Download JB Review Featured Article - Oxford Academic

J. Biochem. 2013;154(6):491–499
doi:10.1093/jb/mvt099
JB Review
How do cells optimize luminal environments of endosomes/
lysosomes for efficient inflammatory responses?
Received September 19, 2013; accepted October 23, 2013; published online October 31, 2013
Department of Molecular Immunology and Inflammation, Research
Institute, National Center for Global Health and Medicine, 1-21-1
Toyama, Shinjuku-ku, Tokyo 162-8655, Japan
*Noriko Toyama-Sorimachi, Department of Molecular
Immunology and Inflammation, Research Institute, National Center
for Global Health and Medicine, 1-21-1 Toyama, Shinjuku-ku,
Tokyo 162-8655, Japan. Tel: þ81-3-3202-7181,
Fax: þ81-3-3202-7364, email: [email protected]
The endosome/lysosome compartments play pivotal
roles in immune cell functions as signalling platforms.
These intracellular compartments can efficiently restrict
the localization of signalling complexes and temporally
regulate signalling events to produce qualitatively
different outcomes. Immune cells also exploit the endosome/lysosome system for signal transduction and
intercellular communication to elicit immune responses.
Antigen-presenting cells such as dendritic cells and
macrophages take up pathogens by endocytosis and
prepare antigens via the endosome/lysosome system.
At the same time, pathogen-derived DNA and RNA
are recognized by immune sensors at the endosome/
lysosome compartments, which transmit signals to
induce immune responses. Recent studies revealed the
importance of controlling the endosomal/lysosomal environment for eliciting efficient signalling events at the
endosomes/lysosomes. Many factors including pH,
membrane potential, amino acid concentrations and
lipid composition are finely tuned at the endosome/lysosome compartments, and dysregulation of these factors
greatly affect immune cell functions. Redox-related
molecules and various types of transporters are involved
in the control of endosomal/lysosomal environment and
could be good therapeutic targets for treating autoimmune diseases.
Keywords: dendritic cells/inflammation/signalling
endosome/toll-like receptor/solute carrier
protein15A4.
Abbreviations: APC, antigen-presenting cells; BCR, B
cell receptors; CpG-ODN, CpG oligo DNA; DC,
dendritic cell; EGFR, epidermal growth factor
receptor; ENU, N-ethyl-N-nitrosourea; ER, endoplasmic reticulum; GPCR, G protein-coupled receptor; IBD, inflammatory bowel disease; iE-DAP, g-Dglutamyl-meso-diaminopimelic acid; IFN-b, interferon-b; IL-6, interleukin-6; LCMV, lymphocytic
choriomeningitis virus; LPS, lipopolysaccharide;
MAPK, mitogen-activated protein kinase; MDP,
muramyl dipeptide; MHC, major histocompatibility
complex; NADPH, reduced nicotinamide adenine
dinucleotide phosphate; NALP3, NACHT, LRR and
PYD domains-containing protein 3; NF-kB, nuclear
factor-kappa B; NOD, nucleotide-binding oligomerization domain; NOX2, NADPH oxidase 2; NGFR,
nerve growth factor receptor; PGN, peptidoglycan;
POT, proton-coupled oligopeptide transporters; SLC,
solute carrier family; SLE, systemic lupus erythematosus; TIRAP, toll/interleukin-1 receptor domaincontaining adapter protein; TLR, Toll-like receptors;
TNFa, tumor necrosis factor alpha; TRAM, translocation associated membrane protein; TRIF, TIRdomain-containing adapter-inducing interferon-b.
Endocytosis is a universal cellular activity to uptake
various materials including nutrition, pathogens and
cell surface receptors into cells. Until a decade ago,
endocytosis of cell surface receptors and the following
lysosomal degradation process were thought to be important regulatory mechanisms for down-modulation
and termination of signalling by cell surface receptors.
Today, endosomes/lysosomes also sometimes play important roles as signalling platforms (1—3). In these
cases, after a surface receptor binds to its ligand, the
receptor—ligand complex is internalized and continuously promotes downstream signalling events via signalling complexes that are spatially restricted to
particular compartments in the cell. Such ‘signalling
endosomes’ can efficiently regulate the localization of
signalling complexes via transportation and enable
particular signalling activities to be sustained for long
periods. Once activated, many receptors, such as epidermal growth factor receptor (EGFR), nerve growth
factor receptor (NGFR) (TrkA), and G proteincoupled receptor (GPCR), use this system (2, 4—6).
In parallel, a growing body of evidence has demonstrated that lysosomes, too, can function as important
signalling platforms and are not only compartments
used for degradation. Cellular nutritional conditions
as well as the metabolic state are sensed at lysosomal
compartments, and signals are transmitted at the lysosomal compartments to induce various gene transcriptions (7—9). Thus, the endosome/lysosome system
plays key roles in various cellular functions.
Immune cells also exploit the signal transduction capacity of the endosome/lysosome system for eliciting
immune responses (10, 11). In particular, some
immune cells, such as dendritic cells (DCs), macrophages, and B cells, take up various antigens, recognize
danger-associated molecular patterns at the endosomes/
lysosomes, and transmit signals from these compartments to produce immune responses (12, 13). Because
ß The Authors 2013. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved
491
Featured Article
Toshihiko Kobayashi, Tsubasa Tanaka and
Noriko Toyama-Sorimachi*
T. Kobayashi et al.
these compartments can restrict signals spatially and
temporally to elicit qualitatively different outcomes,
understanding the molecular bases of endosome/lysosome-dependent signalling events could be helpful for
developing strategies to control immune cell functions.
Recent studies have revealed that efficient signalling at
these compartments is achieved by controlling the endosomal/lysosomal environment. In this review, we highlight the endosome/lysosome-dependent signalling
events and the unique regulation of the endosomal/
lysosomal environment in immune cells.
