Phenotypic Mutation 'Nd6' (pdf version)
AlleleNd6
Mutation Type missense
Chromosome11
Coordinate59,549,354 bp (GRCm38)
Base Change A ⇒ G (forward strand)
Gene Nlrp3
Gene Name NLR family, pyrin domain containing 3
Synonym(s) Cias1, cryopyrin, Pypaf1, NALP3, Mmig1
Chromosomal Location 59,541,568-59,566,955 bp (+)
MGI Phenotype Strain: 3686871
Mice homozygous for null mutations exhibit attenuated inflammatory responses related to decrease secretion of IL-1beta and IL-18. Mice heterozygous for activating mutations suffer from autoinflammatory attacks that lead to organ failure and death before weaning.
Accession Number

NCBI RefSeq: NM_145827; MGI: 2653833

Mapped Yes 
Amino Acid Change Arginine changed to Glycine
Institutional SourceBeutler Lab
Ref Sequences
R586G in Ensembl: ENSMUSP00000078440 (fasta)
Gene Model not available
SMART Domains

DomainStartEndE-ValueType
Pfam:PAAD_DAPIN 4 87 8.6e-24 PFAM
Pfam:NACHT 216 385 1.3e-46 PFAM
low complexity region 533 539 N/A INTRINSIC
low complexity region 688 697 N/A INTRINSIC
LRR_RI 737 764 1.07e-9 SMART
LRR 766 793 5.13e1 SMART
LRR 794 821 3.86e-7 SMART
LRR 823 850 1.62e0 SMART
LRR 851 878 3.39e-3 SMART
LRR 880 907 1.2e2 SMART
LRR 908 935 2.24e-3 SMART
LRR 937 964 2.16e2 SMART
LRR 965 992 8.73e-6 SMART
Predicted Effect possibly damaging

PolyPhen 2 Score 0.586 (Sensitivity: 0.88; Specificity: 0.91)
(Using Ensembl: ENSMUSP00000078440)
Phenotypic Category immune system, NALP3 inflammasome signaling defect
Penetrance  
Alleles Listed at MGI

All alleles(10) : Targeted, knock-out(3) Targeted, other(4) Chemically induced(3)

