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Emerging inflammasome effector
mechanisms
Mohamed Lamkanfi
Abstract | Caspase 1 activation by inflammasome complexes in response to pathogenassociated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs)
induces the maturation and secretion of the pro-inflammatory cytokines interleukin-1β
(IL-1β) and IL-18. Recent reports have begun to identify additional inflammasome effector
mechanisms that proceed independently of IL-1β and IL-18. These include the induction of
pyroptotic cell death, the restriction of bacterial replication, the activation of lipid metabolic
pathways for cell repair and the secretion of DAMPs and leaderless cytokines. These
non-canonical functions of caspase 1 illustrate the diverse mechanisms by which
inflammasomes might contribute to innate immunity, repair responses and host defence.
Cryopyrinopathies
A spectrum of hereditary
autoinflammatory diseases
that are caused by mutations
in the gene encoding NLR
family, pyrin domaincontaining 3 (NLRP3) that
trigger continuous activation
of the NLRP3 inflammasome.
Based on the severity and
spectrum of the symptoms —
which can include urticarial
skin rashes, prolonged
episodes of fever, sensorineural
hearing loss, headaches,
cognitive deficits and renal
amyloidosis — these diseases
are classified as familial cold
autoinflammatory syndrome,
Muckle–Wells syndrome or
chronic infantile neurological
cutaneous articular syndrome.
Department of Biochemistry,
Ghent University, and VIB
Department of Medical
Protein Research, Albert
Baertsoenkaai 3, B‑9000
Ghent, Belgium.
e‑mail: Mohamed.Lamkanfi@
VIB‑UGent.be
doi:10.1038/nri2936
Inflammasomes (BOX 1) are emerging as key regulators
of the innate immune response, and the activity of these
multiprotein complexes has been linked to inflamma‑
tory bowel diseases1–5, vitiligo6, gouty arthritis7, type 1
and type 2 diabetes8,9, and less common autoinflamma‑
tory disorders that are collectively referred to as cryopyrinopathies10,11. Inflammasome complexes are thought to
be assembled around members of the NOD-like receptor
(NLR) or HIN‑200 protein families 12 (FIG. 1). These
receptors are thought to detect microbial pathogenassociated molecular patterns (PAMPs) and endogenous
damage-associated molecular patterns (DAMPs) in intra‑
cellular compartments, similar to the role of mammalian
Toll‑like receptors (TLRs) at the cell surface and within
endosomes13. Although it is incompletely understood
how NLRs detect microbial ligands and DAMPs14,15, it
is evident that inflammasome assembly results in the
activation of caspase 1 (BOX 2). This evolutionarily con‑
served cysteine protease is mainly known for its role
in the maturation of the pro‑inflammatory cytokines
interleukin‑1β (IL‑1β) and IL‑18 (ReFs 16–19).
IL‑1β and IL‑18 are related cytokines that are pro‑
duced as cytosolic precursors and usually require
caspase 1‑mediated cleavage for full activation and
secretion16–19. However, additional proteases, including
caspase 8, myeloblastin (also known as proteinase 3)
and granzyme A, have been shown to convert pro‑IL‑1β
into a biologically active cytokine in several established
mouse models of human disease20–23. This indicates that
caspase 1 is not always required for the maturation of
IL‑1β, and such redundancy in controlling IL‑1β matura‑
tion might safeguard the host immune response against
viral and bacterial pathogens that target caspase 1 activa‑
tion in inflammasomes24. Indeed, IL‑1β and IL‑18 were
recognized early on for their ability to cause a wide variety
of biological effects associated with infection, inflamma‑
tion and autoimmunity 25. IL‑1β regulates systemic and
local responses to infection, injury and immunological
challenge by generating fever, activating lymphocytes and
promoting leukocyte transmigration into sites of injury
or infection25. Although IL‑18 lacks the pyrogenic activity
of IL‑1β, it induces interferon‑γ (IFNγ) production by
activated T cells and natural killer cells in the presence
of IL‑12, thereby contributing to T helper 1 (TH1) cell
polarization25,26. In the absence of IL‑12, IL‑18 can pro‑
mote TH2 cell responses through the production of TH2
cell cytokines such as IL‑4, IL‑5 and IL‑10 (ReFs 26–28).
More recently, IL‑18 has also been implicated in driving
TH17 cell responses because it synergizes with IL‑23
to induce IL‑17 production by already committed
TH17 cells29,30. Thus, IL‑1β and IL‑18 are important
inflammasome effector molecules, as illustrated by the
marked response to therapy with IL‑1 inhibitors found
in patients with cryopyrinopathies, who have increased
inflammasome activation31,32.
However, not all inflammasome functions can be
abrogated by neutralization of IL‑1β and IL‑18. For exam‑
ple, caspase 1‑deficient mice are resistant to lipopoly‑
saccharide (LPS)‑induced shock, whereas mice lacking
both IL‑1β and IL‑18 are susceptible33. Moreover, a recent
study showed that IL‑1β and IL‑18 are not required for
caspase 1‑mediated clearance of several bacterial patho‑
gens (namely, modified Salmonella enterica subsp. enterica
serovar Typhimurium strains that constitutively express
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Box 1 | Inflammasomes
Inflammasomes are intracellular multiprotein complexes that mediate the
proximity-induced autoactivation of caspase 1. Inflammasome-mediated caspase 1
activation has been shown to occur in macrophages, dendritic cells, epithelial cells and
possibly other cell types during bacterial, viral, fungal and parasitic infections.
Inflammasomes are activated in response to stimulation with damage-associated
molecular patterns (DAMPs), such as uric acid and ATP, and upon exposure to crystalline
substances, such as monosodium urate, silica and asbestos particles12,24. The molecular
composition of inflammasome complexes is stimulus dependent, with certain members
of the NOD-like receptor (NLR) and HIN-200 receptor families functioning as the
activating platform in these complexes. Genetic studies in mice indicate the existence of
at least four types of inflammasome (FIG. 1). Three of these contain NLR proteins, namely
NLR family, pyrin domain-containing 1B (NLRP1B), NLR family, CARD-containing 4
(NLRC4) and NLRP3. The fourth type of inflammasome contains the HIN-200 protein
absent in melanoma 2 (AIM2)12. The bipartite adaptor protein ASC (apoptosis-associated
speck-like protein containing a CARD; also known as PYCARD) probably has a key role in
inflammasome assembly and caspase 1 activation by bridging the interaction between
NLRs or HIN-200 proteins and caspase 1, although the precise role of ASC in the activation
of the NLRP1B and NLRC4 inflammasomes is debated. NLRP1B and NLRC4 contain a
caspase recruitment domain (CARD) at their carboxyl and amino termini, respectively
(unlike AIM2 and NLRP3, which have a pyrin domain) and can therefore interact directly
with caspase 1 when overexpressed, without requiring ASC. However, evidence of a role
for ASC in the activation of the endogenous NLRC4 inflammasome is provided by the
observation that robust caspase 1 activation and the production of interleukin-1β (IL-1β)
and IL-18 are markedly decreased in ASC-deficient macrophages infected with viral or
bacterial pathogens, or exposed to a variety of DAMPs and crystalline substances50,63,88.
flagellin, Legionella pneumophila and Burkholderia thailandensis)34. Similarly, caspase 1‑deficient mice are more
susceptible to infection with Francisella tularensis than
mice lacking both IL‑1β and IL‑18 (ReF. 35), and this
indicates that additional caspase 1‑dependent mecha‑
nisms might contribute to the control of infection. In
this regard, several recent publications have begun to
characterize a range of new inflammasome functions and
effector molecules that seem to operate independently
of IL‑1β and IL‑18. In this article, I review these emerg‑
ing non‑canonical inflammasome effector mechanisms
and attempt to illustrate how they might contribute to
immune responses.