Endosomes/Lysosomes Spatiotemporally
Sort TLR4 signalling Components to Induce
Immune Responses
Toll-like receptors (TLRs) are well-characterized
innate immune sensors that play a key role in host
defense (14). They recognize conserved microbial components at the cell surface, or in intracellular compartments such as endosomes/lysosomes, and induce
immune responses (14, 15). At the early stage of inflammation, antigen-presenting cells (APCs), such as
DCs and macrophages, that encounter microbes or
their components are activated through TLRs to produce inflammatory and/or immunomodulatory cytokines, including tumor necrosis factor a (TNFa),
interleukin-6 (IL-6) and type I interferon. At the
same time, the TLR-mediated activation of these
cells leads to a transcription of a panel of genes that
are pivotal for the induction of acquired immune responses following antigen presentation. Importantly,
depending on the type of TLRs triggered, APCs produce different cytokines and induce specific types of
effector T cells that are suitable to eliminate the infecting microbes; that is, which TLRs are triggered determines the type of immune response elicited (16, 17).
TLR4 recognizes lipopolysaccharide (LPS) on gramnegative bacteria at the cell surface and activates two
qualitatively distinct signalling pathways, MyD88dependent and MyD88-independent, at the cell surface
and endosomes, respectively (Fig. 1A) (18, 19). After
LPS binds to TLR4 at the cell surface, the MyD88dependent pathway is activated through the adaptor
proteins TIRAP (toll/interleukin-1 receptor domaincontaining adapter protein) and MyD88, which in
turn activate the transcription factor NF-kB (nuclear
factor-kappa B) and mitogen-activated protein kinase
(MAPK), culminating in the production of pro-inflammatory cytokines such as IL-6 and TNFa.
Subsequently, TLR4 is internalized, the adaptor proteins TRAM (translocation associated membrane protein) and TIR-domain-containing adapter-inducing
interferon-b (TRIF) replace TIRAP and MyD88, and
the MyD88-independent pathway is consequently
initiated at the endosomes. Its activation results in
the generation of interferon-b (IFN-b) and IL-10.
The transition from the TIRAP-MyD88 pathway to
the TRAM-TRIF pathway downstream of TLR4 is
mediated by PI3 kinase-dependent phosphatidylinositol turnover. TIRAP binds PtdIns(4,5)P2 through its
amino-terminal polybasic domain and is localized to
492
PtdIns(4,5)P2-rich
plasma
membrane
regions.
Conversion of PtdIns(4,5)P2 to PtdIns(3,4,5)P3 by
PI3 kinase p110d causes the release of TIRAP into
the cytoplasm, and then TRAM is recruited to
TLR4. These sequential events at the distinct compartments are pivotal for the efficient initiation and termination of inflammatory responses, by shifting the
cytokines released from pro-inflammatory to the antiinflammatory ones. Indeed, PI3Kd-mutant mice were
more susceptible to endotoxic shock than wild-type
controls because of sustained production of TNF,
IL-6 and IL-12 but restrained production of IL-10
and IFN-b (20). Thus, the compartmentalization of
signalling complexes at endosomes confers spatiotemporal properties on TLR4-mediated signalling and is
an important mechanism for mounting an effective
and harmless immune response.
Endosome/Lysosome-Dependent
Recognition of Nucleic Acids by TLRs and
Its Importance in Immune Defense
Some TLRs, including TLR3, 7 and 9, localize to
endosomes, where they recognize nucleic acids and
transmit signals to induce various cytokines (21, 22).
The manner by which nucleic acids are recognized by
these endosomal TLRs appears to enable selective recognition of ingested microbe-derived nucleic acids
while avoiding undesirable responses to self-tissuederived nucleotides in the serum. TLR9 recognizes
unmethylated CpG oligo DNA (CpG-ODN) at the
endosomes and induces pro-inflammatory cytokines
and type I IFN production, which is essential for antiviral immune responses (Fig. 1B) (23). In this process,
endoplasmic reticulum (ER)-resident TLR9 is transported to endosomes, where it binds CpG-ODN, and
then passes to specialized lysosome-related compartments (24). This TLR9 trafficking is crucial for generating functionally active TLR9, given that the limited
proteolysis of TLR9 by lysosomal proteases such as
cathepsins is necessary for CpG-ODN recognition
(25, 26). These lysosomal proteases are most active
under acidic conditions, and therefore the acidification
of endosome/lysosome compartments during their
maturation is important for triggering inflammatory
responses. Intriguingly, the trafficking of TLR9/
CpG-ODN from endosomes to lysosomes is affected
by the CpG-ODN structure, with different structures
leading to different cellular responses (27). The precise
molecular mechanism by which such traffickingcoupled signalling events are spatially and temporally
regulated at the endosomal/lysosomal compartments is
not fully understood.
Endosomes/Lysosomes as Sites for
BCR-TLR Collaboration and
Autoimmune-Response Induction
B cell receptors (BCRs) are composed of a membraneassociated form of immunoglobulin (Ig) that is noncovalently associated with a disulphide-linked heterodimer composed of two integral membrane proteins,
Endosomal/lysosomal signalling and inflammation
Fig. 1 Examples of endosome/lysosome-dependent signalling events in the immune system. (A) Model for endosome-dependent TLR4 signalling.
