Product: NLRP3 Antibody
Catalog: DF7438
Source: Rabbit
Application: WB, IHC, IF/ICC, ELISA(peptide)
Reactivity: Human, Mouse, Rat
Prediction: Pig, Bovine, Horse, Sheep, Rabbit, Dog
Mol.Wt.: 80~120kD; 118kD(Calculated).
Uniprot: Q96P20
RRID: AB_2839376

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Product Info

Source:
Rabbit
Application:
WB 1:500-1:2000, IHC 1:50-1:200, IF/ICC 1:100-1:500, ELISA(peptide) 1:20000-1:40000
*The optimal dilutions should be determined by the end user.
Reactivity:
Human,Mouse,Rat
Prediction:
Pig(90%), Bovine(90%), Horse(90%), Sheep(90%), Rabbit(90%), Dog(89%)
Clonality:
Polyclonal
Specificity:
NLRP3 Antibody detects endogenous levels of total NLRP3.
RRID:
AB_2839376
Cite Format: Affinity Biosciences Cat# DF7438, RRID:AB_2839376.
Purification:
The antiserum was purified by peptide affinity chromatography using SulfoLink™ Coupling Resin (Thermo Fisher Scientific).
Storage:
Rabbit IgG in phosphate buffered saline , pH 7.4, 150mM NaCl, 0.02% sodium azide and 50% glycerol. Store at -20 °C. Stable for 12 months from date of receipt.
Alias:

Fold/Unfold

AGTAVPRL; AII/AVP; Angiotensin/vasopressin receptor AII/AVP like; Angiotensin/vasopressin receptor AII/AVP-like; C1orf7; Caterpiller protein 1.1; CIAS 1; CIAS1; CLR1.1; Cold autoinflammatory syndrome 1; Cold autoinflammatory syndrome 1 protein; Cryopyrin; Familial cold autoinflammatory syndrome; FCAS; FCU; LRR and PYD domains-containing protein 3; Muckle-Wells syndrome; MWS; NACHT; NACHT LRR and PYD containing protein 3; NALP 3; NALP3; NALP3_HUMAN; NLR family pyrin domain containing 3; NLRP3; PYPAF 1; PYPAF1; PYRIN containing APAF1 like protein 1; PYRIN-containing APAF1-like protein 1;

Immunogens

Immunogen:
Uniprot:
Gene(ID):
Expression:
Q96P20 NLRP3_HUMAN:

Predominantly expressed in macrophages. Also expressed in dendritic cells, B- and T-cells (at protein level) (PubMed:11786556) (PubMed:17164409). Expressed in LPS-treated granulocytes, but not in resting cells (at protein level) (PubMed:17164409). Expression in monocytes is very weak (at protein level) (PubMed:17164409). Expressed in stratified non-keratinizing squamous epithelium, including oral, esophageal and ectocervical mucosa and in the Hassall's corpuscles in the thymus. Also, detected in the stratified epithelium covering the bladder and ureter (transitional mucosa) (at protein level) (PubMed:17164409). Expressed in lung epithelial cells (at protein level) (PubMed:23229815). Expressed in chondrocytes (PubMed:12032915). Expressed at low levels in resting osteoblasts (PubMed:17907925).

