4.1. NLRP1
NLR Family Pyrin Domain Containing 1 (NLRP1) is an inflammasome sensor sensing cytosolic proteolytic activity. Anthrax lethal factor, a toxin with endoprotease activity, and IpaH7.8, a
Shigella flexneri effector with ubiquitin ligase activity, trigger NLRP1B degradation and activation [
28,
29]. Human NLRP1, the three murine paralogues NLRP1A, B, and C, each one with different isoforms in different mouse strains, detect different stimuli [
29].
In mice,
Nlrp1a expression is restricted to the hematopoietic compartment and is expressed in hematopoietic stem cells, progenitor cells of both myeloid and lymphoid origins, and terminally differentiated cells such as macrophages [
30].
Expression of
Nlrp1a and
Nlrp1c is positively regulated by SREBP-1A (Sterol regulatory element binding protein-1a), a basic helix–loop–helix leucine zipper (bHLH-LZ) TF [
31]. SREBP-1A binds directly onto a canonical SREBP-1 binding site in the
Nlrp1a proximal promoter [
31].
Srebp-1a expression is itself under the direct control of NF-κB, in synergy with the monocyte/macrophage specific TF PU.1 (an Erythroblast Transformation Specific (ETS) family TF). NF-κB thus indirectly controls
Nlrp1a expression in macrophages (
Figure 2A) [
31]. SREBP-1A also controls LPS-triggered lipogenesis in macrophages, an anabolic pathway required for optimal inflammasome responses. SREBP-1A regulation thus directly couples lipogenesis and control of the NLRP1A inflammasome [
31]. SREBP TFs being key signaling nodes responding to metabolic clues, the control of
Nlrp1a expression might thus participate to the control of metabolic inflammation. Interestingly, mice lacking the three murine
Nlrp1 alleles (or
Il18) develop metabolic syndrome and spontaneous obesity strengthening the functional link between metabolism and
Nlrp1 sensors [
32].
In humans,
NLRP1 is broadly expressed but at particular high levels in keratinocytes, a feature that is not observed in mice [
33]. The molecular basis of the differential expression of
NLRP1 between humans and mice remains unknown. It is controlled at the transcriptional level since inflammasome transcripts are very low in murine keratinocytes [
34].
Endoplasmic reticulum (ER) stress strongly induces
NLRP1 expression in THP-1 monocytes and in various human cell lines. This induction is dependent on two effectors of the unfolded protein response (UPR): PKR-like ER protein kinase (PERK) and Inositol-requiring enzyme 1 (IRE1). Although the mechanisms downstream of IRE1 remain unsolved, PERK activation leads to Activating Transcription Factor 4 (ATF4) expression. ATF4 is a TF from the ATF/cAMP Response Element-binding protein (CREB) family. ATF4 binds to the promoter of human
NLRP1 to induce its expression (
Figure 2A) [
35]. It is thus tempting to speculate that ER stress could prime the NLRP1 inflammasome to increase its ability to detect HAMPs and PAMPs.
4.2. NLRP3
NLRP3 is an inflammasome sensor detecting multiple cellular homeostasis perturbations such as membrane damage, mitochondrial defects, or perturbations of ionic concentrations. NLRP3 inflammasome activation is a two-step process and requires a signal 1 (priming signal, e.g., LPS) and a signal 2 (activation signal, e.g., nigericin). Priming is triggered by pro-inflammatory signals such as Toll-like receptor 4 (TLR4) engagement and has long been associated with transcriptional upregulation of
NLRP3 [
36,
37]. It is now clear that NLRP3 priming is mediated by post-translational modifications independent of transcription (
Figure 2B) [
38,
39,
40]. Yet,
Nlrp3 upregulation accelerates the kinetics and the level of caspase-1 activation following signal 2 addition [
38]. The importance of this upregulation is likely enhanced by the low level of expression of
Nlrp3 at steady state.