At the plasma membrane, LPS-bound TLR4 binds to the sorting adaptor TIRAP, which is anchored with the plasma membrane lipid PI (4,5)
P2. TIRAP then recruits the signalling adaptor MyD88, and MyD88-dependent signalling induces the production of proinflammatory cytokines
including TNFa, IL-6 and IL-12. LPS induces recruitment of the PI3 kinase p110d to the TLR4-TIRAP-MyD88 complex, and p110d then
decreases the amount of PI(4,5)P2, which probably causes TIRAP to be released from the membrane and promotes the switch to endosomal
TRAM signalling. The released TIRAP and MyD88 are degraded through a proteasome-dependent process. This subcellular shuttling of TLR4
results in a cytokine shift from proinflammatory to anti-inflammatory. (B) Recognition of nucleic acids by TLR7 or TLR9 in endosomes. Some
TLRs that sense nucleotides are localized to endosomes and transduce signals to elicit inflammatory responses. TLR7 and TLR9 recognize
single-strand RNA and unmethylated CpG-ODN, respectively, when they enter endolysosomes. These TLRs are localized to the ER in the
steady state and then relocate to endosomes in an AP3- and Unc93B-dependent manner. In endolysosomes, these TLRs are cleaved by
cathepsins, which confers ligand-binding ability on them. The environmental control of the endolysosomes and the proper trafficking and
subcellular localization of the TLRs are required to induce inflammatory responses. (C) Essential role of endocytosis in regulating the outcome of
BCR signalling. The endocytosis and subcellular localization of the BCR following its activation are necessary for BCR-mediated kinase
activation and the downstream gene transcription. BCRs are internalized after cross-linking with antigens, and they traffic from early endosomes
to late endosomes (LAMP-1þ compartments). Lyn and Syk are phosphorylated at the cell surface. The phosphorylation of MAP kinases occurs
at the endosomes, and the phosphorylation of Akt and its downstream targets, resulting from internalized BCRs, occurs at the late endosomes.
Importantly, inhibiting BCR’s endocytosis alters the amount of kinase phosphorylation in the BCR signalling pathway and its outcome.
CD79a and CD79b (28). After ligating with its antigen,
the BCR initiates signalling at the plasma membrane,
which involves the phosphorylation of BCR components by Src family kinases and the recruitment of a
proximal kinase, Syk. The BCR is then internalized
and induces the sequential phosphorylation of
MAPKs and the kinase Akt (Fig. 1C) (29). The inhibition of BCR endocytosis dysregulates these phosphorylation events, leading to gene-transcription defects.
Thus, proper trafficking of the BCR into endosomes
is required for BCR signalling and appropriate
immune responses.
Furthermore, endosomal signalling via BCRs is a
pivotal event for induction of autoimmune response.
Autoantigens include self-DNA, histones, RNA or
ribonucleoproteins, which are thought to be released
from apoptotic cells. The endosomal localization of the
BCRs allows them to engage synergistically with TLR9
in the presence of DNA-containing antigens; this process has been implicated in inducing the activation of
autoimmune B cells (30). In fact, the simultaneous engagement of BCRs with TLR9 and their endosomal
activation contributes to the production of DNAspecific autoantibodies in mouse models of lupus
(30, 31). Thus, signalling endosomes also serve as a
meeting place for multiple signalling pathways
mediated by different immune sensors.
Importance of Controlling the
Endosome/Lysosome Environment in
Inflammatory Cells
Endosomes are gradually acidified by V-ATPase as
they mature (32). As described earlier, the acidic environment of endosomes is important for the cathepsinmediated limited proteolysis of TLR9, which converts
this receptor into its ligand-accessible form. In addition, the binding affinity of CpG-ODN to TLR9
itself is affected by the pH (33). At neutral pH,
TLR9’s binding to CpG-DNA is weak, but lowering
the pH leads to a strong TLR9-CpG-DNA interaction.
Treatment with compounds that interfere with luminal
acidification of endosomes/lysosomes, such as bafilomycin A1 (a specific inhibitor of the vacuolar proton
pump V-ATPase) or chloroquine (a weak base that can
partition into vesicles and increase the pH) impairs the
CpG-DNA-driven signalling via TLR9 (34—36). Thus,
the optimization of luminal pH is an important regulatory mechanism for efficient signalling through
TLR9. From this viewpoint, factors that control the
‘luminal’ pH in immune cells could be useful as targets
for anti-inflammatory drugs to treat autoimmune
diseases. In fact, chloroquine and its derivatives have
already been used to effectively treat autoimmune diseases such as rheumatoid arthritis and systemic lupus
493
T. Kobayashi et al.
erythematosus (SLE) (37, 38). The mechanism underlying chloroquine’s action in autoimmune diseases is
not fully understood, but findings in a murine model
of SLE suggest that it blocks the TLR9-dependent,
chromatin-antibody complex-induced stimulation of
self-reactive B cells (39, 40).
Endosomes finally fuse to lysosomes, which contain
numerous hydrolases for the degradation of proteins,
lipids, and other biomolecules. In lysosomes, these
molecules are completely degraded into their building
blocks, which are recycled for biosynthesis of macromolecules. An impairment in the lysosomes’ degradative function leads to the accumulation of undegraded
molecules, which causes various types of lysosomal
storage diseases (41, 42). DCs, the most potent
APCs, however, limit the lysosomes’ proteolytic activity for antigen presentation to T cells. DCs ingest protein-containing macromolecules and process them into
short peptides, which they present as an antigen together with major histocompatibility complex (MHC)
molecules, a critical event for the induction of the
adaptive immune response (43). To generate peptides
with a certain length and to maximize half-life of the
peptides within lysosomes, DCs possess a unique
mechanism for phagosome maturation. DCs maintain
the phagosomal pH between 7 and 7.5 in the first few
hours after phagocytosis (44). DCs recruit a component of nicotinamide adenine dinucleotide phosphate
(NADPH) oxidase NOX2 to the phagosomes in
Rab27a-dependent fashion (45), where oxygen radicals
generated by the NADPH oxidase cause proton consumption and neutralize the acidic environment, counteracting the actions of the proton pump V-ATPase
(44, 46). Because of this phagosomal alkalization machinery, the phagolysosomal pH still remained above
pH 7 after fusing to the lysosomes. This makes it possible for peptides to be processed to the appropriate
size for loading onto MHC molecules by preventing
the complete degradation of peptides (44). This mechanism is particularly important for antigen processing
for MHC class I, called antigen cross-presentation, in
which exogenously ingested antigens are presented together with MHC class I to activate CD8þ cytotoxic T
cells (47).