Description:
This gene encodes a pyrin-like protein containing a pyrin domain, a nucleotide-binding site (NBS) domain, and a leucine-rich repeat (LRR) motif. This protein interacts with the apoptosis-associated speck-like protein PYCARD/ASC, which contains a caspase recruitment domain, and is a member of the NALP3 inflammasome complex. This complex functions as an upstream activator of NF-kappaB signaling, and it plays a role in the regulation of inflammation, the immune response, and apoptosis. Mutations in this gene are associated with familial cold autoinflammatory syndrome (FCAS), Muckle-Wells syndrome (MWS), chronic infantile neurological cutaneous and articular (CINCA) syndrome, and neonatal-onset multisystem inflammatory disease (NOMID). Multiple alternatively spliced transcript variants encoding distinct isoforms have been identified for this gene. Alternative 5' UTR structures are suggested by available data; however, insufficient evidence is available to determine if all of the represented 5' UTR splice patterns are biologically valid.
Sequence:
MKMASTRCKLARYLEDLEDVDLKKFKMHLEDYPPQKGCIPLPRGQTEKADHVDLATLMIDFNGEEKAWAMAVWIFAAINRRDLYEKAKRDEPKWGSDNARVSNPTVICQEDSIEEEWMGLLEYLSRISICKMKKDYRKKYRKYVRSRFQCIEDRNARLGESVSLNKRYTRLRLIKEHRSQQEREQELLAIGKTKTCESPVSPIKMELLFDPDDEHSEPVHTVVFQGAAGIGKTILARKMMLDWASGTLYQDRFDYLFYIHCREVSLVTQRSLGDLIMSCCPDPNPPIHKIVRKPSRILFLMDGFDELQGAFDEHIGPLCTDWQKAERGDILLSSLIRKKLLPEASLLITTRPVALEKLQHLLDHPRHVEILGFSEAKRKEYFFKYFSDEAQARAAFSLIQENEVLFTMCFIPLVCWIVCTGLKQQMESGKSLAQTSKTTTAVYVFFLSSLLQPRGGSQEHGLCAHLWGLCSLAADGIWNQKILFEESDLRNHGLQKADVSAFLRMNLFQKEVDCEKFYSFIHMTFQEFFAAMYYLLEEEKEGRTNVPGSRLKLPSRDVTVLLENYGKFEKGYLIFVVRFLFGLVNQERTSYLEKKLSCKISQQIRLELLKWIEVKAKAKKLQIQPSQLELFYCLYEMQEEDFVQRAMDYFPKIEINLSTRMDHMVSSFCIENCHRVESLSLGFLHNMPKEEEEEEKEGRHLDMVQCVLPSSSHAACSHGLVNSHLTSSFCRGLFSVLSTSQSLTELDLSDNSLGDPGMRVLCETLQHPGCNIRRLWLGRCGLSHECCFDISLVLSSNQKLVELDLSDNALGDFGIRLLCVGLKHLLCNLKKLWLVSCCLTSACCQDLASVLSTSHSLTRLYVGENALGDSGVAILCEKAKNPQCNLQKLGLVNSGLTSVCCSALSSVLSTNQNLTHLYLRGNTLGDKGIKLLCEGLLHPDCKLQVLELDNCNLTSHCCWDLSTLLTSSQSLRKLSLGNNDLGDLGVMMFCEVLKQQSCLLQNLGLSEMYFNYETKSALETLQEEKPELTVVFEPSW

Predictions

Predictions:

Score>80(red) has high confidence and is suggested to be used for WB detection. *The prediction model is mainly based on the alignment of immunogen sequences, the results are for reference only, not as the basis of quality assurance.

Species
Results
Score
Pig
90
Horse
90
Bovine
90
Sheep
90
Rabbit
90
Dog
89
Xenopus
0
Zebrafish
0
Chicken
0
Model Confidence:
High(score>80) Medium(80>score>50) Low(score<50) No confidence

PTMs - Q96P20 As Substrate

Site PTM Type Enzyme
Ubiquitination
S5 Phosphorylation
Y13 Phosphorylation
S161 Phosphorylation
S163 Phosphorylation
S198 Phosphorylation
T233 Phosphorylation
S295 Phosphorylation
S334 Phosphorylation
S387 Phosphorylation
S436 Phosphorylation
K496 Ubiquitination
S728 Phosphorylation
Y861 Phosphorylation
S975 Phosphorylation

Research Backgrounds

Function:

As the sensor component of the NLRP3 inflammasome, plays a crucial role in innate immunity and inflammation. In response to pathogens and other damage-associated signals, initiates the formation of the inflammasome polymeric complex, made of NLRP3, PYCARD and CASP1 (and possibly CASP4 and CASP5). Recruitment of proCASP1 to the inflammasome promotes its activation and CASP1-catalyzed IL1B and IL18 maturation and secretion in the extracellular milieu. Activation of NLRP3 inflammasome is also required for HMGB1 secretion. The active cytokines and HMGB1 stimulate inflammatory responses. Inflammasomes can also induce pyroptosis, an inflammatory form of programmed cell death. Under resting conditions, NLRP3 is autoinhibited. NLRP3 activation stimuli include extracellular ATP, reactive oxygen species, K(+) efflux, crystals of monosodium urate or cholesterol, amyloid-beta fibers, environmental or industrial particles and nanoparticles, cytosolic dsRNA, etc. However, it is unclear what constitutes the direct NLRP3 activator. Activation in presence of cytosolic dsRNA is mediated by DHX33. Independently of inflammasome activation, regulates the differentiation of T helper 2 (Th2) cells and has a role in Th2 cell-dependent asthma and tumor growth (By similarity). During Th2 differentiation, required for optimal IRF4 binding to IL4 promoter and for IRF4-dependent IL4 transcription. Binds to the consensus DNA sequence 5'-GRRGGNRGAG-3'. May also participate in the transcription of IL5, IL13, GATA3, CCR3, CCR4 and MAF (By similarity).

