin-1 gene variant has been associated with aggravation of inflammatory bowel disease
(IBD) severity2, suggesting that altered sensing of fungi by dectin-1 contributes to aberrant
immune responses in IBD. Nevertheless, some pathogenic fungal strains strategically mask
their β-1,3-glucans to prevent immune recognition, even phagocytosis, and succeed in gaining virulence. The immunologically inert capsule of Cryptocuccos spp. and hydrophobic,
RodA-rich, layer of Aspergillus conidia are considered most extreme examples100,101.
In myeloid cells, dectin-1 transduces downstream signaling via a unique intracellular
signaling domain, which delivers activation signals to Src and Syk family kinases. This
domain resembles an immunoreceptor tyrosine-based activation motif (ITAM), termed
hemITAM, but differs from a conventional ITAM in that it possesses only one of the two Tyrx-x-Leu (YxxL) sequences102. Binding of the tandem Src homology domain 2 (SH2) domains
of Syk to dually phosphorylated ITAMs is crucial for Syk activation. Because of its unusual
ITAM, dectin-1 is thought to dimerize to provide such a docking site103,104, analogous to the
hemITAM-containing receptor Clec-2105. Syk undergoes autophosphorylation at numerous
tyrosines upon binding to dectin-1 in order to initiate downstream signal transduction106
(Figure 2). Two related membrane-associated tyrosine phosphatases (i.e. CD45 and CD148)
mediate Src family kinase activation, but need to be quickly segregated from the dectin-1
synapse to avoid dephosphorylation of the ITAM tyrosine residues, and permit productive
signaling107. Notably, soluble, β-glucan polymers have been found incapable of excluding
CD45 and CD148 activity, even though they bind dectin-1107. This may ensure that dectin-1
signaling is activated solely upon encountering an invading fungus, which should be eliminated, and not harmless shed β-1,3-glucan fragments.
Dectin-1 mediates phagocytic uptake of fungal particles107, directs fungal destruction
(through production of toxic reactive oxygen intermediates) by macrophages, neutrophils
of neutrophils (e.g. degranulation)109. In addition to these immediate antimicrobial effector
responses, dectin-1 signaling promotes efficient MHC class II presentation of fungal-derived
antigens to CD4+ T cells110, and activates nuclear translocation of transcription factor NF-κB
to mediate release of innate response mediators that shape the overall adaptive response.
Syk-dependent signaling. In human DCs, dectin-1 autonomously orchestrates activation
of all NF-κB subunits, through activation of both the classical and noncanonical NF-κB
pathways, and accordingly expression of T h1 and T h17 polarizing cytokines52, via
induction of two independent signaling pathways. Syk-dependent signaling induces
assembly of a trimolecular signaling complex consisting of CARD9, Bcl-10 and MALT1111.
one.
and DCs108, and controls, via Ca2+-dependent NFAT transcription, the microbicidal activity
30
Chapter one
Inducing receptor
Modulating receptor
Mannose
β-glucan
Fungi
Dectin-1
Fucose
DC-SIGN
ROS
(+ Signal 2)
K63
SYK
1
CARD9
Raf-1
NLRP3
ASC
Casp 1
Inflammasomes
Caspase 8
CARD9
p100
NIK
Raf-1
p p
p
p
MALT1 Bcl-10
Caspase 1
CNK1
p
Nucleus
p p
Raf-1
1. Enhanced
p65 activity
TAK1
LSP1
2
Ras
CN
K
Ra 1
f-1
KS
R1
ITAM-like motif y y
p p
KSR1
LSP1
Virus
2. Reduced
transcription
IκBα Canonical
RelB
NF-κB
Non canonical
MALT1 Bcl-10
Casp 8
C
AS
p52 RelB
inactivation
p
a p
p65 RelB
p65 p50
c-Rel
IL-1β
IL23A
NF-κB
p50
IL1B
IL6
TH17 promoting
IL12A
IL12B
TH1 promoting
pro-IL-1β
Figure 2. Dectin-1 provides all transcriptional signals to generate Th1- and Th17- polarizing cytokine
profiles, while signaling induced by DC-SIGN modulates CD4+ T cell responses. (a) Upon β-glucan sensing, dectin-1 activates two independent signaling cascades that integrate at the level of nuclear factor-kB
(NF-κB) activation: Syk- and Raf-1-based pathways. Recruitment of Syk to the phosphorylated (P) dual
tyrosine (Y) motifs of dectin-1 facilitates, via an intermediate kinase (possibly PKCδ; not shown), the assembly of a complex consisting of CARD9, Bcl-10 and MALT1. This CARD9/Bcl-10/MALT1 scaffold then
presumably undergoes non-degenerative Lys63 (K63)-linked poly-ubiqitination (polyUb)170, which can
Introduction
be recognized by cofactors such as TAK1 and TRAF proteins. This leads to the activation of IKK subunit β
(IKKβ; not shown), which phosphorylates the NF-κB inhibitor protein IkBα, and targets it for proteasomal
degradation. Following IkBα degradation, canonical NF-κB subunits (depicted as p65-p50 and c-Rel-p50
dimers) can enter the nucleus, to drive expression of, among other inflammatory mediators, IL-6, IL-1β​
and IL-23, which induce Th17, and the Th1 polarizing cytokine IL-12p70. Syk-dependent signaling also I
C-type lectin receptors orchestrate antifungal immunity
31
 initiates an alternative or non-canonical NF-κB pathway that culminates in the activation of RelB: IKKα
(not depicted) is activated by NIK, and initiates processing of NF-κB inhibitor p100 into p52, which enters
the nucleus together with RelB. CARD9 and Bcl-10 are involved in recruitment of all NF-κB subunits,
whereas the paracaspase MALT1, through its proteolytic activity, strictly targets c-Rel. Besides initiation
of canonical and non-canonical NF-κB signaling (1), Syk mediates generation of reactive oxygen species
(ROS) production for NLRP3/caspase-1 inflammasome activation (2). Raf-1-dependent signaling culminates in phosphorylation at serine 276 (Ser276) and acetylation (a) of p65. Phopshorylated p65 restrains
RelB activity, while acetylated p65 has prolonged nuclear activity and enhances transcription of IL6 and
IL12 genes. The CARD9/Bcl-10/MALT1 scaffold also initiates formation of an alternative caspase-8 inflammasome in which the CARD9/Bcl-10/MALT1 triad is linked to caspase-8 and the adaptor protein ASC for
proteolytic processing of pro-IL-1β. (b) DC-SIGN signaling is ligand-specific and affects Th1 polarization.
(left) A pre-assembled trimeric complex consisting of KSR1, CNK and Raf-1 is, via the adaptor molecule
LSP1, constitutively associated with the cytoplasmic tail of DC-SIGN. Binding of mannose-containing
pathogens (such as fungi) to DC-SIGN induces activation of the serine/threonine kinase Raf-1, which
in turn mediates phosphorylation and acetylation of NF-κB subunit p65. These modifications prolong
the nuclear activity of p65, resulting in increased transcription rates at specific genes, including those
encoding IL-12p70, crucial for antifungal Th1 polarization. (right) Fucose-containing pathogens trigger
an alternative, and LSP1-dependent, pathway, accompanied by disassembly of the KSR1/CNK/Raf-1 triad.
Activation of this cascade attenuates pro-inflammatory cytokine production, thus negatively affects Th1
responses, via a yet unknown mechanism. Please note that the example given here concerns dectin-1
as an inducer of antifungal Th1 and Th17 responses, but DC-SIGN can influence antifungal responses
induced by any other innate receptor.