The alkalized phagolysosomes should be finally
acidified to hydrolyze contents. Recent study further
improved our understanding on how the neutralized
phagolysosomes are safely acidified to fulfil the obligation of lysosomes as degradative compartments (48).
On the surface of phagosomes containing grampositive bacteria, caspase-1 activated via NACHT,
LRR and PYD domains-containing protein 3
(NALP3) inflammasome accumulates and induces
phagosomal acidification by negatively regulating
NADPH oxidase NOX2 activity. An important observation is that phagosomes containing gram-negative
bacteria did not recruit caspase-1 and appeared to be
acidified via different machinery. This implies that the
phagosomal environment is differentially controlled by
bacterial species. Thus environmental regulation
within phagosomes/phagolysosomes is mediated by
multiple complex processes depending on different
cargos.
494
A New Player in Endosomal/Lysosomal
Environmental Control
Immune cells possess a multilayered system for controlling the endosomal/lysosomal environment, involving not only the proton concentration but also the
concentrations of other ions and substances (Fig. 2).
We and another group recently discovered that the
lysosome-resident amino acid transporter, solute carrier family 15A4 (SLC15A4), is critical for signal transduction by TLR7 and TLR9 (Fig. 3) (49, 50). Cellular
transporters are currently categorized into two large
groups: the SLC family, which functions independently
of ATP, and the ATP-binding cassette family, which is
ATP-dependent. SLC15A4 is a member of the SLC
family, which includes roughly 300 genes to date, and
is further divided into many subfamilies (51).
SLC15A4 belongs to the SLC15 subfamily, which
has five members, A1 to A5, and is well conserved
across species (52, 53). These are so-called protoncoupled oligopeptide transporters (POTs) and carry
amino acids and oligopeptides together with a proton
as a common feature, but their functions and expression patterns differ. A1 and A2 are mainly expressed in
the small intestine, liver and kidney, whereas A3 and
A4 have low sequence similarity with A1 and A2 and
are expressed in immune cells and the nervous system
(the localization of A5 is currently unknown).
Moreover, both A1 and A2 have low substrate specificity and the ability to carry 600 different di-peptides
and 8,000 tri-peptides, whereas A3 and A4 transport
only histidine and several histidine-containing di- or
tri-peptides (Fig. 3A) (51). Because of their expression
pattern and substrate specificity, A1 and A2 are
thought to function in the absorption or reabsorption
of digested nutrients. By contrast, A3 and A4 appear
to have a role in immune regulation; however, the
functional importance of these transporters was not
investigated until recently.
The transporter activity of A4 is pH dependent, with
higher transport of histidine/oligopeptides and protons
at pH 5.5 than pH 7.0 (Fig. 3B) (53). Therefore, the
transporter activity of SLC15A4 is maximized in the
acidic environment of lysosomes. Several studies have
demonstrated an association between SLC15A4 function and certain inflammatory disorders, including diabetes and inflammatory bowel disease (IBD) (54, 55).
Importantly, a recent genome-wide association study
identified SLC15A4 as an SLE-susceptibility gene (56),
suggesting a link between the function of SLC15A4
and autoimmune disease in humans. An attractive hypothesis emerging from these observations is that
SLC15A4 contributes to the regulation of inflammatory responses by regulating lysosomal functions or
environments.
We tested this hypothesis using SLC15A4 knockout
mice and found that SLC15A4-deficient DCs showed
an impaired TLR9-mediated production of cytokines
such as IFN-b, IL-12 and IL-15 (50). Consistent with
this finding, SLC15A4-knockout mice exhibited a less
severe form of colitis than wild-type mice. Beutler and
his colleagues also demonstrated using N-ethyl-Nnitrosourea (ENU)-induced SLC15A4 mutant mice
Endosomal/lysosomal signalling and inflammation
Fig. 2 Tuning of the endosomal/lysosomal environment in immune cells. The endosomal/lysosomal compartments play pivotal roles in immune
cell functions as signalling platforms. In addition to the TLRs and BCR mentioned in the text, other important receptor systems such as
chemokine receptors and cytokine receptors use such compartments for their signalling. The ‘luminal’ environments of endosomes/lysosomes are
strictly regulated by various transporters and redox-related molecules, such as V-ATPase, NOX2 and SLC15A4, to optimize endosome/lysosome-dependent signalling events. In addition to these players, phosphatidylinositol turnover is important for generating spatial and temporal
properties of signalling molecules. The lysosomal system in immune cells is also finely tuned to efficiently generate antigen peptides. Phagosomal
alkalization through NOX2-mediated reactive oxygen production limits the vesicles’ proteolytic capacity, thereby preserving peptides from
complete degradation and making it possible to load peptides of the appropriate size onto MHC molecules.
that SLC15A4 plays a crucial role in regulation of the
functions of TLR7 as well as TLR9, which is necessary
for recognizing single-stranded RNA (Fig. 3C).
SLC15A4-mutant mice is highly susceptible to influenza virus infection (49). SLC15A4 is also required
to protect mice from infection by mouse lymphocytic
choriomeningitis virus, in which the DCs prime T cells
in an SLC15A4-dependent manner to clear the virus
(57).
How does SLC15A4 contribute to TLR7- or TLR9mediated signalling? Considering that SLC15A4’s
function is to export histidines/oligopeptides together
with protons from these compartments to the cytosol,
one possibility is that the loss of this transporter substantially affects the proton concentration of the endolysosome. Another possibility is that the loss of this
transporter causes an abnormal histidine concentration in endolysosomes, eventually disrupting the histidine homeostasis. Histidine has an ionizable imidazole
ring in its side chain and the ability, unique among
amino acids, to function as an acid—base catalyst.