NLRP3 upregulation occurs within 2 h of LPS addition but can also be triggered by other TLR ligands, by the nucleotide-binding oligomerization domain 2 (Nod2) ligand (muramyl dipeptide) or by pro-inflammatory cytokines (Tumor necrosis factor (TNF), Interleukin (IL)-1α, IL-1β) in the absence of PAMPs [
36,
37]. This upregulation is dependent on NF-κB as first demonstrated using the BAY 11-7082 inhibitor. Two NF-κB binding sites are present in the
NLRP3 promoter. Upon LPS treatment, the NF-κB subunit RelA/p65 binds the
NLRP3 promoter. LPS-induced
NLRP3 upregulation is lost upon mutation of these two binding sites indicating that these two sites control in a redundant manner the increase in
NLRP3 promoter activity [
41]. Upregulation of
Nlrp3 (and
Il1b) following NF-κB activation is partially dependent on an atypical IκBs (inhibitor of κB), a coactivator of NF-κB in bone marrow-derived macrophages (BMDMs) [
42]. IκBζ binds to another NF-κB subunit, p50, in the
Nlrp3 promoter. The NF-κB heterodimer RelA/p50 may thus be responsible for LPS-induced
Nlrp3 expression. IκBζ recruitment increases H3K4me3, a mark of active transcription [
43] suggesting that this coactivator promotes
Nlrp3 induction through epigenetic modifications. Interestingly, the parasite
Leishmania amazonensis subverts this process by targeting the epigenetic control of NF-κB-related pro-inflammatory genes. During infection, the promoters of these genes display hypoacetylation of the histone H3K9/14 and hypotrimethylation of histone H3K4. These inhibitory epigenetic modifications dampen
Nlrp3 (and also
Aim2,
Nlrc4,
Pycard, Il1b, and
Il18) expression and promotes survival of the parasite within the host [
44].
In addition to its control by pro-inflammatory signals,
Nlrp3 expression is controlled by the circadian clock and presents a peak of expression during the night [
45,
46]. The circadian rhythm in
Nlrp3 transcript level is dependent on a master regulator of the clock, the transcription factor NR1D1 (nuclear receptor subfamily 1 group D member 1, also known as Rev-erbα). Circadian oscillations of
Nlrp3 and
Nr1d1 expressions are in antiphase. Accordingly, NR1D1 binds directly to a Rev-response element (RevRE) in the
Nlrp3 promoter to repress its expression (
Figure 2B). Interestingly, an agonist of NR1D1 attenuates both dextran sodium sulfate (DSS)-induced colitis and D-galactosamine-induced fulminant hepatitis suggesting that NR1D1-mediated circadian control of
Nlrp3 expression controls inflammation in vivo [
45,
46].
Besides NF-κB and NR1D1, direct binding of several TFs on the
NLRP3 promoter have been reported with either negative or positive impact on
NLRP3 expression. Downregulation of
NLRP3 expression is controlled by the aryl hydrocarbon receptor (AhR) [
47] and by B cell lymphoma 6 (BCL6), a transcriptional repressor that antagonizes NF-κB-mediated gene transcription [
48]. AhR is a ligand-activated TF binding numerous environmental contaminants (e.g., dioxin) and endogenous ligands (e.g., the tryptophan derivative, kynurenine). AhR binds two XRE (Xenobiotic Responsive Element) sequences in the
Nrlp3 promoter, and its binding is increased in a ligand-dependent manner. The two XRE sites surround the two NF-κB sites in the
Nlrp3 promoter suggesting that AhR may interfere with NF-κB recruitment either directly or indirectly by modifying the local chromatin architecture. AhR and its ligands also negatively control
Il1b expression indicating that environmental pollutants (and the anti-inflammatory tryptophan catabolites) dampen inflammasome responses (
Figure 2B) [
47].
Growth Factor Independence 1 (GFI1), a protein induced by NF-κB, binds the
Nlrp3 promoter to inhibit
Nlrp3 expression in a negative feedback loop [
49]. In contrast, direct binding of Nuclear factor of activated T cells 5 (NFAT5—a transcription factor induced in response to high salt, hypoxia and mechanical stress) on an osmotic response element (ORE) in the promoter of
Nlrp3 positively regulates
Nlrp3 expression [
50]. Overall, regulation of
Nlrp3 expression emerges as a complex network primarily driven by pro-inflammatory signals but integrating environmental and intrinsic signals.