Downstream intermediate PKCδ is likely to couple Syk activity directly to CARD9
phosphorylation and recruitment81. The CARD9/Bcl-10/MALT1 scaffold subsequently
activates oligomerization of the IκB kinase (IKK) complex to allow nuclear translocation
of canonical NF-κB subunits p65 and c-Rel111. It is not entirely clear how the CARD9/
Bcl-10/MALT1 scaffold targets assembly of the IKK complex, but it presumably occurs
indirectly and involves cofactors such as TRAF proteins (e.g. TRAF2 and TRAF6)112
via CARD9 leads to transcription of genes such as IL1B, IL6, IL23A (encoding the p19
subunit of IL-23), IL12A (IL-12p35), and IL12B (IL-12/IL-23 p40 subunit)74, promoting
both T h1 and T h17 responses52,74. Unlike most PRRs, dectin-1-Syk signaling simultaneously activates the noncanonical NF-κB subunit RelB, which involves NIK. This
pathway partly antagonizes responses induced by dectin-1 through classical NF-κB
signaling, since RelB suppresses IL12B and IL1B transcription, and hence p65- and
c-Rel-mediated IL-12p70 and pro-IL-1β expression, by preventing RNA polymerase II
recruitment74 (Figure 2). Strikingly, dectin-1 activates a second pathway through Raf1 that integrates with Syk-dependent signaling at the level of NF-κB activation, and is
crucial for further fine-tuning of cytokine transcription by dectin-1.
one.
and TAK1, analogous to TCR/BCR signaling81,112. Classical NF-κB signaling by dectin-1
32
Chapter one
The road to Th17 induction via MALT1. As mentioned, dectin-1 is a PRR specialized in
propagation of T h17 responses. Recent studies have given major new insights into two
molecular processes utilized by dectin-1 in human dendritic cells to drive this T h17polarizing cytokine profile. These involve alternative processing of pro-IL-1β 97 and,
as our data in Chapter 2 of this thesis point out, selective activation of NF-κB subunit
c-Rel52. Strikingly, both rely on the MALT1 scaffold protein, underlining a dual role for
MALT1 in shaping T h17 immunity by dectin-197.
MALT1 has been found crucial for pro-IL-1β processing via a noncanonical caspase-8
inflammasome. Production of bioactive IL-1β is strictly regulated and requires proteolytic
processing of pro-IL-1β, formerly thought solely attributable to the NLRP3/caspase-1 inflammasome. Indeed, internalization of some pathogenic fungi by dectin-1 and subsequent
Syk-dependent signaling triggers a conventional NLRP3/caspase-1 inflammasome for
the processing of inactive pro-IL-1β into its 17-kDa mature form. Dectin-1, however, directs
formation of an alternative noncanonical caspase-8 inflammasome for pro-IL-1β cleavage,
without interference of additional cytosolic sensors. This inflammasome consist of the
CARD9/Bcl-10/MALT1 scaffold linked to caspase-8 and the adaptor protein ASC, and is
assembled in response to all (fungal-) pathogens bound by dectin-197. Indirectly, MALT1
has a crucial and indispensable role in caspase-8 inflammasome activity, given that it
directly interacts (presumably dimerization through their respective caspase domains113)
with caspase-8 to prevent autocleavage of caspase-8, which would lead to apoptosis. The
intermediate processing of caspase-8 allows targeted processing of IL-1β97 (Figure 2). Thus,
MALT1 enables dectin-1 to autonomously elicit IL1B transcription as well as rapid IL-1β
maturation, to orchestrate induction of Th17 immunity.
In addition, as we demonstrate in Chapter 2 of this thesis, MALT1 has a specialized
function in regulating expression of Th17-polarizing cytokines IL-23p19 and pro-IL-1β.
Specifically, we demonstrate that MALT1, through the selective targeting of NF-κB subunit
c-Rel, controls a transcriptional subprogram for efficient induction of IL23A and IL1B gene
transcription52, indicating that MALT1 activity is crucial for optimal Th17 effector responses
induced by dectin-1; a notion further discussed in Chapter 6.
Fine-tuning T helper responses by Raf-1. Dectin-1 is also capable of relaying YxxL- and
Syk-independent signaling via the serine/threonine kinase Raf-174, a pathway originally
identified downstream of DC-SIGN (further described later on) 62. Dectin-1-mediated
Raf-1 activation induces selective phosphorylation and subsequent acetylation of p65,
Introduction
which has two important functional consequences. First, phosphorylated p65 can
restrain RelB into inactive p65-RelB dimers to partially reverse the repressing effects
of RelB on IL12B and IL1B transcription. RelB is, however, not completely sequestered
and neutralized by p65; residual RelB can induce moderate production of T h2-related
chemokines CCL17 and CCL22. Acetylation of p65 typically prolongs its activity and
C-type lectin receptors orchestrate antifungal immunity
33
transcription rate, and, downstream dectin-1, results in enhanced expression of IL6,
IL12A and IL10 74 (Figure 2). Overall, the Raf-1 pathway permits and fine-tunes induction of T h1 and T h17 immunity by dectin-1-Syk signaling by balancing p65 and RelB
activities. Clearly, dectin-1-induced NF-κB activation is subject to tight regulation by
distinct pathways which are induced separately but are cooperating at multiple layers
to shape the overall immune response.
DC-SIGN: LIGAND-SPECIFIC SIGNALING FOR TH1 MODULATION
DC-SIGN is a striking example of a signaling CLR that affects adaptive responses induced
by other PRRs at the level of NF-κB activation. Activation of DC-SIGN signaling can have
profound effects on immune responses directed against several pathogens (e.g. mycobacteria)86, and is even exploited by HIV-1 to facilitate productive DC infection and subsequent
in trans infection of T cells - the primary HIV-1 target cells114. Although the role of DC-SIGN in
the host defense against fungi has been studied less extensively, DC-SIGN has the potential
to modulate antifungal effector mechanisms when activated by fungal pathogens. Most
notably, the immunological outcome is dependent on the carbohydrate composition of
the ligand involved; DC-SIGN transduces divergent signaling cascades upon ligation with
mannose- or fucose- containing ligands.
DC-SIGN is predominantly expressed on DC subsets, yet also found on a subpopulation
of macrophages115. Tetrameric surface expression enables DC-SIGN binding to mannose,
fucose, N-acetyl-glucosamine (GlcNAc) and mannan moieties with high-avidity, which
occurs in a Ca2+-dependent manner116. DC-SIGN is involved in the recognition of endogenous
carbohydrates (e.g. ICAM-3 on T cells) as well as ligands derived from numerous pathogens,
including viruses, bateria, helminths, and fungi117. DC-SIGN harbours several internalization motifs in its cytoplasmic tail118, which allow for robust endocytic activity and fungal
uptake; among the fungal pathogens bound via exposed mannose residues are C. albicans,
Despite bearing a classical intracellular YxxL motif, DC-SIGN is an exceptional signaling
CLR in that it does not transduce downstream signals via the Syk-CARD9 axis, and, moreover,
is incapable of activating transcriptional programs (e.g. cytokine expression) on its own62.
A pre-assembled complex consisting of KSR1, CNK and Raf-1 is constitutively attached to
the cytoplasmic domain of DC-SIGN via the adaptor molecule LSP186. Mannose-containing
pathogens such as mycobacteria and HIV-1, and most likely comprising several species of
fungi, induce Raf-1 activation upon DC-SIGN binding. Signaling downstream Raf-1 then leads
to modulation of the transcriptional activity of NF-κB subunit p65, yet only when nuclear
translocation of p65 is induced by any other PRR. Raf-1 phosphorylates p65 at serine (Ser)
276 and controls subsequent acetylation of p65 by two histone acetyltransferases: CBP and
one.
A. fumigatus, capsule-deficient C. neoformans, and Chrysosporium tropicum70,119,120.
34
Chapter one
p300. These post-translational modifications functionally affect p65, defined by prolonged
nuclear activity and increased transcription rates particularly at the IL12A, IL12B, and IL6
genes62. Thus, depending on the PRR coactivated by the fungus, triggering of DC-SIGN/
Raf-1 signaling by mannose-expressing fungi will increase expression of IL-12p70, crucial
for activation of a Th1 polarization program (Figure 2). DC-SIGN-Raf-1 signaling can also
abolish the RelB-dependent suppression of IL12B and IL1B transcription and thereby
directly influence Th17 immunity, as would be the case when dectin-1 is co-ligated. In
contrast, in response to fucose-bearing ligands (i.e. the gastric pathogen Helicobacter pylori
and endogenous Lewis antigens) the KSR1/CNK/Raf-1 triad is selectively disassembled
from the cytoplasmic tail of DC-SIGN for initiation of an alternative, and Raf-1-independent,
signaling cascade that attenuates production of pro-inflammatory cytokines (Figure 2),
with the exact molecular mechansims still being elusive86. This modulatory pathway can
potentially be targeted by pathogenic fungal strains for evading an immune response, with
fucosylated moieties found in selected species of fungi121. Overall, these findings support
the hypothesis that carbohydrate-specific signaling by DC-SIGN can effectively modulate an
antifungal immune response induced by other PRRs, and customize it to the fungal strain
encountered. Whether DC-SIGN indeed affects fungus-specific T helper cell differentiation
awaits further investigation.