Proximity-induced
autoactivation
A process in which two or
more initiator caspases are
brought sufficiently close to
induce their autocatalytic
activation. This process is
thought to occur in large
cytosolic protein complexes
to which caspase zymogens
are recruited by means of
homotypic interactions
between the caspase
recruitment domain (CARD) or
death effector domain (DeD)
motifs in their pro-domains
and several bipartite adaptor
molecules.
Unconventional protein secretion
Secretory proteins usually contain amino‑terminal or
internal signal peptides that target them to the translo‑
cation apparatus of the endoplasmic reticulum (eR)36,37.
From the eR lumen, such proteins are transported to
the Golgi apparatus and then to the extracellular space
in Golgi‑derived secretory vesicles that fuse with the
plasma membrane37. This pathway of protein export is
known as the ‘eR–Golgi’ or ‘classical’ secretory pathway.
However, cytoplasmic and nuclear proteins that lack an
eR‑targeting signal peptide can exit cells through eR‑
and Golgi‑independent pathways38. For example, mature
IL‑1β was shown to be secreted independently of the eR
and the Golgi apparatus more than 20 years ago39. The
number of proteins that have been shown to be released
by unconventional protein secretion has grown to more
than 20, including fibroblast growth factor 2 (FGF2), the
lectins galectin 1 and galectin 3, and possibly the DAMP
high‑mobility group box 1 (HMGB1)38. After their release
into the extracellular space, these effectors can enhance
inflammatory, cell survival and repair responses through
activation of cell surface receptors, such as FGF recep‑
tor 1, the IL‑1 and IL‑18 receptors and the receptor for
advanced glycation end‑products (RAGe).
Although the molecular mechanism by which IL‑1β,
IL‑18 and other proteins that lack signal peptides are
secreted remains obscure, several models have been
proposed to explain the release of such ‘leaderless pro‑
teins’ in microvesicles that are shed from the plasma
membrane, or in secretory lysosomes or exosomes38.
Interestingly, adherent monocytes from caspase 1‑
deficient mice and peritoneal macrophages from mice
lacking two inflammasome components — namely,
NLR family, pyrin domain‑containing 3 (NLRP3) and
apoptosis‑associated speck‑like protein containing a
cARD (ASc; also known as PYcARD) — not only failed
to secrete IL‑1β and IL‑18 after LPS stimulation16,18,19,40,
but were also partially defective in the secretion of the
leaderless cytokine IL‑1α17,40. unlike IL‑1β and IL‑18,
IL‑1α does not undergo caspase 1‑mediated cleavage26.
This might indicate that caspase 1 modulates IL‑1α
secretion indirectly by regulating the secretory path‑
way of this cytokine, and may point to a broader role for
caspase 1 in regulating unconventional protein secretion.
Indeed, pharmacological inhibition, RNA interference
(RNAi)‑mediated downregulation and targeted deletion
of caspase 1 were all recently shown to block the secre‑
tion of IL‑1β, IL‑1α and FGF2 by macrophages, uvA‑
irradiated fibroblasts and uvB‑irradiated keratinocytes,
respectively 41. In addition, caspase 1 activation by either
the NLRP3 inflammasome or the NLR family, cARD‑
containing 4 (NLRc4) inflammasome was required for
secretion of the nuclear DAMP HMGB1 from activated
and infected macrophages33. Because the enzymatic
activity of caspase 1 is required for the secretion of each
of these leaderless proteins33,41, caspase 1 might medi‑
ate the proteolytic activation of a secretion apparatus
of unknown identity. In this context, the small GTPase
RAB39A was recently characterized as a caspase 1 sub‑
strate that is involved in the secretion of IL‑1β from
LPS‑activated human THP‑1 monocytes42. However, it
remains to be determined how caspase 1‑mediated cleav‑
age affects RAB39A function and whether RAB39A has a
role in the secretion of additional leaderless proteins. An
alternative mechanism by which caspase 1 might promote
the release of leaderless proteins such as HMGB1 involves
the induction of a specialized caspase 1‑mediated cell
death programme in activated immune cells, which is
often referred to as pyroptosis (see below). elucidation
of the mechanism awaits the characterization of the
molecular components mediating caspase 1‑dependent
unconventional protein secretion and pyroptosis.
Pyroptosis
Most caspases (BOX 2) — caspases 2, 3 and 6–10 — are
implicated in the induction and execution of apopto‑
sis43,44. This form of programmed cell death is respon‑
sible for organ shaping during embryonic development
and for preserving homeostasis in adult organisms.
Typical morphological features of apoptosis include
plasma membrane blebbing, condensation of the nucleus,
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Figure 1 | Inflammasomes: composition and stimuli. The NOD-like receptor (NLR)
proteins NLR family, pyrin domain-containing 1B (NLRP1B), NLR family, CARD-containing 4
(NLRC4) and NLRP3, and the HIN-200 protein absent in melanoma 2 (AIM2) assemble
inflammasomes in a stimulus-specific manner12. NLRP1B recognizes the cytosolic presence
of the Bacillus anthracis lethal toxin64. The NLRC4 inflammasome is assembled after
detection of bacterial flagellin or the basal body rod component of the bacterial type III
and type IV secretion systems of Salmonella enterica subsp. enterica serovar Typhimurium,
Pseudomonas aeruginosa, Legionella pneumophila and Shigella flexneri12,58,62. NLRP3 is
activated when macrophages are exposed to UV irradiation, microbial pathogenassociated molecular patterns (PAMPs), endogenous damage-associated molecular
patterns (DAMPs) such as ATP, or crystals such as monosodium urate, silica and asbestos.
Recognition of these PAMPs, DAMPs and crystals is thought to involve the detection of a
common secondary messenger, such as K+ fluxes, reactive oxygen species or lysosomal
proteases14,15. By contrast, AIM2 directly binds double-stranded DNA (dsDNA) in the
cytosol to induce caspase 1 activation in cells infected with Francisella tularensis, Listeria
monocytogenes or DNA viruses such as cytomegalovirus and vaccinia virus68–70,72–74. The
adaptor protein ASC (apoptosis-associated speck-like protein containing a CARD; also
known as PYCARD) is probably required for the full activation of all inflammasome
complexes, although its role in the NLRP1B and NLRC4 inflammasomes is still debated45.
CARD, caspase recruitment domain; LRR, leucine-rich repeat. PYD, pyrin domain.
DNA fragmentation and general shrinkage of the cell
volume45. Apoptotic cells usually fail to elicit inflam‑
matory responses because the cytoplasmic content is
shielded from the extracellular environment by packag‑
ing in ‘apoptotic bodies’ (FIG. 2). These apoptotic bodies
are membrane‑bound cell fragments that are rapidly
phagocytosed in vivo by neighbouring cells and resident
phagocytes46,47. The integrity of the envelope surround‑
ing apoptotic bodies is usually preserved until after they
have been engulfed by professional antigen‑presenting
cells or neighbouring cells to prevent accidental spilling
of the intracellular content into the extracellular milieu47.