PTMs:

The disulfide bond in the pyrin domain might play a role in reactive oxygen species-mediated activation.

Ubiquitinated; undergoes both 'Lys-48'- and 'Lys-63'-linked polyubiquitination. Ubiquitination does not lead to degradation, but inhibits inflammasome activation (By similarity). Deubiquitination is catalyzed by BRCC3 and associated with NLRP3 activation and inflammasome assembly. This process can be induced by the activation of Toll-like receptors (by LPS), through a non-transcriptional pathway dependent on the mitochondrial production of reactive oxygen species, and by ATP.

Subcellular Location:

Cytoplasm>Cytosol. Inflammasome. Endoplasmic reticulum. Secreted. Nucleus.
Note: In macrophages, under resting conditions, mainly located in the cytosol, on the endoplasmic reticulum. After stimulation with inducers of the NLRP3 inflammasome, mitochondria redistribute in the vicinity of the endoplasmic reticulum in the perinuclear region, which results in colocalization of NLRP3 on the endoplasmic reticulum and PYCARD on mitochondria, allowing the activation of inflammasome assembly. After the induction of pyroptosis, inflammasome specks are released into the extracellular space where they can further promote IL1B processing and where they can be engulfed by macrophages. Phagocytosis induces lysosomal damage and inflammasome activation in the recipient cells (PubMed:24952504). In the Th2 subset of CD4(+) helper T-cells, mainly located in the nucleus. Nuclear localization depends upon KPNA2. In the Th1 subset of CD4(+) helper T-cells, mainly cytoplasmic (By similarity).

Golgi apparatus membrane.
Note: (Microbial infection) Upon HRSV infection, the protein is mainly located in lipid rafts in the Golgi membrane.

Extracellular region or secreted Cytosol Plasma membrane Cytoskeleton Lysosome Endosome Peroxisome ER Golgi apparatus Nucleus Mitochondrion Manual annotation Automatic computational assertionSubcellular location
Tissue Specificity:

Predominantly expressed in macrophages. Also expressed in dendritic cells, B- and T-cells (at protein level). Expressed in LPS-treated granulocytes, but not in resting cells (at protein level). Expression in monocytes is very weak (at protein level). Expressed in stratified non-keratinizing squamous epithelium, including oral, esophageal and ectocervical mucosa and in the Hassall's corpuscles in the thymus. Also, detected in the stratified epithelium covering the bladder and ureter (transitional mucosa) (at protein level). Expressed in lung epithelial cells (at protein level). Expressed in chondrocytes. Expressed at low levels in resting osteoblasts.

Subunit Structure:

Sensor component of NLRP3 inflammasomes. Inflammasomes are supramolecular complexes that assemble in the cytosol in response to pathogens and other damage-associated signals and play critical roles in innate immunity and inflammation. The core of NLRP3 inflammasomes consists of a signal sensor component (NLRP3), an adapter (ASC/PYCARD), which recruits an effector proinflammatory caspase (CASP1 and, possibly, CASP4 and CASP5). Within the complex, NLRP3 and PYCARD interact via their respective DAPIN/pyrin domains. This interaction initiates speck formation (nucleation) which greatly enhances further addition of soluble PYCARD molecules to the speck in a prion-like polymerization process. NLRP3 localizes at the end of each PYCARD filament. Clustered PYCARD nucleates the formation of CASP1 filaments through the interaction of their respective CARD domains, acting as a platform for CASP1 polymerization. CASP1 filament formation increases local enzyme concentration, resulting in trans-autocleavage and activation. Active CASP1 then processes IL1B and IL18 precursors, leading to the release of mature cytokines in the extracellular milieu and inflammatory response. Reconstituted ternary inflammasomes show star-shaped structures, in which multiple filaments, containing CASP1, protrude radially from a single central hub, containing the sensor protein and PYCARD. In this complex, the sensor protein is sub-stoichiometric to PYCARD, and PYCARD is further substoichiometric to CASP1, suggesting amplifications of signal transduction from the sensor, via the adapter, to the effector. Interacts with MEFV; this interaction targets NLRP3 to degradation by autophagy, hence preventing excessive IL1B- and IL18-mediated inflammation. Interacts with GBP5 (via DAPIN domain); this interaction promotes inflammasome assembly in response to microbial and soluble, but not crystalline, agents. Interacts with EIF2AK2/PKR; this interaction requires EIF2AK2 activity, is accompanied by EIF2AK2 autophosphorylation and promotes inflammasome assembly in response to specific stimuli. Interacts with PML (isoform PML-1) (via the leucine-rich repeat (LRR) domain); PML-mediated increase in NLRP3 inflammasome activation does not depend upon this interaction. Directly interacts with IRF4 (via the LRR domain); this interaction is required for optimal IRF4 binding to IL4 promoter and efficient IL4 transactivation during differentiation of Th2 helper T-cells (By similarity). Interacts (via NACHT domain) with DHX33 (via DEAH box). Interacts with PYDC5.

Family&Domains:

The pyrin domain (also called DAPIN domain or PYD) is involved in PYCARD-binding.

The LRR domain mediates the interaction with IRF4 and PML.

Intramolecular interactions between NACHT and leucine-rich repeat (LRR) domains may be important for autoinhibition in the absence of activating signal.

Belongs to the NLRP family.

Research Fields

· Cellular Processes > Cell growth and death > Necroptosis.   (View pathway)

· Human Diseases > Infectious diseases: Bacterial > Pertussis.

· Human Diseases > Infectious diseases: Viral > Influenza A.

· Organismal Systems > Immune system > NOD-like receptor signaling pathway.   (View pathway)

References

1). Li X et al. Activation of NLRP3 in microglia exacerbates diesel exhaust particles-induced impairment in learning and memory in mice. Environ Int 2020 Mar;136:105487 (PubMed: 31999974) [IF=13.352]

Application: WB    Species: mouse    Sample: BV2 cells

Fig. 5. Reactive oxygen species (ROS) generation, inflammatory markers, and metabolomics analyses of diesel exhaust particles (DEPs)-treated BV2 microglia in vitro. C: Protein levels of molecules involved in the NLRP3 pathway increased following DEPs treatment compared with those in control cells. One-way ANOVA followed by Turkey’s test, n = 3 biological replicates/group.

2). Su P et al. The role of NLRP3 in lead-induced neuroinflammation and possible underlying mechanism. Environ Pollut 2021 Oct 15;287:117520. (PubMed: 34182382) [IF=9.988]

3). Liu X et al. Combination of resolvin E1 and lipoxin A4 promotes the resolution of pulpitis by inhibiting NF-κB activation through upregulating sirtuin 7 in dental pulp fibroblasts. Cell Prolif 2022 Apr 11;e13227. (PubMed: 35411569) [IF=8.755]

Application: WB    Species: rat    Sample: Dental pulp fibroblasts

FIGURE 1 |Effects of combined administration of RvE1 and LXA4 on pro-inflammatory factor expression.(C) NLRP3, (D) caspase-1, (E) IL-1β and (F) IL-18 mRNA levels on LPS-induced DPFs detected by qPCR and (G) their protein levels tested by western blotting (normalized to that of β-tubulin).