DECTIN-2
Apart from being typified as a LC marker122,123, human dectin-2 is more generally expressed
among cells from myeloid origin, specifically DC subtypes, macrophages, monocytes,
and neutrophils14,52,123. Dectin-2 possesses a typical Ca2+-dependent CTLD with affinity for
high-mannose structures (Man9GlcNAc2)124. Consistently, dectin-2 has been implicated in
recognition of α-mannose from a number of fungi: Candida spp., A. fumigatus, capsule-deficient C. neoformans, H. capsulatum, Microsporum audouinii, Malassezia spp. Paracoccoides
brasiliensis, and Trichophyton rubrum52,68,124-126, but also unknown components from house
dust mite allergens, the parasitic worm Schistosoma mansoni, and Mycobacterium tuberculosis124,125,127. In contrast to what its name suggest, dectin-2 is only 27% homologous to dectin-1.
The short cytoplasmic tail of dectin-2 lacks an obvious signaling motif but associates with
the ITAM-bearing Fc receptor common-γ (FcRγ) chain for signal transduction and solid cell
surface expression68. In addition, dectin-2 likely forms a heterodimeric PRR with another,
Introduction
less well-characterized, ITAM-coupled CLR128,129: MCL (also referred to as dectin-3)130. Their
respective homodimers were found to bind α-mannose structures less efficiently, suggesting
that MCL acts as a subunit of a high-affinity PRR complex for sensing fungal infection130.
Engagement of dectin-2 induces phosphorylation of the dual tyrosine residues within
the coupled FcRγ ITAM and, subsequently, recruitment of Src family kinases and Syk68,73. Syk
C-type lectin receptors orchestrate antifungal immunity
35
activation by dectin-2 is a prerequisite not only for CARD9-dependent induction of NF-κBmediated gene transcription, but also for activation of MAP kinase pathways83. Similar to
dectin-1 signaling, canonical NF-κB activation by dectin-2 requires assembly of the CARD9/
Bcl-10/MALT1 scaffold73, and probably involves many of the same intermediate players,
including TRAFs and TAK181. However, as our data in Chapter 2 of this thesis demonstrate,
human DC-expressed dectin-2, in stark contrast to dectin-1, does not equally activate all
NF-κB subunits. We demonstrate that dectin-2-Syk signaling culminates in selective c-Rel
activation via MALT1 for selective production of Th17-polarizing cytokines IL-1β and IL-23
subunit p19 (IL-23p19). Thus, dectin-2 is a representative of a class of signaling CLRs that
modulate antifungal immunity; the implications for Candida albicans-specific Th17 responses
are further discussed in Chapter 6.
Interestingly, dectin-2 has been assigned function in the (dys-) regulation of pulmonary
Th2 responses in a mouse allergy model, further emphasizing a role for dectin-2 as a modulator of T helper cell polarization. Through Syk-dependent generation of proinflammatory
lipid mediators, such as cysteinyl leukotrienes, dectin-2 signaling contributes to pathological airway inflammation in response to extracts derived from house dust mite and A.
fumigatus125,131. Th2 immunity, characterized by alternatively activated macrophages and
antibody class switching to non-opsonizing and IgE subclasses, controls parasitic infections but is considered deleterious during the course of fungal infection16. A. fumigatus is
an exceptional fungus in being a successful opportunistic pathogen and major allergen101,
and might therefore utilize sophisticated strategies to avoid destruction. It remains to be
determined whether dectin-2 signaling in general contributes to Th2-biased immunity,
given its ability to recognize the parasite S. mansoni, and whether dectin-2, in doing so, is
targeted by (virulent) fungal strains for immune evasion. However, consistent with the
immune modulatory functions of dectin-2, this might in fact be dictated by the additional
PRR(s) co-activated by these fungi.
Mincle expression predominates on myeloid cell types, including DCs, macrophages and
neutrophils, yet is also found on B cells132. Through recognition of an ill-defined mannose-rich
ligand, possibly α-mannose, present on glycolipids, mincle recognizes several pathogens
from fungal origin, including Candida spp., Malassezia spp., F. pedrosoi69,84,126,133. Additionally,
mincle exhibits potential to bind the mycobacterial glycolipid trehalose-6,6-dimycolate
(TDM, also known as mycobacterial cordfactor), as well as its synthetic derivate TDB (trehalose-6,6-dibehenate), both well-known for their therapeutic adjuvancy134,135. As such, mincle
is held responsible for the Th1/Th17 adjuvancy of TDM and TDB in mice135,136, yet it is still
unclear whether mincle has a role in controlling Mycobacterium tuberculosis infection137,138.
one.
MINCLE
36
Chapter one
In addition, mincle has been implicated in anti-bacterial defenses; mincle appears to have
a non-redundant role in the protective response to Klebseilla pneumonia infection in mice,
by preventing hyperinflammation, although direct binding of mincle to these bacteria has
not been proven139. While mincle is not directly involved in the phagocytic engulfment of
particulate cargo82,84, it does localize to the phagocytic synapse when macrophages interact
with C. albicans84.
Mincle possesses a single extracellular CRD for the recognition of microbial carbo-
hydrates in a Ca2+-dependent fashion69,140. The mincle CRD contains a primary glutamic
acid/proline-asparagine (EPN) motif, predictive for its mannose specificity, flanked by a
hydrophobic groove, for binding branched acyl chains (fatty acids)140,141. This additional
binding domain probably fine-tunes the specificity of mincle towards the sugar-proximal
parts of lipid-moieties. Its cytoplasmic tail is devoid of any classic signaling motifs, and via
a positively charged arginine residue, mincle associates with the FcRγ chain for transduction of ITAM-coupled signaling, in analogy to dectin-282. Moreover, and similar to dectin-2,
mincle heterodimerizes with MCL for cell surface expression142. Although structural analyses have demonstrated that MCL also contains a hydrophobic domain for binding the
branched acyl chains present in TDM and TDB, its CRD lacks a primary EPN motif140, critical
for TDM binding by mincle134. Given that MCL binds TDM with much lower affinity142, and
that heterodimer formation with MCL is not unique to mincle, the mincle/MCL interaction
possibly confers additional function to each of the molecules. Most likely, heterodimerization
with MCL enhances the ligand binding affinity, allowing mincle to detect small numbers
of glycolipids on the fungal surface. Furthermore, coupling to MCL might ensure efficient
signal transduction, and could also reflect a mechanism by which mincle acquires endocytic receptor function143, with MCL demonstrated to exhibit endocytic activity128, in stark
contrast to mincle82,84. Whether mincle and MCL indeed function cooperatively, also in the
context of human antifungal immunity, remains to be demonstrated.
Next to its role as a microbial sensor, mincle is involved in the innate recognition of dam-
aged-self. As such, mincle binds the ribonuceoprotein SAP-130, derived from necrotic cells
that have lost their membrane integrity82. The mincle-mediated response to damaged-self
comprises infiltration of neutrophils for cellular clean-up82, with a possible role for mincle in
the induction of pathological inflammatory events following ischemic stroke144. Strikingly,
mincle recognizes SAP-130 in the absence of Ca2+ and independent of the CRD residues
involved in fungal binding82, implying dual ligand specificity. Also, the proinflammatory
response to necrotic cell death by mincle can be considered a sterile reaction, linked to tis-
Introduction
sue repair, rather than infection control and full-blown activation of adaptive immunity145.
It remains to be established if mincle, analogous DC-SIGN, transduces divergent signaling
pathways to influence these distinct proinflammatory processes, or whether this is actually
dependent on the presence or absence of co-stimulated PRRs.
Ligation of murine mincle activates the highly conserved Syk-CARD9 signaling axis81,82,134,
C-type lectin receptors orchestrate antifungal immunity
37
whereas its activity in the context of murine Fonsecaea infection has been shown inadequate
for induction of protective antifungal responses85. Although costimulation of other PRRs
was demonstrated sufficient for subsequent clearance of the pathogen, these protective
responses were dependent on mincle signaling as well, indicative of a modulatory function for mincle during establishment of antifungal immunity85. Our work on the functional
characterization of human mincle described in this thesis confirmed and extended these
previous findings, by demonstrating that mincle-dependent Syk signaling in human DCs
culminates in assembly of the complete CARD9/Bcl-10/MALT1 signaling module (Chapter
3). Most strikingly, though, we found that mincle does not signal for NF-κB activation (Chapter
4), and that mincle does not exhibit the potential to activate cytokine responses on its own
(Chapters 3 and 4). Instead, our data uncover an alternate mechanism of action: mincle couples Syk-CARD9 signaling to a PI(3)K-PKB pathway for modulation of cytokine responses
induced by other PRRs (Chapters 3, 4 and 5), having serious implications for the ability of
human DCs to promote concurrent Th1 and Th17 responses during fungal infection, an
issue further discussed in Chapter 6 of this thesis.