Apoptotic cells can also prevent the accidental induction
of inflammation by inactivating the immunostimulatory
activity of the DAMP HMGB1 through the oxidation of
residue cys106 (ReF. 48). Finally, the uptake of apoptotic
bodies by macrophages and dendritic cells was recently
proposed to actively suppress antigen presentation and
the secretion of inflammatory cytokines by these cells49.
unlike most caspases, caspase 1 is not involved in the
induction of apoptosis. Instead, caspase 1 activation in
neurons, macrophages and dendritic cells drives a spe‑
cialized form of cell death known as pyroptosis34,50–53.
caspase 1‑dependent cell death was first reported to
occur in macrophages infected with Shigella flexneri, the
causative agent of bacillary dysentery 54,55. Pyroptosis was
subsequently observed in macrophages and dendritic
cells infected with the facultative intracellular pathogens
S. Typhimurium, Pseudomonas aeruginosa and L. pneumophila45,56,57. each of these pathogens induces caspase 1
activation through the NLRc4 inflammasome (FIG. 1).
These pathogens are recognized when bacterial flagel‑
lin is transferred through specialized bacterial type III
and type IV secretion systems into the host cell cytosol
or upon detection of the basal body rod component of
the S. flexneri or P. aeruginosa type III secretion appara‑
tus58–63. The induction of pyroptosis is not restricted to
the NLRc4 inflammasome, as Bacillus anthracis infec‑
tion induces the pyroptotic cell death of mouse macro‑
phages through the NLRP1B inflammasome64,65. This
occurs when the anthrax metalloprotease lethal factor
gains access to the cytosol of susceptible macrophages64.
Notably, lethal factor‑induced pyroptosis was shown to
confer resistance to infection with B. anthracis spores
in vivo65, highlighting the importance of pyroptosis for
host defence against pathogens. The pyroptotic cell death
of macrophages infected with Staphylococcus aureus
requires activation of the NLRP3 inflammasome66,67.
Although the precise mechanism is still debated, acti‑
vation of this inflammasome could proceed through
several mutually non‑exclusive mechanisms, including
K+ efflux, the generation of reactive oxygen species, lyso‑
somal destabilization and the translocation of microbial
ligands into the host cytosol14,15. Finally, infections with
the bacterial pathogens Listeria monocytogenes and
F. tularensis induce pyroptosis upon their recognition
in the host cell cytosol by the absent in melanoma 2
(AIM2) inflammasome68–74. The role of caspase 1 in the
immune response to F. tularensis, the causative agent
of tularaemia, is illustrated by the observation that
caspase 1‑deficient mice have increased susceptibility
to infection with this pathogen75. Notably, mice lacking
the canonical inflammasome substrates IL‑1β and IL‑18
are less susceptible to infection with F. tularensis than
caspase 1‑deficient mice 35, and this indicates that
additional caspase 1‑dependent mechanisms, such as
pyroptosis, might make an important contribution to
the control of F. tularensis infection. Indeed, although
caspase 1 activity is required for pyroptosis51,76, this
form of cell death proceeds independently of IL‑1β
and IL‑18 (ReF. 56). A recent study established the cru‑
cial in vivo role of pyroptosis in clearing a modified
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Box 2 | Caspases
Caspases are an evolutionarily conserved family of metazoan cysteine proteases with
11 representatives in humans: caspases 1–10 and caspase 14. Caspases cleave various
cellular substrates after aspartic acid residues and have essential roles in apoptosis,
inflammation, cell proliferation and cell differentiation89. For example, caspase 3mediated cleavage of mammalian STE20-like kinase 1 (MST1; also known as STK4) was
shown to be crucial for skeletal muscle differentiation, and caspase 8-mediated
cleavage of the long splice variant of cellular FLICE-like inhibitory protein (cFLIPL)
regulates the balance between apoptosis induction and nuclear factor-κB
(NF-κB)-driven T cell proliferation90. All caspases are synthesized as zymogens consisting
of an amino-terminal pro-domain of variable length and a carboxy-terminal protease
domain. Caspases can be subdivided according to the length of their pro-domain
(see the figure). Initiator caspases (such as caspases 1 and 8) have large pro-domains
containing homotypic protein–protein interaction motifs of the death domain
superfamily, specifically either a caspase recruitment domain (CARD) or a death effector
domain (DED). These interaction motifs allow the recruitment of pro-caspases into
multiprotein complexes (such as the inflammasomes) by homotypic interactions with
adaptor molecules such as ASC (apoptosis-associated speck-like protein containing a
CARD; also known as PYCARD). Within these complexes, pro-caspases undergo the
conformational changes and/or autoprocessing required for their activation91. By
contrast, the effector caspases (caspases 3, 6, 7 and 14) have short pro-domains of only a
few amino acids and they lack any homotypic interaction motifs. These caspases require
proteolytic maturation by the initiator caspases or other proteases to achieve maximum
enzymatic activity. Unlike the ‘true’ caspases listed above, human caspase 12 is devoid
of enzymatic activity because crucial catalytic residues have been mutated. In addition,
most people express a truncated form of caspase 12 that resembles the human
CARD-only proteins CARD17 (also known as INCA), CARD18 (also known as ICEBERG)
and CARD16 (also known as COP or pseudo-ICE)92.
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NOD-like receptor
(NLR). The human NLR family
comprises 22 members. They
share a domain organization
that usually includes an
amino-terminal caspase
recruitment domain (CARD)
or pyrin domain (PYD),
followed by an intermediary
nucleotide-binding
oligomerization domain (NOD)
and carboxy-terminal
leucine-rich repeat motifs.
NLRs are thought to survey the
host cytosol and intracellular
compartments for pathogenand damage-associated
molecular patterns to activate
signalling pathways that
contribute to the host innate
immune response.
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S. Typhimurium
strain that constitutively expressed
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bacterial flagellin34. The pyroptotic cell death of infected
macrophages exposed intracellular bacteria to extracel‑
lular immune surveillance and allowed their destruction
by neutrophils. Pyroptosis also conferred protection
against the bacterial pathogens L. pneumophila and
B. thailandensis, which establishes this inflammasome
effector mechanism as a crucial component of host
defence against intracellular bacterial pathogens34.
Although the molecular mechanisms controlling
pyroptosis are still poorly defined, this cell death mode
differs morphologically from apoptosis in that pores with
a diameter of 1–2 nm appear in the plasma membrane of
pyroptotic cells at early time points52. The resulting ion
fluxes could explain some of the hallmarks of pyroptotic
cells, including cytoplasmic swelling, osmotic lysis and
the release of the intracellular content into the extracel‑
lular milieu77 (FIG. 2). In addition to eliminating infected
immune cells, this process can enhance innate and adap‑
tive immune responses by exposing microbial antigens
to surveillance by the immune system. Together with the
fact that caspase 1 activation is linked with the produc‑
tion of mature IL‑1β and IL‑18 and the release of leader‑
less cytokines and DAMPs (such as HMGB1 and S100
proteins), these characteristics are thought to render
pyroptosis (and associated inflammasome functions)
an inherently pro‑inflammatory cell death mode.