The pKa value for the imidazole ring is 7.0. In fact,
histidine’s buffering capacity is so high that it is used as
a buffer in histidine-tryptophan-ketoglutarate solution, which is used for organ preservation. Therefore,
in the course of endolysosomal acidification, too much
histidines within endolysosomes may affect endolysosomal pH. This idea is supported by the observation
that histidine administration in mice leads to the amelioration of inflammatory responses (50). We also
showed that free histidine, but not alanine, in a certain
concentration range can inhibit the enzymatic activities
of cathepsins B and L, both of which are important for
TLR9’s processing and activation in lysosomes (25,
26). Because the addition of exogenous histidine did
not exacerbate the impaired TLR9-mediated cytokine
production of DCs lacking SLC15A4, the endolysosomal histidine homeostasis of these cells may already
have been disrupted.
The precise mechanism for the suppression of inflammatory responses in the absence of this transporter needs to be clarified. Besides DCs, we revealed
that B cells also express SLC15A4 and B cell’s
SLC15A4 is crucial for autoimmune symptom (manuscript under revision). Nevertheless, the finding that
disruption of a lysosome-resident transporter greatly
affects lysosome-dependent immunological events revealed the importance of controlling the environment
of endolysosome compartments for adequate immune
response induction.
Another important finding obtained from SLC15A4knockout mice is that SLC15A4 is responsible for transporting a certain type of microbial component from
inside the lysosomes to the cytosol for initiating cytokine production and inflammatory responses (50). The
bacterial cell wall peptidoglycan (PGN) contains unique
495
T. Kobayashi et al.
Fig. 3 Roles of the lysosomal histidine/oligopeptide transporter, SLC15A4, in innate immunity. (A, B) SLC15A4 contains 12 transmembrane
domains, with both the N- and C-termini facing the cytosol, and transports histidines/oligopeptides uphill using proton-motive force. The
transporter activity of SLC15A4 is pH dependent. In mature lysosomes, histidines/oligopeptides derived from lysosomal degradation products
are actively transported out by SLC15A4. (C) The ligand binding and signalling of TLR9 are strictly regulated by the lysosomal state and
functions. Histidine in lysosomes might be maintained within a defined concentration, which contributes to the maximal functional efficiency of
lysosomal components, including TLR9. SLC15A4 deficiency causes homeostasis failure of the lysosomal environment and disrupts TLR9
function. Furthermore, SLC15A4 is responsible for transporting the peptide glycan-like NOD1 ligand, tri-DAP, from lysosomes to the cytosol,
and induces IL-1b production to eliminate bacterial invaders. Thus, this lysosomal transporter also functions as a receiving clerk for pathogenic
components.
structure such as g-D-glutamyl-meso-diaminopimelic
acid (iE-DAP) or muramyl dipeptide (MDP). These
components are recognized by cytoplasmic pathogen
sensors to initiate the activation of NF-kB and MAP
kinases and the induction of innate immune responses.
Nucleotide-binding oligomerization domain (NOD) 1
and NOD2 are well characterized cytoplasmic sensors
and recognize iE-DAP and MDP, respectively (58). The
importance of these molecules is underscored by their
genetic association with IBD (59). Although both
NOD1 and NOD2 detect PGN components in the cytosol, how such microbially derived components reach the
host cytosol is still unclear. SLC15A4-knockout mice
exhibit a severe defect in NOD1-dependent IL-1b production in vivo, which is caused by the unresponsiveness
of SLC15A4-deficient DCs and macrophages to triDAP (Fig. 3C) (50). Thus, this lysosomal transporter
also functions as a receiving clerk for pathogenic components. Because the lysosomes are the site at which
phagocytosed pathogens are digested, the transport of
the digestion products by a pH-dependent lysosomal
transporter seems to be a reasonable and safe way for
the cell to monitor pathogen invasion.
496
Conclusion and Perspectives
In summary, the endosome/lysosome compartments
play pivotal roles in immune cell functions as signalling
platforms. Their environments are strictly regulated as
the occasion demands by various transporters and
redox-related molecules such as V-ATPase, NOX2
and SLC15A4 to optimize endosome/lysosomedependent signalling events. The endosome/lysosome
system is an attractive target for controlling immune
diseases, although our understanding of it is still
quite primitive. Although not discussed here in
detail, certain immune cells, such as natural killer
cells and cytotoxic T cells, develop specialized lysosome-related organelles, called secretory lysosomes,
which contain cytotoxic components including granzymes and perforin and are essential for the cells’ cytotoxic function. Thus, the environmental control of
secretory lysosomes is another interesting and important issue.
Because lysosome functions are closely related to
immune cell functions, further investigations of the
endolysosome system should reveal novel regulatory
Endosomal/lysosomal signalling and inflammation
mechanisms for the immune system. We still have
more to learn about which molecules control the signalling-competent endolysosomal environment, and
how and when such molecules are recruited to and
regulate compartments. To better understand the molecular bases of these processes, a comprehensive
understanding of dynamic organization of proteins
and lipid species at the endosomes/lysosomes is
necessary.
Funding
This work was supported by the Funding Program for Next
Generation World-Leading Researchers (Next Program) (for
N.T.-S., LS134), grants-in-aid for Scientific Research from the
Ministry of Education, Science, Sports and Culture of Japan (for
N.T.-S., 21390123, for T.K., 25871165), and a grant from the
National Center for Global Health and Medicine (for N.T.-S.,
23S001).
Conflict of Interest
None declared.