CONCLUDING REMARKS AND OUTLOOK
As defined and discussed here, immunity to fungal infection is orchestrated by multiple CLRs
expressed on dendritic cells. Two types of antifungal CLRs can be recognized: (i) receptors
that evoke a T helper polarization program autonomously and (ii) receptors that modulate
signals from other receptors to fine-tune a particular response. Unbalanced inflammation
can have deleterious effects on host immunity, and it is becoming evident that these types
of CLRs together transduce collaborative signaling to ensure tightly controlled and tailored
effector responses upon fungal intrusion. Further characterization of the molecular mechanisms by which CLRs control these processes may therefore give important insights into
defense mechanisms deployed against pathogenic fungi, and will facilitate tailor made
Antiviral effectors for antifungal control? Our understanding of antifungal immune
responses has been expanded recently by studies supporting a possible role for type
I IFNs -classic antiviral effectors- during fungal-induced inflammation. Type I IFNs are
produced by murine DCs in response to cytoplasmic fungal nucleic acids 146,147, but also
it has been demonstrated that murine dectin-1 directly signals for IFN-β expression148,
though precise functional contribution to the overall antifungal immune response
remains enigmatic. Moreover, defective expression of human type I IFN genes is
associated with susceptibility to CMC, albeit through unknown mechanisms149. Our
study described in Chapter 5 corroborates these initial findings by providing evidence
one.
vaccines and novel targets to combat fungal infections.
38
for contribution of a type I IFN response to
Chapter one
REFERENCES
human antifungal inflammation, which
also sheds new light on the molecular prin-
1. Brown, G.D., Denning, D.W.,
ciples of CLR cooperation.
Gow, N.A., Levitz, S.M. et al.
Hidden killers: human fungal
infections. Sci. Transl. Med. 4,
165rv113 (2012).
2. Iliev, I.D., Funari, V.A., Taylor,
K.D., Nguyen, Q. et al.
Interactions between
commensal fungi and the
C-type lectin receptor Dectin-1
influence colitis. Science 336,
1314-1317 (2012).
3. Miceli, M.H., Diaz, J.A. & Lee,
S.A. Emerging opportunistic
yeast infections. Lancet Infect.
Dis. 11, 142-151 (2011).
4. Hoogeboom, R., van Kessel,
K.P., Hochstenbach, F.,
Wormhoudt, T.A. et al. A
mutated B cell chronic
lymphocytic leukemia subset
that recognizes and responds
to fungi. J. Exp. Med. 210, 59-70
(2013).
5. Kronstad, J.W., Attarian, R.,
Cadieux, B., Choi, J. et al.
Expanding fungal pathogenesis: Cryptococcus breaks out of
Introduction
the opportunistic box. Nat. Rev.
Microbiol. 9, 193-203 (2011).
6. Sommer, F. & Backhed, F. The
gut microbiota--masters of host
39
C-type lectin receptors orchestrate antifungal immunity
18. van de Veerdonk, F.L. &
development and physiology.
Leibundgut-Landmann, S.
Nat. Rev. Microbiol. 11, 227-238
Cutting Edge: IL-17-Secreting
Netea, M.G. T-cell Subsets and
(2013).
Innate Lymphoid Cells Are
Antifungal Host Defenses.
Essential for Host Defense
Current fungal infection reports
7. Iliev, I.D. & Underhill, D.M.
against Fungal Infection. J.
4, 238-243 (2010).
Striking a balance: fungal
Immunol. 190, 521-525 (2013).
19. Schroder, K., Hertzog, P.J.,
commensalism versus
pathogenesis. Curr. Opin.
13. Brown, G.D. Innate antifun-
Microbiol. 16, 366-373 (2013).
gal immunity: the key role of
Interferon-gamma: an overview
phagocytes. Annu. Rev.
of signals, mechanisms and
8. Hooper, L.V. & Macpherson,
Immunol. 29, 1-21 (2011).
A.J. Immune adaptations that
Ravasi, T. & Hume, D.A.
functions. J. Leukoc. Biol. 75,
163-189 (2004).
maintain homeostasis with the
14. Taylor, P.R., Roy, S., Leal, S.M.,
intestinal microbiota. Nat. Rev.
Jr., Sun, Y. et al. Activation of
Immunol. 10, 159-169 (2010).
neutrophils by autocrine
Prostaglandin E2 enhances and
IL-17A-IL-17RC interactions
gamma interferon inhibits germ
20. Kalo-Klein, A. & Witkin, S.S.
9. de Repentigny, L., Aumont, F.,
during fungal infection is
tube formation in Candida
Bernard, K. & Belhumeur, P.
regulated by IL-6, IL-23,
albicans. Infect. Immun. 58,
Characterization of binding of
RORgammat and dectin-2. Nat.
260-262 (1990).
Candida albicans to small
Immunol. (2013).
21. Korn, T., Bettelli, E., Oukka, M.
intestinal mucin and its role in
adherence to mucosal
15. Casadevall, A. & Pirofski, L.A.
& Kuchroo, V.K. IL-17 and Th17
epithelial cells. Infect. Immun.
Immunoglobulins in defense,
Cells. Annu. Rev. Immunol. 27,
68, 3172-3179 (2000).
pathogenesis, and therapy of
485-517 (2009).
fungal diseases. Cell host &
microbe 11, 447-456 (2012).
Cleavage of E-cadherin: a
22. De Luca, A., Zelante, T.,
D’A ngelo, C., Zagarella, S. et al.
mechanism for disruption of
16. Wuthrich, M., Deepe, G.S., Jr.
IL-22 defines a novel immune
the intestinal epithelial barrier
& Klein, B. Adaptive immunity
pathway of antifungal
by Candida albicans. Transl.
to fungi. Annu. Rev. Immunol.
resistance. Mucosal Immunol. 3,
Res. 149, 211-222 (2007).
30, 115-148 (2012).
361-373 (2010).
11. van Spriel, A.B., Sofi, M.,
17. Park, B.J., Wannemuehler,
23. Kolls, J.K., McCray, P.B., Jr. &
Gartlan, K.H., van der Schaaf, A.
K.A., Marston, B.J., Govender, N.
Chan, Y.R. Cytokine-mediated
et al. The tetraspanin protein
et al. Estimation of the current
regulation of antimicrobial
CD37 regulates IgA responses
global burden of cryptococcal
proteins. Nat. Rev. Immunol. 8,
and anti-fungal immunity. PLoS
menigitis among persons living
829-835 (2008).
Pathog. 5, e1000338 (2009).
with HIV/AIDS. AIDS 23,
525-530 (2009).
24. Milner, J.D., Brenchley, J.M.,
12. Gladiator, A., Wangler, N.,
Laurence, A., Freeman, A.F. et al.
Trautwein-Weidner, K. &
Impaired T(H)17 cell differentia-
one.
10. Frank, C.F. & Hostetter, M.K.
40
Chapter one
tion in subjects with autosomal
29. Kisand, K., Boe Wolff, A.S.,
35. LeibundGut-Landmann, S.,
dominant hyper-IgE syndrome.
Podkrajsek, K.T., Tserel, L. et al.
Wuthrich, M. & Hohl, T.M.
Chronic mucocutaneous
Immunity to fungi. Curr. Opin.
candidiasis in APECED or
Immunol. 24, 449-458 (2012).
Nature 452, 773-776 (2008).
25. Ma, C.S., Chew, G.Y., Simpson,
thymoma patients correlates
N., Priyadarshi, A. et al.
with autoimmunity to
36. Zhu, J., Yamane, H. & Paul,
Deficiency of Th17 cells in
Th17-associated cytokines. J.
W.E. Differentiation of effector
hyper IgE syndrome due to
Exp. Med. 207, 299-308 (2010).
CD4 T cell populations (*). Annu.
mutations in STAT3. J. Exp. Med.
205, 1551-1557 (2008).
Rev. Immunol. 28, 445-489
30. Puel, A., Cypowyj, S.,
(2010).
Bustamante, J., Wright, J.F. et al.
26. Liu, L., Okada, S., Kong, X.F.,
Chronic mucocutaneous
37. Iwasaki, A. & Medzhitov, R.
Kreins, A.Y. et al. Gain-of-
candidiasis in humans with
Regulation of adaptive
function human STAT1
inborn errors of interleukin-17
immunity by the innate
mutations impair IL-17
immunity. Science 332, 65-68
immune system. Science 327,
immunity and underlie chronic
(2011).
291-295 (2010).