Despite the different immunological outcomes of
apoptosis and pyroptosis (FIG. 2), apoptotic and pyrop‑
totic cells share several prominent morphological and
biochemical features. Nuclear condensation and oligo‑
nucleosomal DNA fragmentation are observed in both
cell death modes19,46,51,52,76. Moreover, the DNA damage
sensor poly(ADP‑ribose) polymerase 1 (PARP1) is proc‑
essed into an 89 kDa fragment during both apoptosis and
pyroptosis46,78. under homeostatic conditions, PARP1
participates in genomic DNA repair and DNA replica‑
tion by catalysing the synthesis of poly(ADP‑ribose) in a
process that consumes NAD+ and the ATP energy stores
of the cell. Thus, cleavage of PARP1 during both apop‑
tosis and pyroptosis indicates that PARP1 inactivation
might be a general strategy used by cells undergoing pro‑
grammed cell death, possibly to preserve energy stores in
order to allow for proper dismantling of the cell. Finally,
maturation of caspase 3 and caspase 7 is observed during
both apoptosis and pyroptosis51,78,79, although pyroptosis‑
associated DNA fragmentation, PARP1 processing and
plasma membrane permeabilization are not affected
in macrophages lacking either of these executioner
caspases51,55,78. This might not come as a complete surprise
given that caspase 3 and caspase 7 are partially redundant
and that deletion of both caspases was necessary for pro‑
tection against apoptosis80. Nevertheless, S. Typhimurium
has been shown to induce pyroptosis in macrophages
lacking both caspase 3 and caspase 7 (ReF. 45), and this
confirms that these executioner caspases are not neces‑
sary for pyroptosis. Thus, although apoptosis and pyrop‑
tosis share the morphological and biochemical features
described above (FIG. 2), the signalling pathways involved
are distinct. Despite recent progress in characterizing the
molecular features of pyroptotic cell death and its role in
host defence against bacterial pathogens, much remains
to be learned about the signalling mechanism by which
caspase 1 induces pyroptosis.
Inhibition of glycolysis
In an attempt to identify new caspase 1 substrates
that could explain the phenotype of pyroptotic cells, a
caspase 1 digestome analysis was carried out, and this
identified several crucial enzymes of bioenergetic
pathways as potential caspase 1 targets81. Biochemical
studies confirmed that the glycolysis enzymes fructose‑
bisphosphate aldolase, glyceraldehyde‑3‑phosphate dehy‑
drogenase, α‑enolase and pyruvate kinase can be cleaved
by recombinant caspase 1. These enzymes operate in a
metabolic pathway that replenishes cellular ATP energy
stores through the conversion of glucose to pyruvate.
caspase 1‑mediated processing of glyceraldehyde‑3‑
phosphate dehydrogenase inhibited its enzymatic activ‑
ity 81, and this indicated that caspase 1 might decrease
the metabolic rate of infected cells. To address whether
caspase 1‑mediated processing of these metabolic
enzymes occurred during infection, peritoneal macro‑
phages of wild‑type and caspase 1‑deficient mice were
infected with S. Typhimurium. As expected, aldolase was
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2[TQRVQUKU
Pathogen-associated
molecular pattern
(PAMP). A conserved pathogen
molecule that is usually
essential for microbial survival,
and that contains either nucleic
acid structures that are unique
to microorganisms or cell wall
components (such as
lipopolysaccharide and
flagellin) that are not found in
mammalian cells. PAMPs are
ligands for receptors of the
host innate immune system.
Damage-associated
molecular pattern
(DAMP). A molecule that is
produced or released from host
cells upon cellular stress,
damage or non-physiological
cell death. DAMPs are also
referred to as ‘alarmins’ and are
thought to be responsible for
the initiation and perpetuation
of inflammatory responses and
tissue repair under
non-infectious (sterile)
conditions. examples include
high-mobility group box 1
(HMGB1), ATP, uric acid and
heat-shock proteins.
Unconventional protein
secretion
The secretion of cytoplasmic
and nuclear proteins into the
extracellular space through
an incompletely understood
mechanism that does not
require the translocation
apparatus of the classical
endoplasmic reticulum
(eR)–Golgi secretion pathway.
Proteins that are secreted
through this route include
interleukin-1α (IL-1α), IL-1β,
IL-18, fibroblast growth
factor 2, galectin 1, galectin 3
and possibly high-mobility
group box 1.
Pyroptosis
A specialized form of
programmed cell death that
requires caspase 1 activity.
It is characterized by
cytoplasmic swelling, early
plasma membrane rupture,
nuclear condensation and
internucleosomal DNA
fragmentation. The
cytoplasmic content is released
into the extracellular space,
and this is thought to augment
inflammatory and repair
responses. Pyroptosis occurs
in myeloid cells infected with
pathogenic bacteria, and it
might affect cells of the central
nervous system and the
cardiovascular system under
ischaemic conditions.
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+OOWPQNQIKECNN[
UKNGPVEGNNFGCVJ
Figure 2 | main features of pyroptosis. The molecular mechanisms underlying pyroptosis are still poorly defined.
Morphologically, pyroptotic cells are characterized by the early loss of plasma membrane integrity77, and this is accompanied
0CVWTG4GXKGYU^+OOWPQNQI[
by the shedding of membrane vesicles93. Pyroptotic and apoptotic cells share several prominent
features (shown in blue
boxes), including nuclear condensation and internucleosomal DNA fragmentation, cleavage of the DNA damage repair
enzyme poly(ADP-ribose) polymerase 1 (PARP1) and activation of the executioner caspases caspase 3 and caspase 7 (ReF. 45).
However, the volume of the cytoplasmic compartment of pyroptotic cells increases, whereas apoptosis is characterized by
general shrinkage of the cell volume. Together with the role of caspase 1 in cytokine maturation and unconventional protein
secretion, the release of the cytoplasmic content into the extracellular milieu during pyroptosis is thought to render this
form of cell death inherently pro-inflammatory45. By contrast, apoptosis is usually considered to be immunologically silent
because the cytoplasmic content is packaged in apoptotic bodies and these membrane-bound cell fragments are rapidly
phagocytosed in vivo by neighbouring cells and resident phagocytes47. HMGB1, high mobility group box 1; IL, interleukin.
processed in infected wild‑type macrophages, but not in
those lacking caspase 1 (ReF. 81). concurrently, the rate of
glycolysis in the caspase 1‑deficient cells was markedly
higher, further supporting an inhibitory role for caspase 1
in the regulation of glycolysis. Because myeloid cells are
highly dependent on glycolysis for ATP production82,
caspase 1‑mediated inactivation of glycolysis enzymes
might restrict intracellular pathogen replication by quickly
depleting energy sources and by preparing infected host
cells to undergo pyroptosis. However, it remains to
be determined whether and to what extent caspase 1‑
mediated inactivation of glycolysis enzymes contributes to
protection against bacterial pathogenicity in vivo.
Cell survival
Another mechanism by which caspase 1 might protect
host cells is by repairing the damage caused by bacte‑
rial pore‑forming toxins that are released by pathogenic
bacteria83. The pores formed by these toxins can range
significantly in diameter, depending on the nature of the
toxin. For example, S. aureus α‑toxin and Aeromonas
hydrophila aerolysin typically produce holes in the host
cell membrane with a diameter of only 2 nm, whereas
Streptococcus pneumoniae and L. monocytogenes pro‑
duce toxins that can create perforations of up to 50 nm
in diameter 84. consequently, the latter toxins allow trans‑
location of large proteins across the plasma membrane,
whereas S. aureus α‑toxin and A. hydrophila aerolysin
only render the plasma membrane permeable to small
inorganic ions84. Nevertheless, depending on the concen‑
tration of these toxins and the targeted cell type, the dam‑
age elicited by these toxins can range from irreparable cell
destruction to temporal perforations that can quickly be
resealed by the host cell’s dedicated repair machinery.