Nlrp3 expression has been mostly studied in macrophages and displays a cell-type-specific regulation in dendritic cells (DCs). Of note, the DC-specific gene regulation affects not only
Nlrp3 but also other inflammasome genes as detailed below. Indeed, plasmacytoid dendritic cells (pDC) express neither
Nlrp3 nor
Il1b and are resistant to inflammasome activation [
51]. Conventional DCs (cDC) have a limited ability to respond to inflammasome stimuli [
21,
51]. Interferon Regulatory Factors (IRFs) belong to a family of TFs, which includes 9 members in human and mice. pDCs express high levels of IRF8, while cDC1 and cDC2 express high levels of IRF8 and IRF4, respectively. IRF4 and IRF8 bind the promoter regions of
Nlrp3,
Il1b, and
Aim2 and intronic regions of
Pycard and
Nlrc4, which correlate with the low expression of these genes in cDC1 and cDC2. Haploinsufficiency in
Irf8 increases
Nlrc4,
Pycard, and
Il1b expression in cDC1 and increases the inflammasome response to the NLRC4-engaging pathogen
Salmonella typhimurium (
S. typhimurium). In contrast, ectopic expression of
Irf8 in macrophages decreases
Nlrp3,
Nlrc4, and
il1b expression, while ectopic expression of
Irf4 decreases
Nlrc4,
Il1b, and
Pycard expression [
21]. Importantly, the low inflammasome activity in cDC limits their pyroptosis in response to bacterial pathogens and favors antigen presentation and T cell priming.
4.3. NAIP and NLRC4
The Neuronal apoptosis inhibitory protein (NAIP)/NLRC4 inflammasome is atypical since NLRC4 is not a direct sensor but requires a NAIP protein to sense PAMPs. A single NAIP protein is encoded in the human genome and binds bacterial type III secretion system (T3SS) needle proteins [
52] and flagellin [
53]. Several NAIP paralogs are present in mice and bind T3SS needle proteins (NAIP1), T3SS inner rod proteins (NAIP2), or flagellin (NAIP5/6) [
54,
55].
IRF8 was identified in mice as a positive regulator of
Naip2, 5, 6, and
Nlrc4. IRF8 binds the promoter regions of
Naip2, 5, 6, and an intronic region of
Nlrc4. IRF8 has a very low intrinsic DNA-binding activity and, at steady state, binds with PU.1 at Ets-IRF composite elements (EICE) (
Figure 2C) [
56]. NAIP/NLRC4 inflammasomes are the major inflammasomes allowing mice to control
S. typhimurium infection. Accordingly,
Irf8−/− mice are highly susceptible to
S. typhimurium [
25]. Interestingly, although the effects of IRF8 were demonstrated at steady state,
Irf8 is induced upon
Legionella infection and could participate in the NAIP/NLRC4-dependent response against this flagellin-expressing pathogen [
57]. Surprisingly, overexpression of
Irf8 in bone marrow-derived macrophages decreases
Nlrc4 expression. Furthermore, as presented above, in cDC1, IRF8 inhibits
Nlrc4 expression (
Figure 2C) [
21]. IRF8 concentration modulates its ability to cooperate with other TFs and engage different binding sites [
58] possibly explaining its reported opposed role on
Nlrc4 at steady state and upon overexpression.
4.4. AIM2
AIM2 is a receptor of cytosolic DNA [
59,
60]. Since, in healthy cells, DNA is restricted to the nucleus, the presence of cytosolic DNA is either indicative of an infection or of a cellular stress associated with the loss of nuclear membrane integrity [
61].