31. Conti, H.R. & Gaffen, S.L. Host
38. Matzinger, P. The danger
responses to Candida albicans:
model: a renewed sense of self.
Th17 cells and mucosal
Science 296, 301-305 (2002).
mucocutaneous candidiasis. J.
Exp. Med. 208, 1635-1648
(2011).
27. van de Veerdonk, F.L.,
candidiasis. Microbes Infect. 12,
Plantinga, T.S., Hoischen, A.,
518-527 (2010).
Smeekens, S.P. et al. STAT1
Medzhitov, R. Innate immune
mutations in autosomal
32. Hernandez-Santos, N. &
recognition. Annu. Rev.
dominant chronic mucocutane-
Gaffen, S.L. Th17 cells in
Immunol. 20, 197-216 (2002).
ous candidiasis. N. Engl. J. Med.
immunity to Candida albicans.
365, 54-61 (2011).
Cell host & microbe 11, 425-435
40. Geijtenbeek, T.B. & Gringhuis,
(2012).
S.I. Signalling through C-type
28. Puel, A., Doffinger, R.,
lectin receptors: shaping
Natividad, A., Chrabieh, M. et al.
33. Littman, D.R. & Rudensky,
immune responses. Nat. Rev.
Autoantibodies against IL-17A,
A.Y. Th17 and regulatory T cells
Immunol. 9, 465-479 (2009).
IL-17F, and IL-22 in patients
in mediating and restraining
with chronic mucocutaneous
inflammation. Cell 140, 845-858
41. Takeuchi, O. & Akira, S.
candidiasis and autoimmune
(2010).
Pattern recognition receptors
polyendocrine syndrome type
Introduction
39. Janeway, C.A., Jr. &
and inflammation. Cell 140,
I. J. Exp. Med. 207, 291-297
34. Romani, L. Immunity to
(2010).
fungal infections. Nat. Rev.
Immunol. 11, 275-288 (2011).
805-820 (2010).
42. Philpott, D.J., Sorbara, M.T.,
Robertson, S.J., Croitoru, K. &
41
C-type lectin receptors orchestrate antifungal immunity
Girardin, S.E. NOD proteins:
48. Curotto de Lafaille, M.A. &
sour treat for toll-like receptors.
regulators of inflammation in
Lafaille, J.J. Natural and
Front. Cell. Infect. Microbiol. 2,
142 (2012).
health and disease. Nat. Rev.
adaptive foxp3+ regulatory T
Immunol. 14, 9-23 (2014).
cells: more of the same or a
division of labor? Immunity 30,
55. Wuthrich, M., Gern, B., Hung,
43. Kapsenberg, M.L. Dendritic-
626-635 (2009).
C.Y., Ersland, K. et al. Vaccine-
cell control of pathogen-driven
induced protection against 3
T-cell polarization. Nat. Rev.
49. Latge, J.P. Tasting the fungal
systemic mycoses endemic to
Immunol. 3, 984-993 (2003).
cell wall. Cell. Microbiol. 12,
North America requires Th17
863-872 (2010).
cells in mice. J. Clin. Invest. 121,
44. Ghosh, S., May, M.J. & Kopp,
554-568 (2011).
E.B. NF-kappa B and Rel
50. Bowman, S.M. & Free, S.J.
proteins: evolutionarily
The structure and synthesis of
56. von Bernuth, H., Picard, C.,
conserved mediators of
the fungal cell wall. Bioessays
Jin, Z., Pankla, R. et al. Pyogenic
immune responses. Annu. Rev.
28, 799-808 (2006).
Immunol. 16, 225-260 (1998).
bacterial infections in humans
with MyD88 deficiency. Science
51. Levitz, S.M. Innate recogni45. Moser, M. & Murphy, K.M.
tion of fungal cell walls. PLoS
Dendritic cell regulation of
Pathog. 6, e1000758 (2010).
321, 691-696 (2008).
57. Plantinga, T.S., Johnson, M.D.,
Scott, W.K., van de Vosse, E. et al.
Th1-Th2 development. Nat.
Immunol. 1, 199-205 (2000).
52. Gringhuis, S.I., Wevers, B.A.,
Toll-like receptor 1 polymor-
Kaptein, T.M., van Capel, T.M. et
phisms increase susceptibility
46. Thieu, V.T., Yu, Q., Chang, H.C.,
al. Selective C-Rel activation via
to candidemia. J. Infect. Dis. 205,
Yeh, N. et al. Signal transducer
MALT1 controls anti-fungal
934-943 (2012).
and activator of transcription 4
T(H)-17 immunity by dectin-1
is required for the transcription
and dectin-2. PLoS Pathog. 7,
58. Bochud, P.Y., Chien, J.W.,
factor T-bet to promote T helper
e1001259 (2011).
Marr, K.A., Leisenring, W.M. et al.
Immunity 29, 679-690 (2008).
53. Lemaitre, B., Nicolas, E.,
phisms and aspergillosis in
Michaut, L., Reichhart, J.M. &
stem-cell transplantation. N.
47. Zielinski, C.E., Mele, F.,
Hoffmann, J.A. The dorsoven-
Engl. J. Med. 359, 1766-1777
Aschenbrenner, D., Jarrossay, D.
tral regulatory gene cassette
(2008).
et al. Pathogen-induced human
spatzle/Toll/cactus controls the
Th17 cells produce IFN-gamma
potent antifungal response in
59. Akira, S., Uematsu, S. &
or IL-10 and are regulated by
Drosophila adults. Cell 86,
Takeuchi, O. Pathogen
IL-1beta. Nature 484, 514-518
973-983 (1996).
recognition and innate
immunity. Cell 124, 783-801
(2012).
54. Bourgeois, C. & Kuchler, K.
Fungal pathogens-a sweet and
(2006).
one.
Toll-like receptor 4 polymor-
1 cell-fate determination.
42
Chapter one
60. Sutmuller, R.P., den Brok,
that induces the formation of
activating receptor for
M.H., Kramer, M., Bennink, E.J. et
Birbeck granules. Immunity 12,
pathogenic fungus, Malassezia.
al. Toll-like receptor 2 controls
71-81 (2000).
expansion and function of
regulatory T cells. J. Clin. Invest.
65. Zhang, J., Zhu, J., Bu, X.,
116, 485-494 (2006).
Cushion, M. et al. Cdc42 and
70. Cambi, A., Gijzen, K., de Vries
RhoB activation are required for
l, J., Torensma, R. et al. The
61. Veldhoen, M., Hocking, R.J.,
mannose receptor-mediated
C-type lectin DC-SIGN (CD209)
Atkins, C.J., Locksley, R.M. &
phagocytosis by human
is an antigen-uptake receptor
Stockinger, B. TGFbeta in the
alveolar macrophages. Mol. Biol.
for Candida albicans on
context of an inflammatory
Cell 16, 824-834 (2005).
dendritic cells. Eur. J. Immunol.
cytokine milieu supports de
33, 532-538 (2003).
novo differentiation of
66. Cambi, A., Netea, M.G.,
IL-17-producing T cells.
Mora-Montes, H.M., Gow, N.A. et
71. Heinsbroek, S.E., Taylor, P.R.,
Immunity 24, 179-189 (2006).
al. Dendritic cell interaction
Martinez, F.O., Martinez-
with Candida albicans critically
Pomares, L. et al. Stage-specific
62. Gringhuis, S.I., den Dunnen,
depends on N-linked mannan. J.
sampling by pattern recogni-
J., Litjens, M., van Het Hof, B. et al.
Biol. Chem. 283, 20590-20599
tion receptors during Candida
C-type lectin DC-SIGN
(2008).
albicans phagocytosis. PLoS
Pathog. 4, e1000218 (2008).
modulates Toll-like receptor
signaling via Raf-1 kinase-de-
67. Taylor, P.R., Tsoni, S.V.,
pendent acetylation of
Willment, J.A., Dennehy, K.M. et
72. LeibundGut-Landmann, S.,
transcription factor NF-kappaB.
al. Dectin-1 is required for
Gross, O., Robinson, M.J., Osorio,
Immunity 26, 605-616 (2007).
63. de Jong, M.A., Vriend, L.E.,
beta-glucan recognition and
F. et al. Syk- and CARD9-
control of fungal infection. Nat.
dependent coupling of innate
Immunol. 8, 31-38 (2007).