Repairing bacterial toxin‑induced damage to the
plasma membrane requires the activation of lipid bio‑
genesis pathways, the molecular machinery of which is
controlled by two transcription factors known as sterol
regulatory element‑binding protein 1 (SReBP1) and
SReBP2 (ReF. 84). Interestingly, activation of SReBP1
and SReBP2 is controlled by the NLRP3 and NLRc4
inflammasomes in fibroblasts that have been intoxicated
with S. aureus α‑toxin or A. hydrophila aerolysin, or
infected with live Aeromonas trota bacteria83. This path‑
way is thought to promote host cell survival because inhi‑
bition or transcriptional downregulation of SReBP1 and
SReBP2 enhanced cell death responses83. Inflammasome‑
induced activation of SReBPs is thought to be indirect,
but the caspase 1 substrates that drive this pathway
remain to be found. It would be interesting to determine
whether caspase 1 activates SReBPs in macrophages
infected with live A. hydrophila, because these bacteria
were recently shown to activate caspase 1 through the
NLRP3, but not the NLRc4, inflammasome85. This might
reveal differential signalling mechanisms induced by the
three A. hydrophila cytotoxins85.
NATuRe RevIewS | Immunology
voLuMe 11 | MARcH 2011 | 217
© 2011 Macmillan Publishers Limited. All rights reserved
REVIEWS
Glycolysis
A metabolic pathway that
generates the cellular
high-energy store ATP by
oxidizing glucose to pyruvate.
In eukaryotic cells, pyruvate is
further oxidized into CO2 and
H2O in a process known as
‘aerobic respiration’. This
results in a net yield of
36–38 molecules of ATP per
metabolized molecule of
glucose.
Autophagosome
A double-membrane-bound
vesicle that is used by
eukaryotic cells to target
protein aggregates, damaged
organelles and invading
microorganisms for digestion
by lysosomal hydrolases. This
catabolic process allows
recycling of cellular
components and is thought to
contribute to cell death, cell
survival during starvation,
cellular differentiation and host
defence against infectious
agents.
#EVKXG
ECURCUG
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NGCFGTNGUUE[VQMKPGU
CPF&#/2U
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ITQYVJHCEVQTU
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+PȯCOOCVKQP
the NLRP3 and NLRc4 inflammasomes in inducing
proteolytic maturation of caspase 7 in activated immune
cells that were exposed to ATP and nigericin or infected
with live S. Typhimurium51. In contrast to caspase 7, the
activation of caspase 3 was not affected in caspase 1‑
deficient macrophages51, and this indicates that the
activation of caspase 3 and caspase 7 is differentially reg‑
ulated during inflammation. These observations identi‑
fied an alternative mechanism by which inflammasomes
might control bacterial infections. Indeed, caspase 7
activation downstream of the NLRc4 inflammasome
was subsequently reported in macrophages infected
with L. pneumophila 79. Moreover, caspase 7‑deficient
macrophages are less capable of restricting intracellular
L. pneumophila replication, possibly owing to defects
in the fusion of bacteria‑containing phagosomes with
lysosomes and the delayed induction of macrophage
cell death79. Importantly, caspase 1 and caspase 7 regu‑
late L. pneumophila growth in the lungs of orally infected
mice61,79, demonstrating the importance of this inflamma‑
some effector pathway in host defence against this bac‑
terial pathogen. However, it remains to be determined
%#4&
Two of at least six specialized
secretion systems by which
Gram-negative pathogens can
deliver virulence factors into
eukaryotic host cells.
Pathogenic bacteria such as
Shigella, Salmonella, Yersinia,
Chlamydia and Pseudomonas
spp. all make use of a type III
secretion system to infect host
cells and to modulate signalling
pathways. By contrast,
pathogens such as
Helicobacter pylori, Legionella
pneumophila and Bordetella
pertussis make use of a type IV
secretion system for the
horizontal transfer of plasmid
DNA containing antibiotic
resistance genes and to inject
effector proteins into
eukaryotic host cells.
Caspase 7 activation
Activation of the NLRc4 inflammasome was recently
shown to restrict the intracellular replication of L. pneumophila, the causative agent of a severe form of bacte‑
rial pneumonia known as Legionnaires’ disease61,86,87.
Inflammasome activation in resistant mouse strains
results in the rapid caspase 1‑dependent delivery of
L. pneumophila to lysosomes, where the bacteria are
degraded. By contrast, defective inflammasome activa‑
tion in Nlrc4–/– and Casp1–/– mice allows bacterial repli‑
cation in specialized intracellular vesicles that resemble
autophagosomes61,87. Notably, inflammasome‑mediated
restriction of L. pneumophila replication proceeds
independently of IL‑1β and IL‑18 (ReFs 61,87), but the
caspase 1 substrates that are responsible for this proc‑
ess remain unclear. A proteome‑wide screen for new
caspase 1 targets identified caspase 7, an effector caspase,
as a direct substrate of caspase 1, and biochemical studies
confirmed that caspase 7 is cleaved by caspase 1 after
the canonical activation sites Asp23 and Asp198 (ReF. 51).
Importantly, studies in macrophages from Nlrp3–/–,
Nlrc4–/–, Asc–/– or Casp1–/– mice confirmed the role of
%#4&
Type III and type IV
secretion
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GPGTI[UQWTEGU
/GODTCPG
RGTOGCDKNK\CVKQP
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HTCIOGPVCVKQP
%CURCUG
4GRCKTCPFJGCNKPI
2[TQRVQUKU
4GOQXCNQH
KPVTCEGNNWNCT
TGRNKECVKQPPKEJGU
+OOWPG
UWTXGKNNCPEG
QHOKETQDKCN
EQORQPGPVU
4GUVTKEVKQPQH
.GIKQPGNNC
TGRNKECVKQPD[
VCTIGVKPIVQ
N[UQUQOGU
Figure 3 | Caspase 1 effector mechanisms. Pathogen invasion of macrophages and dendritic cells triggers the assembly
of inflammasome complexes and caspase 1 activation. Active caspase 1 induces inflammation by mediating the
0CVWTG4GXKGYU^+OOWPQNQI[
extracellular secretion or release of leaderless cytokines such as interleukin-1β (IL-1β), IL-18 and IL-1α, and possibly
damage-associated molecular patterns (DAMPs) such as high mobility group box 1 (HMGB1), through an as yet unknown
mechanism. Caspase 1 also promotes repair and healing responses by inducing lipid membrane biogenesis through the
activation of sterol regulatory element binding proteins (SREBPs) and through the secretion or release of growth factors
such as fibroblast growth factor 2. The latter contributes to repair through ligation of cell surface receptors on target cells.
Caspase 1 cleaves poly(ADP-ribose) polymerase 1 (PARP1) and glycolysis enzymes, possibly to prepare host cells to
undergo pyroptosis. This specialized cell death programme removes intracellular niches for microbial replication and
eliminates infected immune cells. Moreover, it might help to tune immune responses by releasing microbial components
into the extracellular milieu, where they can be detected by the immune system. It is probable that caspase 1 cleaves
additional as yet unidentified substrates that are responsible for early membrane permeabilization and oligonucleosomal
DNA fragmentation during pyroptosis. Finally, inflammasome-mediated activation of caspase 7 (an effector caspase)
restricts bacterial replication in Legionella-infected macrophages by targeting the infectious agent to lysosomes. CARD,
caspase recruitment domain.
218 | MARcH 2011 | voLuMe 11
www.nature.com/reviews/immunol
© 2011 Macmillan Publishers Limited. All rights reserved
REVIEWS
whether inflammasome‑mediated activation of caspase 7
also restricts the replication of S. Typhimurium and other
bacterial pathogens, and whether this inflammasome
effector pathway is activated during viral infection.