References
1. Platta, H.W. and Stenmark, H. (2011) Endocytosis and
signaling. Curr. Opin. Cell Biol. 23, 393—403
2. Murphy, J.E., Padilla, B.E., Hasdemir, B., Cottrell, G.S.,
and Bunnett, N.W. (2009) Endosomes: a legitimate platform for the signaling train. Proc. Natl. Acad. Sci. U S A.
106, 17615—17622
3. Hupalowska, A. and Miaczynska, M. (2012) The new
faces of endocytosis in signaling. Traffic 13, 9—18
4. Delcroix, J.D., Valletta, J.S., Wu, C., Hunt, S.J., Kowal,
A.S., and Mobley, W.C. (2003) NGF signaling in sensory
neurons: evidence that early endosomes carry NGF
retrograde signals. Neuron 39, 69—84
5. Wiley, H.S. and Burke, P.M. (2001) Regulation of receptor tyrosine kinase signaling by endocytic trafficking.
Traffic 2, 12—18
6. Irannejad, R., Tomshine, J.C., Tomshine, J.R., Chevalier,
M., Mahoney, J.P., Steyaert, J., Rasmussen, S.G.,
Sunahara, R.K., El-Samad, H., Huang, B., and von
Zastrow, M. (2013) Conformational biosensors reveal
GPCR signalling from endosomes. Nature 495, 534—538
7. Settembre, C., Zoncu, R., Medina, D.L., Vetrini, F.,
Erdin, S., Huynh, T., Ferron, M., Karsenty, G.,
Vellard, M.C., Facchinetti, V., Sabatini, D.M., and
Ballabio, A. (2012) A lysosome-to-nucleus signalling
mechanism senses and regulates the lysosome via
mTOR and TFEB. EMBO J. 31, 1095—1108
8. Sancak, Y., Bar-Peled, L., Zoncu, R., Markhard, A.L.,
Nada, S., and Sabatini, D.M. (2010) Ragulator-Rag complex targets mTORC1 to the lysosomal surface and is necessary for its activation by amino acids. Cell 141, 290—303
9. Cang, C., Zhou, Y., Navarro, B., Seo, Y.J., Aranda, K.,
Shi, L., Battaglia-Hsu, S., Nissim, I., Clapham, D.E.,
and Ren, D. (2013) mTOR regulates lysosomal ATP-sensitive two-pore Na(þ) channels to adapt to metabolic
state. Cell 152, 778—790
10. Watts, C. (2012) The endosome-lysosome pathway and
information generation in the immune system. Biochim.
Biophys. Acta 1824, 14—21
11. Kagan, J.C. (2012) Signaling organelles of the innate
immune system. Cell 151, 1168—1178
12. Niedergang, F. and Chavrier, P. (2004) Signaling and
membrane dynamics during phagocytosis: many roads
lead to the phagos(R)ome. Curr. Opin. Cell Biol. 16,
422—428
13. Savina, A. and Amigorena, S. (2007) Phagocytosis and
antigen presentation in dendritic cells. Immunol. Rev.
219, 143—156
14. Kawai, T. and Akira, S. (2008) Toll-like receptor and RIGI-like receptor signaling. Ann. N Y Acad. Sci. 1143, 1—20
15. Gilliet, M., Cao, W., and Liu, Y.J. (2008) Plasmacytoid
dendritic cells: sensing nucleic acids in viral infection and
autoimmune diseases. Nat. Rev. Immunol. 8, 594—606
16. Dabbagh, K. and Lewis, D.B. (2003) Toll-like receptors
and T-helper-1/T-helper-2 responses. Curr. Opin. Infect.
Dis. 16, 199—204
17. Steinman, R.M. and Hemmi, H. (2006) Dendritic cells:
translating innate to adaptive immunity. Curr. Top.
Microbiol. Immunol. 311, 17—58
18. Kagan, J.C., Su, T., Horng, T., Chow, A., Akira, S., and
Medzhitov, R. (2008) TRAM couples endocytosis of
Toll-like receptor 4 to the induction of interferon-beta.
Nat. Immunol. 9, 361—368
19. Tanimura, N., Saitoh, S., Matsumoto, F., AkashiTakamura, S., and Miyake, K. (2008) Roles for LPSdependent interaction and relocation of TLR4 and
TRAM in TRIF-signaling. Biochem. Biophys. Res.
Commun. 368, 94—99
20. Aksoy, E., Taboubi, S., Torres, D., Delbauve, S.,
Hachani, A., Whitehead, M.A., Pearce, W.P.,
Berenjeno-Martin, I., Nock, G., Filloux, A., Beyaert, R.,
Flamand, V., and Vanhaesebroeck, B. (2012) The
p110delta isoform of the kinase PI(3)K controls the subcellular compartmentalization of TLR4 signaling and protects from endotoxic shock. Nat. Immunol. 13, 1045—1054
21. Blasius, A.L. and Beutler, B. (2010) Intracellular toll-like
receptors. Immunity 32, 305—315
22. Blasius, A.L. and Beutler, B. (2010) Intracellular toll-like
receptors. Immunity 32, 305—315
23. Kumagai, Y., Takeuchi, O., and Akira, S. (2008) TLR9
as a key receptor for the recognition of DNA. Adv. Drug
Deliv. Rev. 60, 795—804
24. Latz, E., Schoenemeyer, A., Visintin, A., Fitzgerald,
K.A., Monks, B.G., Knetter, C.F., Lien, E., Nilsen,
N.J., Espevik, T., and Golenbock, D.T. (2004) TLR9
signals after translocating from the ER to CpG DNA
in the lysosome. Nat. Immunol. 5, 190—198
25. Matsumoto, F., Saitoh, S., Fukui, R., Kobayashi, T.,
Tanimura, N., Konno, K., Kusumoto, Y., AkashiTakamura, S., and Miyake, K. (2008) Cathepsins are
required for Toll-like receptor 9 responses. Biochem.