Theelen, B., Taylor, M.E. et al.
immunity to the induction of T
helper cells that produce
C-type lectin Langerin is a
68. Sato, K., Yang, X.L., Yudate, T.,
interleukin 17. Nat. Immunol. 8,
630-638 (2007).
beta-glucan receptor on human
Chung, J.S. et al. Dectin-2 is a
Langerhans cells that recog-
pattern recognition receptor for
nizes opportunistic and
fungi that couples with the Fc
73. Robinson, M.J., Osorio, F.,
pathogenic fungi. Mol.
receptor gamma chain to
Rosas, M., Freitas, R.P. et al.
Immunol. 47, 1216-1225 (2010).
induce innate immune
Dectin-2 is a Syk-coupled
responses. J. Biol. Chem. 281,
pattern recognition receptor
38854-38866 (2006).
crucial for Th17 responses to
K. et al. Langerin, a novel C-type
69. Yamasaki, S., Matsumoto, M.,
206, 2037-2051 (2009).
lectin specific to Langerhans
Takeuchi, O., Matsuzawa, T. et al.
cells, is an endocytic receptor
C-type lectin Mincle is an
64. Valladeau, J., Ravel, O.,
Introduction
Proc. Natl. Acad. Sci. U. S. A. 106,
1897-1902 (2009).
Dezutter-Dambuyant, C., Moore,
fungal infection. J. Exp. Med.
43
C-type lectin receptors orchestrate antifungal immunity
74. Gringhuis, S.I., den Dunnen,
Spriel, A.B. et al. Early stop
83. Saijo, S., Ikeda, S., Yamabe, K.,
J., Litjens, M., van der Vlist, M. et
polymorphism in human
Kakuta, S. et al. Dectin-2
al. Dectin-1 directs T helper cell
DECTIN-1 is associated with
recognition of alpha-mannans
differentiation by controlling
increased candida colonization
and induction of Th17 cell
noncanonical NF-kappaB
in hematopoietic stem cell
differentiation is essential for
activation through Raf-1 and
transplant recipients. Clin.
host defense against Candida
Syk. Nat. Immunol. 10, 203-213
Infect. Dis. 49, 724-732 (2009).
albicans. Immunity 32, 681-691
(2009).
(2010).
79. Glocker, E.O., Hennigs, A.,
75. Ferwerda, B., Ferwerda, G.,
Nabavi, M., Schaffer, A.A. et al. A
84. Wells, C.A., Salvage-Jones,
Plantinga, T.S., Willment, J.A. et
homozygous CARD9 mutation
J.A., Li, X., Hitchens, K. et al. The
al. Human dectin-1 deficiency
in a family with susceptibility to
macrophage-inducible C-type
and mucocutaneous fungal
fungal infections. N. Engl. J. Med.
lectin, mincle, is an essential
infections. N. Engl. J. Med. 361,
361, 1727-1735 (2009).
component of the innate
80. Drewniak, A.A., Gazendam,
albicans. J. Immunol. 180,
76. Chai, L.Y., de Boer, M.G., van
R.P., Tool, A.T., van Houdt, M. et al.
7404-7413 (2008).
der Velden, W.J., Plantinga, T.S.
Invasive fungal infection and
1760-1767 (2009).
immune response to Candida
et al. The Y238X stop codon
impaired neutrophil killing in
85. Sousa Mda, G., Reid, D.M.,
polymorphism in the human
human CARD9 deficiency.
Schweighoffer, E., Tybulewicz,
beta-glucan receptor dectin-1
Blood (2013).
V. et al. Restoration of pattern
and susceptibility to invasive
recognition receptor costimula81. Strasser, D., Neumann, K.,
tion to treat chromoblastomy-
Bergmann, H., Marakalala, M.J.
cosis, a chronic fungal infection
et al. Syk kinase-coupled C-type
of the skin. Cell host & microbe
77. Cunha, C., Di Ianni, M., Bozza,
lectin receptors engage protein
9, 436-443 (2011).
S., Giovannini, G. et al. Dectin-1
kinase C-sigma to elicit Card9
Y238X polymorphism
adaptor-mediated innate
86. Gringhuis, S.I., den Dunnen,
associates with susceptibility to
immunity. Immunity 36, 32-42
J., Litjens, M., van der Vlist, M. &
invasive aspergillosis in
(2012).
Geijtenbeek, T.B. Carbohydrate-
736-743 (2011).
specific signaling through the
hematopoietic transplantation
through impairment of both
82. Yamasaki, S., Ishikawa, E.,
DC-SIGN signalosome tailors
recipient- and donor-dependent
Sakuma, M., Hara, H. et al. Mincle
immunity to Mycobacterium
mechanisms of antifungal
is an ITAM-coupled activating
tuberculosis, HIV-1 and
immunity. Blood 116, 5394-
receptor that senses damaged
Helicobacter pylori. Nat.
cells. Nat. Immunol. 9, 1179-
Immunol. 10, 1081-1088 (2009).
5402 (2010).
1188 (2008).
78. Plantinga, T.S., van der
87. Willment, J.A., Marshall, A.S.,
Velden, W.J., Ferwerda, B., van
Reid, D.M., Williams, D.L. et al.
one.
aspergillosis. J. Infect. Dis. 203,
44
The human beta-glucan
Chapter one
fungal pathogen Cryptococcus
IL-1beta via a noncanonical
receptor is widely expressed
neoformans. Infect. Immun. 77,
caspase-8 inflammasome. Nat.
and functionally equivalent to
3491-3500 (2009).
Immunol. 13, 246-254 (2012).
murine Dectin-1 on primary
cells. Eur. J. Immunol. 35,
93. Sorgi, C.A., Secatto, A.,
98. Wheeler, R.T., Kombe, D.,
1539-1547 (2005).
Fontanari, C., Turato, W.M. et al.
Agarwala, S.D. & Fink, G.R.
Histoplasma capsulatum cell
Dynamic, morphotype-specific
88. Martin, B., Hirota, K., Cua, D.J.,
wall {beta}-glucan induces lipid
Candida albicans beta-glucan
Stockinger, B. & Veldhoen, M.
body formation through CD18,
exposure during infection and
Interleukin-17-producing
TLR2, and dectin-1 receptors:
drug treatment. PLoS Pathog. 4,
gammadelta T cells selectively
correlation with leukotriene B4
e1000227 (2008).
expand in response to
generation and role in HIV-1
pathogen products and
infection. J. Immunol. 182,
99. Blander, J.M. & Sander, L.E.
environmental signals.
4025-4035 (2009).
Beyond pattern recognition:
94. Steele, C., Marrero, L., Swain,
scaling the microbial threat. Nat.
Immunity 31, 321-330 (2009).
five immune checkpoints for
89. Steele, C., Rapaka, R.R., Metz,
S., Harmsen, A.G. et al. Alveolar
Rev. Immunol. 12, 215-225
A., Pop, S.M. et al. The beta-glu-
macrophage-mediated killing
(2012).
can receptor dectin-1 recog-
of Pneumocystis carinii f. sp.
nizes specific morphologies of
muris involves molecular
Aspergillus fumigatus. PLoS
recognition by the Dectin-1
K.J. Complementation of a
Pathog. 1, e42 (2005).
beta-glucan receptor. J. Exp.
capsule-deficient mutation of
Med. 198, 1677-1688 (2003).
Cryptococcus neoformans
90. Brown, G.D., Herre, J.,
restores its virulence. Mol. Cell.
Williams, D.L., Willment, J.A. et
95. Viriyakosol, S., Jimenez
al. Dectin-1 mediates the
Mdel, P., Gurney, M.A.,
biological effects of beta-glu-
Ashbaugh, M.E. & Fierer, J.
101. Aimanianda, V., Bayry, J.,
cans. J. Exp. Med. 197, 1119-
Dectin-1 is required for
Bozza, S., Kniemeyer, O. et al.
1124 (2003).
resistance to coccidioidomyco-
Surface hydrophobin prevents
sis in mice. mBio 4(2013).
immune recognition of
91. Viriyakosol, S., Fierer, J.,
Biol. 14, 4912-4919 (1994).
airborne fungal spores. Nature
Brown, G.D. & Kirkland, T.N.
96. Yadav, M. & Schorey, J.S. The
Innate immunity to the
beta-glucan receptor dectin-1
460, 1117-1121 (2009).
pathogenic fungus
functions together with TLR2
102. Ariizumi, K., Shen, G.L.,
Coccidioides posadasii is
to mediate macrophage
Shikano, S., Xu, S. et al. Identi-
dependent on Toll-like receptor
activation by mycobacteria.
fication of a novel, dendritic
2 and Dectin-1. Infect. Immun.
Blood 108, 3168-3175 (2006).
cell-associated molecule,
73, 1553-1560 (2005).