Conclusions and perspectives
It has become evident in recent years that inflamma‑
somes have important roles in innate immune signalling
and host defence. In particular, our knowledge of how
inflammasome complexes of distinct composition are
assembled in a stimulus‑dependent manner has grown
significantly. Now, the different effector mechanisms (in
addition to IL‑1β and IL‑18 secretion) by which inflam‑
masomes might contribute to immunity and host defence
are also starting to emerge. As described above, recent
studies have highlighted a range of new inflammasome
functions and effector mechanisms (FIG. 3). caspase 1
has been shown to control the secretion of leaderless
cytokines and proteins such as IL‑1α and FGF2, as well
as the release of endogenous DAMPs such as HMGB1.
Moreover, excessive caspase 1 activation in damaged neu‑
rons and infected myeloid cells induces pyroptotic cell
death. Furthermore, caspase 1 dampens the metabolic
rate of infected cells by cleaving key enzymes of the glyco‑
lysis pathway and regulates lipid metabolic pathways for
cell repair. Finally, activation of the executioner caspase 7
downstream of inflammasomes contributes to restriction
of Legionella replication in infected macrophages.
Interestingly, most of these emerging functions of
caspase 1 seem to operate independently of the canoni‑
cal substrates IL‑1β and IL‑18, and this indicates that
inflammasomes contribute to innate immune responses
in a variety of ways. Indeed, these effector mechanisms
probably function together to mount a fast and effec‑
tive innate immune response against the pathogen and
Villani, A. C. et al. Common variants in the NLRP3
region contribute to Crohn’s disease susceptibility.
Nature Genet. 41, 71–76 (2009).
2.
Zaki, M. H. et al. The NLRP3 inflammasome protects
against loss of epithelial integrity and mortality during
experimental colitis. Immunity 32, 379–391 (2010).
3.
Allen, I. C. et al. The NLRP3 inflammasome functions
as a negative regulator of tumorigenesis during colitisassociated cancer. J. Exp. Med. 207, 1045–1056
(2010).
4.
Bauer, C. et al. Colitis induced in mice with dextran
sulfate sodium (DSS) is mediated by the NLRP3
inflammasome. Gut 59, 1192–1199 (2010).
5.
Dupaul-Chicoine, J. et al. Control of intestinal
homeostasis, colitis, and colitis-associated colorectal
cancer by the inflammatory caspases. Immunity
32, 367–378 (2010).
6.
Jin, Y. et al. NALP1 in vitiligo-associated multiple
autoimmune disease. N. Engl. J. Med. 356,
1216–1225 (2007).
7.
Martinon, F., Petrilli, V., Mayor, A., Tardivel, A. &
Tschopp, J. Gout-associated uric acid crystals activate
the NALP3 inflammasome. Nature 440, 237–241
(2006).
8.
Magitta, N. F. et al. A coding polymorphism in NALP1
confers risk for autoimmune Addison’s disease and
type 1 diabetes. Genes Immun. 10, 120–124 (2009).
9.
Larsen, C. M. et al. Interleukin-1-receptor antagonist
in type 2 diabetes mellitus. N. Engl. J. Med. 356,
1517–1526 (2007).
10. Agostini, L. et al. NALP3 forms an IL-1β-processing
inflammasome with increased activity in Muckle–Wells
autoinflammatory disorder. Immunity 20, 319–325
(2004).
1.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
might instruct the adaptive immune system regarding
the systemic risk posed by the invading microbial agent.
At an early phase of infection, activation of caspase 7
downstream of the inflammasome might halt pathogen
replication in intracellular replication niches, while at
the same time inflammasome‑mediated activation of
SReBPs could repair pathogen‑induced damage to the
plasma membrane. when bacterial (or viral) loads fur‑
ther increase, inflammasomes can induce pyroptosis
and instruct macrophages and dendritic cells to initi‑
ate the unconventional secretion or passive release of
pro‑inflammatory cytokines, growth factors, DAMPs
and microbial antigens to alert the immune system of
an imminent threat and to initiate repair responses.
In this context, the crucial role of pyroptosis in host
defence was recently demonstrated in mice infected
with the Gram‑negative bacterial pathogens L. pneumophila and B. thailandensis 34. Pyroptosis also confers
resistance against Gram‑positive pathogens such as
B. anthracis in vivo65, but its role in clearing viral and
fungal infections remains to be determined. In addi‑
tion, much remains to be learned regarding exactly how
caspase 1 induces pyroptosis and how inflammasomes
regulate the unconventional secretion of leaderless pro‑
teins. what are the crucial components of the molecular
machinery driving the secretion of leaderless proteins
and how are they regulated by inflammasomes? Are
unconventional protein secretion and pyroptosis intrin‑
sically linked or can they be uncoupled? what is the
relative contribution of each of these inflammasome
effector mechanisms to innate and adaptive immune
responses during microbial infections? Answering
these and other questions will undoubtedly illuminate
intriguing new mechanisms by which inflammasomes
contribute to host defence and immunity.
Hoffman, H. M., Mueller, J. L., Broide, D. H.,
Wanderer, A. A. & Kolodner, R. D. Mutation of a
new gene encoding a putative pyrin-like protein
causes familial cold autoinflammatory syndrome
and Muckle–Wells syndrome. Nature Genet. 29,
301–305 (2001).
Lamkanfi, M. & Dixit, V. M. The inflammasomes.
PLoS Pathog. 5, e1000510 (2009).
Kawai, T. & Akira, S. TLR signaling. Cell Death Differ.
13, 816–825 (2006).
Tschopp, J. & Schroder, K. NLRP3 inflammasome
activation: the convergence of multiple signalling
pathways on ROS production? Nature Rev. Immunol.
10, 210–215 (2010).
Lamkanfi, M. & Dixit, V. M. Inflammasomes:
guardians of cytosolic sanctity. Immunol. Rev. 227,
95–105 (2009).
Gu, Y. et al. Activation of interferon-γ inducing factor
mediated by interleukin-1β converting enzyme.
Science 275, 206–209 (1997).
Kuida, K. et al. Altered cytokine export and apoptosis
in mice deficient in interleukin-1β converting enzyme.
Science 267, 2000–2003 (1995).
Ghayur, T. et al. Caspase-1 processes IFN-γ-inducing
factor and regulates LPS-induced IFN-γ production.
Nature 386, 619–623 (1997).
Li, P. et al. Mice deficient in IL-1β-converting enzyme
are defective in production of mature IL-1β and
resistant to endotoxic shock. Cell 80, 401–411 (1995).
Joosten, L. A. et al. Inflammatory arthritis in caspase 1
gene-deficient mice: contribution of proteinase 3 to
caspase 1-independent production of bioactive
interleukin-1β. Arthritis Rheum. 60, 3651–3662
(2009).
NATuRe RevIewS | Immunology
21. Irmler, M. et al. Granzyme A is an interleukin 1βconverting enzyme. J. Exp. Med. 181, 1917–1922
(1995).
22. Maelfait, J. et al. Stimulation of Toll-like receptor 3
and 4 induces interleukin-1β maturation by caspase-8.
J. Exp. Med. 205, 1967–1973 (2008).
23. Mayer-Barber, K. D. et al. Caspase-1 independent
IL-1β production is critical for host resistance to
Mycobacterium tuberculosis and does not require
TLR signaling in vivo. J. Immunol. 184, 3326–3330
(2010).
24. Kanneganti, T. D. Central roles of NLRs and
inflammasomes in viral infection. Nature Rev.
Immunol. 10, 688–698 (2010).
25. Sims, J. E. & Smith, D. E. The IL-1 family: regulators of
immunity. Nature Rev. Immunol. 10, 89–102 (2010).
26. Dinarello, C. A. Immunological and inflammatory
functions of the interleukin-1 family. Annu. Rev.
Immunol. 27, 519–550 (2009).