Biophys. Res. Commun. 367, 693—699
26. Park, B., Brinkmann, M.M., Spooner, E., Lee, C.C.,
Kim, Y.M., and Ploegh, H.L. (2008) Proteolytic cleavage
in an endolysosomal compartment is required for activation of Toll-like receptor 9. Nat. Immunol. 9, 1407—1414
27. Honda, K., Ohba, Y., Yanai, H., Negishi, H., Mizutani,
T., Takaoka, A., Taya, C., and Taniguchi, T. (2005) Spatiotemporal regulation of MyD88-IRF-7 signalling for
robust type-I interferon induction. Nature 434,
1035—1040
28. Hsueh, R.C. and Scheuermann, R.H. (2000) Tyrosine
kinase activation in the decision between growth, differentiation, and death responses initiated from the B cell
antigen receptor. Adv. Immunol. 75, 283—316
29. Chaturvedi, A., Martz, R., Dorward, D., Waisberg, M.,
and Pierce, S.K. (2011) Endocytosed BCRs sequentially
497
T. Kobayashi et al.
30.
31.
32.
33.
34.
35.
36.
37.
38.
39.
40.
41.
42.
43.
44.
45.
498
regulate MAPK and Akt signaling pathways from intracellular compartments. Nat. Immunol. 12, 1119—1126
Chaturvedi, A., Dorward, D., and Pierce, S.K. (2008)
The B cell receptor governs the subcellular location of
Toll-like receptor 9 leading to hyperresponses to DNAcontaining antigens. Immunity 28, 799—809
Rawlings, D.J., Schwartz, M.A., Jackson, S.W., and
Meyer-Bahlburg, A. (2012) Integration of B cell responses through Toll-like receptors and antigen receptors. Nat. Rev. Immunol. 12, 282—294
Mindell, J.A. (2012) Lysosomal acidification mechanisms. Annu. Rev. Physiol. 74, 69—86
Rutz, M., Metzger, J., Gellert, T., Luppa, P., Lipford,
G.B., Wagner, H., and Bauer, S. (2004) Toll-like receptor
9 binds single-stranded CpG-DNA in a sequence- and
pH-dependent manner. Eur. J. Immunol. 34, 2541—2550
Hacker, H., Mischak, H., Miethke, T., Liptay, S.,
Schmid, R., Sparwasser, T., Heeg, K., Lipford, G.B.,
and Wagner, H. (1998) CpG-DNA-specific activation
of antigen-presenting cells requires stress kinase activity
and is preceded by non-specific endocytosis and endosomal maturation. EMBO J. 17, 6230—6240
Yi, A.K., Tuetken, R., Redford, T., Waldschmidt, M.,
Kirsch, J., and Krieg, A.M. (1998) CpG motifs in bacterial DNA activate leukocytes through the pH-dependent generation of reactive oxygen species. J. Immunol.
160, 4755—4761
Ahmad-Nejad, P., Hacker, H., Rutz, M., Bauer, S.,
Vabulas, R.M., and Wagner, H. (2002) Bacterial CpGDNA and lipopolysaccharides activate Toll-like receptors at distinct cellular compartments. Eur. J. Immunol.
32, 1958—1968
Katz, S.J. and Russell, A.S. (2011) Re-evaluation of antimalarials in treating rheumatic diseases: re-appreciation
and insights into new mechanisms of action. Curr. Opin.
Rheumatol. 23, 278—281
Lee, S.J., Silverman, E., and Bargman, J.M. (2011) The
role of antimalarial agents in the treatment of SLE and
lupus nephritis. Nat. Rev. Nephrol. 7, 718—729
Leadbetter, E.A., Rifkin, I.R., Hohlbaum, A.M.,
Beaudette, B.C., Shlomchik, M.J., and MarshakRothstein, A. (2002) Chromatin-IgG complexes activate
B cells by dual engagement of IgM and Toll-like receptors. Nature 416, 603—607
Viglianti, G.A., Lau, C.M., Hanley, T.M., Miko, B.A.,
Shlomchik, M.J., and Marshak-Rothstein, A. (2003)
Activation of autoreactive B cells by CpG dsDNA.
Immunity 19, 837—847
Platt, F.M., Boland, B., and van der Spoel, A.C. (2012)
The cell biology of disease: lysosomal storage disorders:
the cellular impact of lysosomal dysfunction. J. Cell Biol
199, 723—734
Schultz, M.L., Tecedor, L., Chang, M., and Davidson,
B.L. (2011) Clarifying lysosomal storage diseases. Trends
Neurosci. 34, 401—410
Neefjes, J., Jongsma, M.L., Paul, P., and Bakke, O.
(2011) Towards a systems understanding of MHC class
I and MHC class II antigen presentation. Nat. Rev.
Immunol. 11, 823—836
Savina, A., Jancic, C., Hugues, S., Guermonprez, P.,
Vargas, P., Moura, I.C., Lennon-Dumenil, A.M., Seabra,
M.C., Raposo, G., and Amigorena, S. (2006) NOX2 controls phagosomal pH to regulate antigen processing during
crosspresentation by dendritic cells. Cell 126, 205—218
Jancic, C., Savina, A., Wasmeier, C., Tolmachova, T.,
El-Benna, J., Dang, P.M., Pascolo, S., Gougerot-
46.
47.
48.
49.
50.
51.
52.
53.
54.
55.
56.
Pocidalo, M.A., Raposo, G., Seabra, M.C., and
Amigorena, S. (2007) Rab27a regulates phagosomal pH
and NADPH oxidase recruitment to dendritic cell
phagosomes. Nat. Cell Biol. 9, 367—378
Savina, A., Peres, A., Cebrian, I., Carmo, N., Moita, C.,
Hacohen, N., Moita, L.F., and Amigorena, S. (2009) The
small GTPase Rac2 controls phagosomal alkalinization
and antigen crosspresentation selectively in CD8(þ) dendritic cells. Immunity 30, 544—555
Ramachandra, L., Simmons, D., and Harding, C.V.