Introduction
100. Chang, Y.C. & Kwon-Chung,
dectin-1, by subtractive cDNA
97. Gringhuis, S.I., Kaptein, T.M.,
cloning. J. Biol. Chem. 275,
92. Giles, S.S., Dagenais, T.R.,
Wevers, B.A., Theelen, B. et al.
20157-20167 (2000).
Botts, M.R., Keller, N.P. & Hull,
Dectin-1 is an extracellular
C.M. Elucidating the pathogene-
pathogen sensor for the
103. Kerrigan, A.M. & Brown, G.D.
sis of spores from the human
induction and processing of
Syk-coupled C-type lectin
45
C-type lectin receptors orchestrate antifungal immunity
receptors that mediate cellular
Calcineurin regulates innate
infection of dendritic cells. Nat.
activation via single tyrosine
antifungal immunity in
Immunol. 11, 419-426 (2010).
based activation motifs.
neutrophils. J. Exp. Med. 207,
Immunol. Rev. 234, 335-352
923-931 (2010).
115. Soilleux, E.J., Morris, L.S.,
110. Ma, J., Becker, C., Lowell, C.A.
Constitutive and induced
(2010).
Leslie, G., Chehimi, J. et al.
104. Rogers, N.C., Slack, E.C.,
& Underhill, D.M. Dectin-1-
expression of DC-SIGN on
Edwards, A.D., Nolte, M.A. et al.
triggered recruitment of light
dendritic cell and macrophage
Syk-dependent cytokine
chain 3 protein to phagosomes
subpopulations in situ and in
induction by Dectin-1 reveals a
facilitates major histocompati-
vitro. J. Leukoc. Biol.71, 445-457
novel pattern recognition
bility complex class II
(2002).
pathway for C type lectins.
presentation of fungal-derived
Immunity 22, 507-517 (2005).
antigens. J. Biol. Chem. 287,
116. Koppel, E.A., van Gisbergen,
34149-34156 (2012).
K.P., Geijtenbeek, T.B. & van
105. Hughes, C.E., Pollitt, A.Y.,
Kooyk, Y. Distinct functions of
Mori, J., Eble, J.A. et al. CLEC-2
111. Gross, O., Gewies, A., Finger,
activates Syk through
K., Schafer, M. et al. Card9
L-SIGN (DC-SIGNR) and
dimerization. Blood 115,
controls a non-TLR signalling
mSIGNR1 in pathogen
2947-2955 (2010).
pathway for innate anti-fungal
recognition and immune
DC-SIGN and its homologues
immunity. Nature 442, 651-656
regulation. Cell. Microbiol. 7,
(2006).
157-165 (2005).
tyrosine kinase: a crucial player
112. Rawlings, D.J., Sommer, K. &
117. van den Berg, L.M.,
in diverse biological functions.
Moreno-Garcia, M.E. The
Gringhuis, S.I. & Geijtenbeek, T.B.
Nat. Rev. Immunol. 10, 387-402
CARMA1 signalosome links the
An evolutionary perspective on
(2010).
signalling machinery of
C-type lectins in Infect. Immun..
adaptive and innate immunity
Ann. N. Y. Acad. Sci.1253,
107. Goodridge, H.S., Reyes, C.N.,
in lymphocytes. Nat. Rev.
149-158 (2012).
Becker, C.A., Katsumoto, T.R. et
Immunol. 6, 799-812 (2006).
106. Mocsai, A., Ruland, J. &
Tybulewicz, V.L. The SYK
immune receptor Dectin-1
113. Kawadler, H., Gantz, M.A.,
T.B., van Vliet, S.J., Wijers, M. et al.
upon formation of a ‘phagocytic
Riley, J.L. & Yang, X. The
The dendritic cell-specific
synapse’. Nature 472, 471-475
paracaspase MALT1 controls
adhesion receptor DC-SIGN
(2011).
caspase-8 activation during
internalizes antigen for
lymphocyte proliferation. Mol.
presentation to T cells. J.
108. Goodridge, H.S., Wolf, A.J. &
Cell 31, 415-421 (2008).
Underhill, D.M. Beta-glucan
Immunol. 168, 2118-2126
(2002).
recognition by the innate
114. Gringhuis, S.I., van der Vlist,
immune system. Immunol. Rev.
M., van den Berg, L.M., den
230, 38-50 (2009).
Dunnen, J. et al. HIV-1 exploits
Dominguez-Soto, A., Ancochea,
innate signaling by TLR8 and
J., Jimenez-Heffernan, J.A. et al.
DC-SIGN for productive
Dendritic cell-specific
109. Greenblatt, M.B., Aliprantis,
A., Hu, B. & Glimcher, L.H.
119. Serrano-Gomez, D.,
intercellular adhesion
one.
118. Engering, A., Geijtenbeek,
al. Activation of the innate
46
Chapter one
molecule 3-grabbing noninteg-
124. McGreal, E.P., Rosas, M.,
endocytic receptor. Eur. J.
rin mediates binding and
Brown, G.D., Zamze, S. et al. The
Immunol. 34, 210-220 (2004).
internalization of Aspergillus
carbohydrate-recognition
fumigatus conidia by dendritic
domain of Dectin-2 is a C-type
129. Graham, L.M., Gupta, V.,
cells and macrophages. J.
lectin with specificity for high
Schafer, G., Reid, D.M. et al. The
Immunol. 173, 5635-5643
mannose. Glycobiology 16,
C-type lectin receptor CLECSF8
(2004).
422-430 (2006).
(CLEC4D) is expressed by
120. Mansour, M.K., Latz, E. &
125. Barrett, N.A., Maekawa, A.,
cellular activation through Syk
Levitz, S.M. Cryptococcus
Rahman, O.M., Austen, K.F. &
kinase. J. Biol. Chem. 287,
neoformans glycoantigens are
Kanaoka, Y. Dectin-2 recogni-
25964-25974 (2012).
myeloid cells and triggers
captured by multiple lectin
tion of house dust mite triggers
receptors and presented by
cysteinyl leukotriene genera-
130. Zhu, L.L., Zhao, X.Q., Jiang,
dendritic cells. J. Immunol. 176,
tion by dendritic cells. J.
C., You, Y. et al. C-type lectin
Immunol. 182, 1119-1128
receptors Dectin-3 and Dectin-2
(2009).
form a heterodimeric pat-
3053-3061 (2006).
121. Takashima, M., Hamamoto,
M. & Nakase, T. Taxonomic
tern-recognition receptor for
126. Ishikawa, T., Itoh, F., Yoshida,
host defense against fungal
significance of fucose in the
S., Saijo, S. et al. Identification of
infection. Immunity 39,
class Urediniomycetes:
Distinct Ligands for the C-type
324-334 (2013).
distribution of fucose in cell
Lectin Receptors Mincle and
wall and phylogeny of
Dectin-2 in the Pathogenic
131. Clarke, D.L., Davis, N.H.,
urediniomycetous yeasts. Syst.
Fungus Malassezia. Cell host &
Campion, C.L., Foster, M.L. et al.
Appl. Microbiol. 23, 63-70 (2000).
microbe 13, 477-488 (2013).
Dectin-2 sensing of house dust
mite is critical for the initiation
122. Ariizumi, K., Shen, G.L.,
127. Ritter, M., Gross, O., Kays, S.,
of airway inflammation.
Shikano, S., Ritter, R., 3rd et al.
Ruland, J. et al. Schistosoma
Mucosal Immunol. (2013).
Cloning of a second dendritic
mansoni triggers Dectin-2,
cell-associated C-type lectin
which activates the Nlrp3
132. Vijayan, D., Radford, K.J.,
(dectin-2) and its alternatively
inflammasome and alters
Beckhouse, A.G., Ashman, R.B. &
spliced isoforms. J. Biol. Chem.
adaptive immune responses.
Wells, C.A. Mincle polarizes
275, 11957-11963 (2000).
Proc. Natl. Acad. Sci. U.S.A. 107,
human monocyte and
20459-20464 (2010).
Introduction
123. Kanazawa, N., Tashiro, K.,
neutrophil responses to
Candida albicans. Immunol.
Inaba, K., Lutz, M.B. & Miyachi, Y.
128. Arce, I., Martinez-Munoz, L.,
Molecular cloning of human
Roda-Navarro, P. & Fernandez-
dectin-2. J. Invest. Dermatol. 122,
Ruiz, E. The human C-type
133. Bugarcic, A., Hitchens, K.,
lectin CLECSF8 is a novel
Beckhouse, A.G., Wells, C.A. et al.
monocyte/macrophage
Human and mouse macro-
1522-1524 (2004).