27. Hoshino, T. et al. Cutting edge: IL-18-transgenic mice:
in vivo evidence of a broad role for IL-18 in
modulating immune function. J. Immunol. 166,
7014–7018 (2001).
28. Nakanishi, K., Yoshimoto, T., Tsutsui, H. & Okamura, H.
Interleukin-18 regulates both Th1 and Th2 responses.
Annu. Rev. Immunol. 19, 423–474 (2001).
29. Weaver, C. T., Harrington, L. E., Mangan, P. R.,
Gavrieli, M. & Murphy, K. M. Th17: an effector CD4
T cell lineage with regulatory T cell ties. Immunity 24,
677–688 (2006).
30. Harrington, L. E. et al. Interleukin 17-producing CD4+
effector T cells develop via a lineage distinct from the
T helper type 1 and 2 lineages. Nature Immunol.
6, 1123–1132 (2005).
voLuMe 11 | MARcH 2011 | 219
© 2011 Macmillan Publishers Limited. All rights reserved
REVIEWS
31. Hoffman, H. M. et al. Efficacy and safety of rilonacept
(interleukin-1 trap) in patients with cryopyrinassociated periodic syndromes: results from two
sequential placebo-controlled studies. Arthritis
Rheum. 58, 2443–2452 (2008).
32. Lachmann, H. J. et al. Use of canakinumab in the
cryopyrin-associated periodic syndrome.
N. Engl. J. Med. 360, 2416–2425 (2009).
33. Lamkanfi, M. et al. Inflammasome-dependent
release of the alarmin HMGB1 in endotoxemia.
J. Immunol. 185, 4385–4392 (2010).
This paper describes the role of the NLRP3 and
NLRC4 inflammasomes in extracellular release of
HMGB1.
34. Miao, E. A. et al. Caspase-1-induced pyroptosis is
an innate immune effector mechanism against
intracellular bacteria. Nature Immunol. 11,
1136–1142 (2010).
This paper describes the crucial role of
caspase 1‑mediated cell death as a host defence
mechanism against bacterial pathogens.
35. Henry, T. & Monack, D. M. Activation of the
inflammasome upon Francisella tularensis infection:
interplay of innate immune pathways and virulence
factors. Cell. Microbiol. 9, 2543–2551 (2007).
This paper describes the increased susceptibility
of caspase 1‑deficient mice to infection with
Francisella tularensis compared with mice lacking
IL‑1β and IL‑18.
36. Trombetta, E. S. & Parodi, A. J. Quality control and
protein folding in the secretory pathway. Annu. Rev.
Cell. Dev. Biol. 19, 649–676 (2003).
37. Lee, M. C., Miller, E. A., Goldberg, J., Orci, L. &
Schekman, R. Bi-directional protein transport between
the ER and Golgi. Annu. Rev. Cell. Dev. Biol. 20,
87–123 (2004).
38. Nickel, W. & Rabouille, C. Mechanisms of regulated
unconventional protein secretion. Nature Rev. Mol.
Cell Biol. 10, 148–155 (2009).
39. Rubartelli, A., Cozzolino, F., Talio, M. & Sitia, R.
A novel secretory pathway for interleukin-1β, a protein
lacking a signal sequence. EMBO J. 9, 1503–1510
(1990).
40. Sutterwala, F. S. et al. Critical role for NALP3/CIAS1/
Cryopyrin in innate and adaptive immunity through
its regulation of caspase-1. Immunity 24, 317–327
(2006).
41. Keller, M., Ruegg, A., Werner, S. & Beer, H. D.
Active caspase-1 is a regulator of unconventional
protein secretion. Cell 132, 818–831 (2008).
This paper characterizes the role of caspase 1 in
the secretion of IL‑1α and FGF2.
42. Becker, C. E., Creagh, E. M. & O’Neill, L. A.
Rab39a binds caspase-1 and is required for
caspase-1-dependent interleukin-1β secretion.
J. Biol. Chem. 284, 34531–34537 (2009).
43. Salvesen, G. S. & Riedl, S. J. Caspase mechanisms.
Adv. Exp. Med. Biol. 615, 13–23 (2008).
44. Riedl, S. J. & Shi, Y. Molecular mechanisms of caspase
regulation during apoptosis. Nature Rev. Mol. Cell
Biol. 5, 897–907 (2004).
45. Lamkanfi, M. & Dixit, V. M. Manipulation of host cell
death pathways during microbial infections. Cell Host
Microbe 8, 44–54 (2010).
46. Strasser, A., O’Connor, L. & Dixit, V. M. Apoptosis
signaling. Annu. Rev. Biochem. 69, 217–245 (2000).
47. Taylor, R. C., Cullen, S. P. & Martin, S. J. Apoptosis:
controlled demolition at the cellular level. Nature Rev.
Mol. Cell Biol. 9, 231–241 (2008).
48. Kazama, H. et al. Induction of immunological
tolerance by apoptotic cells requires caspasedependent oxidation of high-mobility group box-1
protein. Immunity 29, 21–32 (2008).
49. Savill, J., Dransfield, I., Gregory, C. & Haslett, C.
A blast from the past: clearance of apoptotic cells
regulates immune responses. Nature Rev. Immunol.
2, 965–975 (2002).
50. Mariathasan, S. et al. Differential activation of the
inflammasome by caspase-1 adaptors ASC and Ipaf.
Nature 430, 213–218 (2004).
51. Lamkanfi, M. et al. Targeted peptidecentric
proteomics reveals caspase-7 as a substrate of the
caspase-1 inflammasomes. Mol. Cell. Proteomics
7, 2350–2363 (2008).
This paper describes caspase 7 as a downstream
effector of the NLRP3 and NLRC4 inflammasomes.
52. Fink, S. L. & Cookson, B. T. Caspase-1-dependent
pore formation during pyroptosis leads to osmotic
lysis of infected host macrophages. Cell. Microbiol.
8, 1812–1825 (2006).
53. Lamkanfi, M. et al. Glyburide inhibits the Cryopyrin/
Nalp3 inflammasome. J. Cell Biol. 187, 61–70 (2009).
54. Chen, Y., Smith, M. R., Thirumalai, K. & Zychlinsky, A.
A bacterial invasin induces macrophage apoptosis by
binding directly to ICE. EMBO J. 15, 3853–3860
(1996).
55. Hilbi, H. et al. Shigella-induced apoptosis is
dependent on caspase-1 which binds to IpaB.
J. Biol. Chem. 273, 32895–32900 (1998).
56. Monack, D. M., Detweiler, C. S. & Falkow, S.
Salmonella pathogenicity island 2-dependent
macrophage death is mediated in part by the host
cysteine protease caspase-1. Cell. Microbiol.
3, 825–837 (2001).
This paper reports that pyroptosis proceeds
independently of IL‑1β and IL‑18.
57. van der Velden, A. W., Velasquez, M. & Starnbach,
M. N. Salmonella rapidly kill dendritic cells via a
caspase-1-dependent mechanism. J. Immunol. 171,
6742–6749 (2003).
58. Suzuki, T. et al. Differential regulation of caspase-1
activation, pyroptosis, and autophagy via Ipaf and
ASC in Shigella-infected macrophages. PLoS Pathog.
3, e111 (2007).
59. Miao, E. A. et al. Cytoplasmic flagellin activates
caspase-1 and secretion of interleukin 1β via Ipaf.
Nature Immunol. 7, 569–575 (2006).
60. Franchi, L. et al. Cytosolic flagellin requires Ipaf for
activation of caspase-1 and interleukin 1β in
salmonella-infected macrophages. Nature Immunol.