4). Li XX et al. Protective effects of acarbose against vascular endothelial dysfunction through inhibiting Nox4/NLRP3 inflammasome pathway in diabetic rats. Free Radic Biol Med 2019 Sep 18 (PubMed: 31541678) [IF=8.101]

Application: WB    Species: rat    Sample: RAECs

Fig.1. |Acarbose inhibited HG-induced NLRP3 inflammasome activation in RAECs .RAECs were incubated with 30mM glucose for 24, 48, 72 hours. Summarized data showed cell viability by MTT assay (A) and IL-1β levels in the supernatant by ELISA (B). Summarized data showed cell viability of RAECs treated with acarbose at the concentration of 1, 3, 9µM (C), and IL-1β concentration secreted from RAECs treated with 3, 9µM acarbose under 30mM glucose stimulation (D). RAECs were incubated with 30mM glucose for 24 hours and pretreated with 3µM acarbose, the protein expression of NLRP3 (E) and p20/pro-caspase-1 (F), the activity of caspase-1 (G), and IL-1β concentration (H).

5). Zhu JJ et al. Neferine Protects against Hypoxic-Ischemic Brain Damage in Neonatal Rats by Suppressing NLRP3-Mediated Inflammasome Activation. Oxid Med Cell Longev 2021 May 8;2021:6654954. (PubMed: 34046147) [IF=7.310]

Application: WB    Species: Rat    Sample: brain tissue

Figure 4 Neferine attenuates NLRP3/caspase-1/IL-1β activation in neonatal rat brain tissue after HI. (a) The protein levels of cleaved caspase-1, GSDMD, ASC, NLRP3, IL-1β, and IL-18 were evaluated by Western blotting in brain tissues. (b–g) Analyses of cleaved caspase-1, GSDMD, ASC, NLRP3, IL-1β, and IL-18 (normalized to β-actin). ∗P < 0.05, ∗∗P < 0.01, and ∗∗∗P < 0.001. n = 4.

6). Hong Z et al. The ROS/GRK2/HIF-1α/NLRP3 Pathway Mediates Pyroptosis of Fibroblast-Like Synoviocytes and the Regulation of Monomer Derivatives of Paeoniflorin. Oxid Med Cell Longev 2022 Jan 29;2022:4566851. (PubMed: 35132350) [IF=7.310]

7). Zhao X et al. Quercetin Protects Ethanol-Induced Hepatocyte Pyroptosis via Scavenging Mitochondrial ROS and Promoting PGC-1α-Regulated Mitochondrial Homeostasis in L02 Cells. Oxid Med Cell Longev 2022 Jul 16;2022:4591134. (PubMed: 35879991) [IF=7.310]

8). Ding S et al. Resveratrol alleviates chronic "real-world" ambient particulate matter-induced lung inflammation and fibrosis by inhibiting NLRP3 inflammasome activation in mice. Ecotoxicol Environ Saf 2019 Oct 30;182:109425 (PubMed: 31295660) [IF=7.129]

9). Lin JQ et al. Zinc provides neuroprotection by regulating NLRP3 inflammasome through autophagy and ubiquitination in a spinal contusion injury model. CNS Neurosci Ther 2020 Oct 9. (PubMed: 33034415) [IF=7.035]

Application: WB    Species: mouse    Sample:

FIGURE 4 |Treatment with zinc suppresses the NLRP3 inflammasome after SCI Mice were sacrificed on the third day after zinc administration.(A)Representative Western blot analysis of NLRP3 inflammasome in Sham, SCI + vehicle and SCI + zinc group(n = 6).

Application: IF/ICC    Species: mouse    Sample:

FIGURE 4 |Treatment with zinc suppresses the NLRP3 inflammasome after SCI Mice were sacrificed on the third day after zinc administration.(A)Representative Western blot analysis of NLRP3 inflammasome in Sham, SCI + vehicle and SCI + zinc group(n = 6). (B-E) The result of WB analysis of NLRP3, ASC, Caspase-1, IL-1β from each group. (F) Immunofluorescence staining was used to detect the level of NLRP3 from each group (n = 8, scale bar = 50 µm).

10). Du L et al. Novel biphenyl diester derivative AB-38b inhibits NLRP3 inflammasome through Nrf2 activation in diabetic nephropathy. Cell Biol Toxicol 2019 Nov 25 (PubMed: 31768838) [IF=6.819]

Application: IHC    Species: mouse    Sample: kidney

Fig. 4 Immunohistochemical staining for NLRP3, cleaved caspase-1, and IL-18 of diabetic mice kidneys. Representative views for NLRP3 (a), cleaved caspase-1 (c), and IL-18 e) were shown.

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