Cell Biol. 90, 889-895 (2012).
47
C-type lectin receptors orchestrate antifungal immunity
phage-inducible C-type lectin
138. Lang, R. Recognition of the
rial cord factor. Immunity 38,
(Mincle) bind Candida albicans.
mycobacterial cord factor by
1050-1062 (2013).
Glycobiology 18, 679-685
Mincle: relevance for granu-
(2008).
loma formation and resistance
143. Lobato-Pascual, A., Saether,
to tuberculosis. Front. Immunol.
P.C., Fossum, S., Dissen, E. &
134. Ishikawa, E., Ishikawa, T.,
4, 5 (2013).
Daws, M.R. Mincle, the receptor
Morita, Y.S., Toyonaga, K. et al.
for mycobacterial cord factor,
Direct recognition of the
139. Sharma, A., Steichen, A.L.,
forms a functional receptor
mycobacterial glycolipid,
Jondle, C.N., Mishra, B.B. &
complex with MCL and
trehalose dimycolate, by C-type
Sharma, J. Protective role of
FcepsilonRI-gamma. Eur. J.
lectin Mincle. J. Exp. Med. 206,
Mincle in bacterial pneumonia
Immunol. (2013).
by regulation of neutrophil
mediated phagocytosis and
144. Suzuki, Y., Nakano, Y.,
135. Schoenen, H., Bodendorfer,
extracellular trap formation. J.
Mishiro, K., Takagi, T. et al.
B., Hitchens, K., Manzanero, S. et
Infect. Dis. (2013).
Involvement of Mincle and Syk
al. Cutting edge: Mincle is
in the changes to innate
essential for recognition and
140. Furukawa, A.,
immunity after ischemic stroke.
adjuvanticity of the mycobacte-
Kamishikiryo, J., Mori, D.,
Sci. Rep. 3, 3177 (2013).
rial cord factor and its synthetic
Toyonaga, K. et al. Structural
analog trehalose-dibehenate. J.
analysis for glycolipid
145. Sancho, D. & Reis, E.S.C.
Immunol. 184, 2756-2760
recognition by the C-type
Sensing of cell death by
(2010).
136. Desel, C., Werninghaus, K.,
lectins Mincle and MCL. Proc.
myeloid C-type lectin receptors.
Natl. Acad. Sci. U.S.A. 110,
Curr. Opin. Immunol. (2013).
17438-17443 (2013).
146. Bourgeois, C., Majer, O.,
Ritter, M., Jozefowski, K. et al.
The Mincle-activating adjuvant
141. Feinberg, H., Jegouzo, S.A.,
Frohner, I.E., Lesiak-Markowicz,
TDB induces MyD88-
Rowntree, T.J., Guan, Y. et al.
I. et al. Conventional dendritic
dependent Th1 and Th17
Mechanism for recognition of
cells mount a type I IFN
responses through IL-1R
an unusual mycobacterial
response against Candida spp.
signaling. PLoS One 8, e53531
glycolipid by the macrophage
requiring novel phagosomal
receptor mincle. J. Biol. Chem.
TLR7-mediated IFN-beta
288, 28457-28465 (2013).
signaling. J. Immunol. 186,
(2013).
137. Heitmann, L., Schoenen, H.,
3104-3112 (2011).
Ehlers, S., Lang, R. & Holscher, C.
142. Miyake, Y., Toyonaga, K.,
Mincle is not essential for
Mori, D., Kakuta, S. et al. C-type
147. Biondo, C., Signorino, G.,
controlling Mycobacterium
lectin MCL is an FcRgamma-
Costa, A., Midiri, A. et al.
tuberculosis infection.
coupled receptor that mediates
Recognition of yeast nucleic
Immunobiology 218, 506-516
the adjuvanticity of mycobacte-
acids triggers a host-protective
(2013).
type I interferon response. Eur.
one.
2879-2888 (2009).
48
Chapter one
J. Immunol. 41, 1969-1979
globulin production. J. Exp. Med. 161. Palm, N.W. & Medzhitov, R.
(2011).
192, 1545-1552 (2000).
Pattern recognition receptors
and control of adaptive
148. del Fresno, C., Soulat, D.,
153. O’Shea, J.J. & Paul, W.E.
immunity. Immunol. Rev. 227,
Roth, S., Blazek, K. et al.
Mechanisms underlying
221-233 (2009).
Interferon-beta production via
lineage commitment and
162. Kawai, T. & Akira, S. Toll-like
Dectin-1-Syk-IRF5 signaling in
plasticity of helper CD4+ T cells.
dendritic cells is crucial for
Science 327, 1098-1102 (2010).
receptors and their crosstalk
immunity to C. albicans.
154. Reiner, S.L. Development in
with other innate receptors in
Immunity 38, 1176-1186 (2013).
motion: helper T cells at work.
Infect. Immun.. Immunity 34,
Cell 129, 33-36 (2007).
637-650 (2011).
Kumar, V., Johnson, M.D. et al.
155. Abbas, A.K., Murphy, K.M. &
163. Drickamer, K. C-type
Functional genomics identifies
Sher, A. Functional diversity of
lectin-like domains. Curr. Opin.
type I interferon pathway as
helper T lymphocytes. Nature
Struct. Biol. 9, 585-590 (1999).
central for host defense against
383, 787-793 (1996).
149. Smeekens, S.P., Ng, A.,
164. Zhang, J.G., Czabotar, P.E.,
Candida albicans. Nat.
Commun. 4, 1342 (2013).
156. Sakaguchi, S., Miyara, M.,
Policheni, A.N., Caminschi, I. et
Costantino, C.M. & Hafler, D.A.
al. The dendritic cell receptor
150. Mosmann, T.R., Cherwinski,
FOXP3+ regulatory T cells in
Clec9A binds damaged cells via
H., Bond, M.W., Giedlin, M.A. &
the human immune system.
exposed actin filaments.
Coffman, R.L. Two types of
Nat. Rev. Immunol. 10, 490-500
Immunity 36, 646-657 (2012).
murine helper T cell clone. I.
(2010).
165. Ahrens, S., Zelenay, S.,
Definition according to profiles
of lymphokine activities and
157. Crotty, S. Follicular helper
Sancho, D., Hanc, P. et al. F-actin
secreted proteins. J. Immunol.
CD4 T cells (TFH). Annu. Rev.
is an evolutionarily conserved
136, 2348-2357 (1986).
Immunol. 29, 621-663 (2011).
damage-associated molecular
151. Harrington, L.E., Hatton,
158. Zhang, N. & Bevan, M.J.
a receptor for dead cells.
Immunity 36, 635-645 (2012).
pattern recognized by DNGR-1,
R.D., Mangan, P.R., Turner, H. et
CD8(+) T cells: foot soldiers of
al. Interleukin 17-producing
the immune system. Immunity
CD4+ effector T cells develop
35, 161-168 (2011).
via a lineage distinct from the T
The C-type lectin-like domain
helper type 1 and 2 lineages.
159. Pepper, M. & Jenkins, M.K.
superfamily. FEBS J. 272,
Nat. Immunol. 6, 1123-1132
Origins of CD4(+) effector and
6179-6217 (2005).
(2005).
central memory T cells. Nat.
Immunol. 12, 467-471 (2011).
152. Breitfeld, D., Ohl, L.,
Kremmer, E., Ellwart, J. et al.
Introduction
166. Zelensky, A.N. & Gready, J.E.
167. Wen, H., Miao, E.A. & Ting,
J.P. Mechanisms of NOD-like
160. Bevan, M.J. Understand
receptor-associated inflam-
Follicular B helper T cells
memory, design better
masome activation. Immunity
express CXC chemokine
vaccines. Nat. Immunol. 12,
39, 432-441 (2013).
receptor 5, localize to B cell
463-465 (2011).
follicles, and support immuno-
C-type lectin receptors orchestrate antifungal immunity
49
168. Loo, Y.M. & Gale, M., Jr.
Immune signaling by RIG-I-like
receptors. Immunity 34,
680-692 (2011).
169. Broz, P. & Monack, D.M.
Newly described pattern
recognition receptors team up
against intracellular pathogens.
Nat. Rev. Immunol. 13, 551-565
(2013).
170. Sun, L., Deng, L., Ea, C.K., Xia,
Z.P. & Chen, Z.J. The TRAF6
ubiquitin ligase and TAK1
kinase mediate IKK activation
by BCL10 and MALT1 in T
lymphocytes. Mol. Cell 14,
one.
289-301 (2004).