7, 576–582 (2006).
61. Amer, A. et al. Regulation of Legionella phagosome
maturation and infection through flagellin and host
Ipaf. J. Biol. Chem. 281, 35217–35223 (2006).
62. Miao, E. A. et al. Innate immune detection of the
type III secretion apparatus through the NLRC4
inflammasome. Proc. Natl Acad. Sci. USA 107,
3076–3080 (2010).
63. Sutterwala, F. S. et al. Immune recognition of
Pseudomonas aeruginosa mediated by the IPAF/
NLRC4 inflammasome. J. Exp. Med. 204,
3235–3245 (2007).
64. Boyden, E. D. & Dietrich, W. F. Nalp1b controls mouse
macrophage susceptibility to anthrax lethal toxin.
Nature Genet. 38, 240–244 (2006).
65. Terra, J. K. et al. Cutting edge: resistance to Bacillus
anthracis infection mediated by a lethal toxin sensitive
allele of Nalp1b/Nlrp1b. J. Immunol. 184, 17–20
(2010).
66. Munoz-Planillo, R., Franchi, L., Miller, L. S. &
Nunez, G. A critical role for hemolysins and bacterial
lipoproteins in Staphylococcus aureus-induced
activation of the Nlrp3 inflammasome. J. Immunol.
183, 3942–3948 (2009).
67. Mariathasan, S. et al. Cryopyrin activates the
inflammasome in response to toxins and ATP.
Nature 440, 228–232 (2006).
68. Sauer, J. D. et al. Listeria monocytogenes triggers
AIM2-mediated pyroptosis upon infrequent
bacteriolysis in the macrophage cytosol.
Cell Host Microbe 7, 412–419 (2010).
69. Fernandes-Alnemri, T. et al. The AIM2 inflammasome
is critical for innate immunity to Francisella tularensis.
Nature Immunol. 11, 385–393 (2010).
70. Rathinam, V. A. et al. The AIM2 inflammasome is
essential for host defense against cytosolic bacteria
and DNA viruses. Nature Immunol. 11, 395–402
(2010).
71. Jones, J. W. et al. Absent in melanoma 2 is required
for innate immune recognition of Francisella
tularensis. Proc. Natl Acad. Sci. USA 107,
9771–9776 (2010).
72. Wu, J., Fernandes-Alnemri, T. & Alnemri, E. S.
Involvement of the AIM2, NLRC4, and NLRP3
inflammasomes in caspase-1 activation by Listeria
monocytogenes. J. Clin. Immunol. 30, 693–702
(2010).
73. Warren, S. E. et al. Cutting edge: cytosolic bacterial
DNA activates the inflammasome via Aim2.
J. Immunol. 185, 818–821 (2010).
74. Tsuchiya, K. et al. Involvement of absent in melanoma 2
in inflammasome activation in macrophages infected
with Listeria monocytogenes. J. Immunol. 185,
1186–1195 (2010).
75. Mariathasan, S., Weiss, D. S., Dixit, V. M. &
Monack, D. M. Innate immunity against Francisella
tularensis is dependent on the ASC/caspase-1 axis.
J. Exp. Med. 202, 1043–1049 (2005).
76. Monack, D. M., Raupach, B., Hromockyj, A. E. &
Falkow, S. Salmonella typhimurium invasion induces
220 | MARcH 2011 | voLuMe 11
77.
78.
79.
80.
81.
82.
83.
84.
85.
86.
87.
88.
89.
90.
91.
92.
93.
apoptosis in infected macrophages. Proc. Natl Acad.
Sci. USA 93, 9833–9838 (1996).
Fernandes-Alnemri, T. et al. The pyroptosome: a
supramolecular assembly of ASC dimers mediating
inflammatory cell death via caspase-1 activation.
Cell Death Differ. 14, 1590–1604 (2007).
Malireddi, R. K., Ippagunta, S., Lamkanfi, M. &
Kanneganti, T. D. Cutting edge: proteolytic inactivation
of poly(ADP-ribose) polymerase 1 by the Nlrp3 and
Nlrc4 inflammasomes. J. Immunol. 185, 3127–3130
(2010).
This paper describes the inflammasome‑dependent
cleavage of PARP1 during pyroptosis.
Akhter, A. et al. Caspase-7 activation by the Nlrc4/Ipaf
inflammasome restricts Legionella pneumophila
infection. PLoS Pathog. 5, e1000361 (2009).
This paper describes the role of caspase 7
activation by the NLRC4 inflammasome in
restricting Legionella replication.
Lakhani, S. A. et al. Caspases 3 and 7: key mediators
of mitochondrial events of apoptosis. Science 311,
847–851 (2006).
Shao, W., Yeretssian, G., Doiron, K., Hussain, S. N. &
Saleh, M. The caspase-1 digestome identifies the
glycolysis pathway as a target during infection and
septic shock. J. Biol. Chem. 282, 36321–36329
(2007).
This paper describes glycolysis enzymes as
substrates of caspase 1.
Cramer, T. et al. HIF-1α is essential for myeloid cellmediated inflammation. Cell 112, 645–657 (2003).
Gurcel, L., Abrami, L., Girardin, S., Tschopp, J. &
van der Goot, F. G. Caspase-1 activation of lipid
metabolic pathways in response to bacterial poreforming toxins promotes cell survival. Cell 126,
1135–1145 (2006).
This paper reports that inflammasomes activate
lipid metabolic pathways.
Gonzalez, M. R., Bischofberger, M., Pernot, L.,
van der Goot, F. G. & Freche, B. Bacterial pore-forming
toxins: the (w)hole story? Cell. Mol. Life Sci. 65,
493–507 (2008).
McCoy, A. J. et al. Cytotoxins of the human pathogen
Aeromonas hydrophila trigger, via the NLRP3
inflammasome, caspase-1 activation in macrophages.
Eur. J. Immunol. 40, 2797–2803 (2010).
Lamkanfi, M. et al. The Nod-like receptor family
member Naip5/Birc1e restricts Legionella
pneumophila growth independently of caspase-1
activation. J. Immunol. 178, 8022–8027 (2007).
Zamboni, D. S. et al. The Birc1e cytosolic patternrecognition receptor contributes to the detection and
control of Legionella pneumophila infection. Nature
Immunol. 7, 318–325 (2006).
Broz, P. et al. Redundant roles for inflammasome
receptors NLRP3 and NLRC4 in host defense
against Salmonella. J. Exp. Med. 207, 1745–1755
(2010).
Fischer, U., Janicke, R. U. & Schulze-Osthoff, K.
Many cuts to ruin: a comprehensive update of
caspase substrates. Cell Death Differ. 10, 76–100
(2003).
Lamkanfi, M., Festjens, N., Declercq, W.,
Vanden Berghe, T. & Vandenabeele, P. Caspases in cell
survival, proliferation and differentiation. Cell Death
Differ. 14, 44–55 (2007).
Boatright, K. M. et al. A unified model for apical
caspase activation. Mol. Cell 11, 529–541 (2003).
Martinon, F. & Tschopp, J. Inflammatory caspases
and inflammasomes: master switches of inflammation.
Cell Death Differ. 14, 10–22 (2007).
Pizzirani, C. et al. Stimulation of P2 receptors causes
release of IL-1β-loaded microvesicles from human
dendritic cells. Blood 109, 3856–3864 (2007).
Acknowledgements
This work was supported by European Union Framework
Program 7 (Marie Curie grant 256432) and by the Fund for
Scientific Research – Flanders.
Competing interests statement
The author declares no competing financial interests.
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