AIM2 is an IFN-inducible gene. In murine BMDMs,
Aim2 is expressed at steady state and its expression is slightly induced by IFN, poly(dA:dT) treatment, or infections [
60]. In mice, AIM2 levels are sufficient to promote its inflammasome functions, and the AIM2 inflammasome does not require a priming step [
62,
63]. Its role in myeloid human cells is less clear than in the murine context. Indeed, a cGAS-STING-lysosomal-NLRP3 pathway triggers cell death in response to cytosolic DNA in human myeloid cells [
64], while in the presence of IFN-γ, AIM2 is involved in
T. gondii responses. These results suggest that AIM2 functionality in human cells is variable depending on the context [
65]. In contrast to murine macrophages, AIM2 levels are very low at steady state in human macrophages but are strongly induced by LPS [
66] or IFN-γ [
65]. The upregulation of
AIM2 transcript in inflammatory conditions is likely relevant in human diseases since
AIM2 expression level is increased in keratinocytes from psoriatic lesions compared to healthy skin [
67].
In human cells,
AIM2 expression is dynamically controlled by B lymphocyte-induced maturation protein-1 (BLIMP1 also known as PR domain zinc finger protein 1-PRDM1), IRF1/2 and Signal Transducer and Activator of Transcription 1 (STAT1), a transcription factor activated by interferons [
68,
69]. BLIMP1 binding site overlaps with IRF1/2 binding site and these transcription factors compete to repress or activate
AIM2 expression, respectively. Indeed,
BLIMP1 knock-down increases IRF1/2 binding on the
AIM2 promoter and AIM2 expression (
Figure 2D) [
68]. To our knowledge, it is unknown whether BLIMP1 modulation may be involved in licensing AIM2 function in certain human cell types. In addition, two tandem Gamma-activated site (GAS) sequences are present 220 nt upstream of
AIM2 TSS. These GAS belong to the long terminal repeat (LTR) sequence of an endogenous retrovirus termed
MER41. Treatment with IFN-γ triggers an increase in H3K27ac and STAT1 binding at the
MER41.AIM2 site. Accordingly,
MER41 sequence is required for
AIM2 upregulation in HeLa cells exposed to IFN-γ. The conservation of the
MER41.AIM2 site across anthropoid primates (but not in mice) suggests that this retrovirus sequence was co-opted for
AIM2 regulation in an ancestor of anthropoid primates [
69] and illustrates how past infections have shaped the species-specific regulation of inflammasome genes. Interestingly, a
MER41 sequence displaying IRF1 and STAT1 binding in monocytes is also found in proximity of
GSDMD, suggesting that the regulation of several inflammasome genes may result from retrovirus integration.
4.5. Pyrin
Pyrin is a sensor detecting Rho GTPase inhibition. Rho GTPases are altered by numerous bacterial toxins and bacterial effectors. Pyrin thus acts as a guard of this important cytoskeleton regulator [
70]. Gain of function mutations in
MEFV, the gene encoding Pyrin, cause several inflammatory syndromes, including Familial Mediterranean Fever (FMF) [
71].
Pyrin is constitutively expressed in human neutrophils, monocytes, and M-CSF-differentiated monocyte-derived macrophages [
72]. In monocytes,
MEFV expression is positively regulated by IFN-α, IFN-γ, and TNF, while IL-4 and IL-10 repress it [
73,
74]. TNF-dependent upregulation of
MEFV expression is due to the binding of CCAAT/enhancer binding protein β (C/EBPβ) in the
MEFV promoter, which acts in synergy with NF-κB p65/RelA subunit. Indeed, NF-κB p65/RelA binds the
MEFV promoter both through a canonical NF-κB binding site and indirectly through its binding to C/EBP
β (
Figure 2E) [
75]. The mechanism underlying IFN-mediated upregulation is unclear although IFN-stimulated response element (ISRE) and GAS consensus sequences have been identified in silico in the
MEFV promoter [
73].
In mice,
Mefv expression is induced by TLR ligands and inflammatory cytokines (IFN-β + TNF). TNF signaling contribution to inflammation in a murine model of FMF strongly suggests that transcriptional regulation of inflammasome sensors is important not only for the detection of pathogens but also for the prevention of deleterious inflammatory reactions [
76].