(2009) MHC molecules and microbial antigen processing
in phagosomes. Curr. Opin. Immunol. 21, 98—104
Sokolovska, A., Becker, C.E., Ip, W.K., Rathinam,
V.A., Brudner, M., Paquette, N., Tanne, A., Vanaja,
S.K., Moore, K.J., Fitzgerald, K.A., Lacy-Hulbert, A.,
and Stuart, L.M. Activation of caspase-1 by the NLRP3
inflammasome regulates the NADPH oxidase NOX2 to
control phagosome function. Nat. Immunol. 14, 543—553
Blasius, A.L., Arnold, C.N., Georgel, P., Rutschmann,
S., Xia, Y., Lin, P., Ross, C., Li, X., Smart, N.G., and
Beutler, B. (2010) Slc15a4, AP-3, and HermanskyPudlak syndrome proteins are required for Toll-like receptor signaling in plasmacytoid dendritic cells. Proc.
Natl. Acad. Sci. U S A. 107, 19973—19978
Sasawatari, S., Okamura, T., Kasumi, E., TanakaFuruyama, K., Yanobu-Takanashi, R., Shirasawa, S.,
Kato, N., and Toyama-Sorimachi, N. (2011) The solute
carrier family 15A4 regulates TLR9 and NOD1 functions in the innate immune system and promotes colitis
in mice. Gastroenterology 140, 1513—1525
Yamashita, T., Shimada, S., Guo, W., Sato, K.,
Kohmura, E., Hayakawa, T., Takagi, T., and
Tohyama, M. (1997) Cloning and functional expression
of a brain peptide/histidine transporter. J. Biol. Chem.
272, 10205—10211
Daniel, H. and Kottra, G. (2004) The proton oligopeptide cotransporter family SLC15 in physiology and
pharmacology. Pflugers Arch. 447, 610—618
Bhardwaj, R.K., Herrera-Ruiz, D., Eltoukhy, N., Saad,
M., and Knipp, G.T. (2006) The functional evaluation of
human peptide/histidine transporter 1 (hPHT1) in transiently transfected COS-7 cells. Eur. J. Pharm. Sci. 27,
533—542
Takeuchi, F., Ochiai, Y., Serizawa, M., Yanai, K.,
Kuzuya, N., Kajio, H., Honjo, S., Takeda, N.,
Kaburagi, Y., Yasuda, K., Shirasawa, S., Sasazuki, T.,
and Kato, N. (2008) Search for type 2 diabetes susceptibility genes on chromosomes 1q, 3q and 12q. J. Hum.
Genet. 53, 314—324
Lee, J., Tattoli, I., Wojtal, K.A., Vavricka, S.R.,
Philpott, D.J., and Girardin, S.E. (2009) pH-dependent
internalization of muramyl peptides from early endosomes enables Nod1 and Nod2 signaling. J. Biol.
Chem. 284, 23818—23829
Han, J.W., Zheng, H.F., Cui, Y., Sun, L.D., Ye, D.Q.,
Hu, Z., Xu, J.H., Cai, Z.M., Huang, W., Zhao, G.P.,
Xie, H.F., Fang, H., Lu, Q.J., Li, X.P., Pan, Y.F.,
Deng, D.Q., Zeng, F.Q., Ye, Z.Z., Zhang, X.Y., Wang,
Q.W., Hao, F., Ma, L., Zuo, X.B., Zhou, F.S., Du,
W.H., Cheng, Y.L., Yang, J.Q., Shen, S.K., Li, J.,
Sheng, Y.J., Zuo, X.X., Zhu, W.F., Gao, F., Zhang,
P.L., Guo, Q., Li, B., Gao, M., Xiao, F.L., Quan, C.,
Zhang, C., Zhang, Z., Zhu, K.J., Li, Y., Hu, D.Y., Lu,
W.S., Huang, J.L., Liu, S.X., Li, H., Ren, Y.Q., Wang,
Z.X., Yang, C.J., Wang, P.G., Zhou, W.M., Lv, Y.M.,
Zhang, A.P., Zhang, S.Q., Lin, D., Low, H.Q., Shen, M.,
Zhai, Z.F., Wang, Y., Zhang, F.Y., Yang, S., Liu, J.J.,
Endosomal/lysosomal signalling and inflammation
and Zhang, X.J. (2009) Genome-wide association study
in a Chinese Han population identifies nine new susceptibility loci for systemic lupus erythematosus. Nat. Genet.
41, 1234—1237
57. Blasius, A.L., Krebs, P., Sullivan, B.M., Oldstone, M.B.,
and Popkin, D.L. (2012) Slc15a4, a gene required for
pDC sensing of TLR ligands, is required to control persistent viral infection. PLoS Pathog. 8, e1002915
58. Franchi, L., Warner, N., Viani, K., and Nunez, G. (2009)
Function of Nod-like receptors in microbial recognition
and host defense. Immunol. Rev. 227, 106—128
59. Hampe, J., Cuthbert, A., Croucher, P.J., Mirza, M.M.,
Mascheretti, S., Fisher, S., Frenzel, H., King, K.,
Hasselmeyer, A., MacPherson, A.J., Bridger, S., van
Deventer, S., Forbes, A., Nikolaus, S., Lennard-Jones,
J.E., Foelsch, U.R., Krawczak, M., Lewis, C., Schreiber,
S., and Mathew, C.G. (2001) Association between insertion mutation in NOD2 gene and Crohn’s disease in
German and British populations. Lancet 357, 1925—1928
499