Lab Alleles
AlleleSourceChrCoordTypePredicted EffectPPH Score
IGL00421:Nlrp3 APN 11 59565943 missense probably damaging 0.99
IGL00573:Nlrp3 APN 11 59565116 missense possibly damaging 0.93
IGL01025:Nlrp3 APN 11 59551887 missense probably benign 0.21
IGL01637:Nlrp3 APN 11 59549378 missense probably damaging 0.99
IGL02010:Nlrp3 APN 11 59549535 missense probably benign
IGL02334:Nlrp3 APN 11 59565083 missense probably benign
IGL02417:Nlrp3 APN 11 59566023 unclassified 0.00
IGL02578:Nlrp3 APN 11 59548401 missense probably damaging 1.00
IGL02710:Nlrp3 APN 11 59565976 missense probably damaging 0.99
IGL02816:Nlrp3 APN 11 59555782 missense probably benign 0.03
IGL03157:Nlrp3 APN 11 59549546 missense possibly damaging 0.80
IGL03334:Nlrp3 APN 11 59549016 missense probably damaging 1.00
Nd1 UTSW 11 59565974 missense probably benign 0.33
Nd14 UTSW 11 59555875 missense
Nd3 UTSW 11 59565974 missense probably benign 0.33
Nd5 UTSW 11 59565879 missense probably benign 0.00
Nd7 UTSW 11 59555875 missense probably damaging 0.99
Nd9 UTSW 11 59549354 missense possibly damaging 0.59
Park2 UTSW 11 59565128 nonsense probably null
Park3 UTSW 11 59565850 missense probably benign 0.02
Park4 UTSW 11 59549531 missense probably benign 0.19
Park5 UTSW 11 59548476 missense probably damaging 0.99
Park6 UTSW 11 59549036 missense probably damaging 1.00
Park7 UTSW 11 59548010 nonsense probably null
Park8 UTSW 11 59566199 missense probably benign 0.19
R0008:Nlrp3 UTSW 11 59558448 missense probably benign 0.00
R0008:Nlrp3 UTSW 11 59558448 missense probably benign 0.00
R0052:Nlrp3 UTSW 11 59565128 nonsense probably null
R0362:Nlrp3 UTSW 11 59548797 missense possibly damaging 0.49
R0416:Nlrp3 UTSW 11 59555924 splice donor site probably benign
R0649:Nlrp3 UTSW 11 59548542 missense possibly damaging 0.83
R0740:Nlrp3 UTSW 11 59548256 missense probably benign 0.01
R0863:Nlrp3 UTSW 11 59565850 missense probably benign 0.02
R1300:Nlrp3 UTSW 11 59555768 missense possibly damaging 0.86
R1622:Nlrp3 UTSW 11 59548476 missense probably damaging 0.99
R1654:Nlrp3 UTSW 11 59543123 missense probably benign 0.03
R1715:Nlrp3 UTSW 11 59543351 missense probably damaging 1.00
R1754:Nlrp3 UTSW 11 59558402 missense possibly damaging 0.80
R1837:Nlrp3 UTSW 11 59548916 missense probably benign 0.00
R1905:Nlrp3 UTSW 11 59549036 missense probably damaging 1.00
R2281:Nlrp3 UTSW 11 59549136 missense possibly damaging 0.70
R4296:Nlrp3 UTSW 11 59549661 missense possibly damaging 0.89
R4305:Nlrp3 UTSW 11 59548010 nonsense probably null
R4540:Nlrp3 UTSW 11 59551899 missense possibly damaging 0.83
R4591:Nlrp3 UTSW 11 59549222 missense probably benign 0.00
R4816:Nlrp3 UTSW 11 59548301 missense probably benign 0.32
R4913:Nlrp3 UTSW 11 59549238 missense probably benign 0.09
R4970:Nlrp3 UTSW 11 59548728 missense probably damaging 1.00
R5051:Nlrp3 UTSW 11 59566199 missense probably benign 0.19
R5112:Nlrp3 UTSW 11 59548728 missense probably damaging 1.00
R5185:Nlrp3 UTSW 11 59565084 missense probably benign 0.05
R5417:Nlrp3 UTSW 11 59549063 missense probably damaging 1.00
R5869:Nlrp3 UTSW 11 59548134 missense probably damaging 1.00
R5898:Nlrp3 UTSW 11 59546852 missense probably benign 0.00
R5953:Nlrp3 UTSW 11 59546791 missense probably benign
R5979:Nlrp3 UTSW 11 59548971 missense probably benign 0.06
Z1088:Nlrp3 UTSW 11 59551860 missense possibly damaging 0.67
Mode of Inheritance Autosomal Semidominant
Local Stock Sperm, gDNA
Repository

none

Last Updated 03/02/2017 12:13 PM by Katherine Timer
Record Created 06/05/2010 8:25 PM by Hua Huang
Record Posted 10/18/2010
Phenotypic Description
Figure 1.

The ND6 phenotype was initially identified among G3 mice (G5801 and G5806) homozygous for mutations induced by N-ethyl-N-nitrosourea (ENU) and tested in the NALP3 Inflammasome Screen.  Peritoneal macrophages isolated from these mice secreted reduced amounts of the proinflammatory cytokine interleukin (IL)-1β in response to priming with lipopolysaccharide (LPS) followed by nigericin treatment (Figure 1).  Macrophages from G5801 produced normal levels of tumor necrosis factor (TNF)-α in response to LPS stimulation alone, suggesting that signaling from the Toll-like receptor 4 (TLR4), which senses LPS, was unimpaired.

 

 

 

Nature of Mutation
The Nlrp3 gene was directly sequenced as a candidate gene and an A to G transition was found at position 1982 of the Nlrp3 transcript in exon 4 of 10 total exons using Genbank record NM_145827. The mutation is located in the third coding exon.
 
1967 CTTGTAAACCAGGAGAGAACCTCTTATTTGGAG
581 -L--V--N--Q--E--R--T--S--Y--L--E-
 
The mutated nucleotide is indicated in red lettering, and causes an arginine to glycine substitution at residue 586 of the NLRP3 protein.
Protein Prediction

Figure 2. Domain structure of NLRP3. Shown are the pyrin domain (PYD), NACHT domain, NACHT associated domain (NAD), and leucine-rich repeat (LRR) domain. Together, the NACHT and NAD domains are known as the nucleotide-binding domain (NBD). The amino acid altered by the ND7 mutation is shown. Click on the image to view other mutations found in NLRP3. Click on each mututation for more specific information.

The ND6 mutation results in an arginine to glycine change in coding exon 3, and occurs in the region between the nucleotide-binding domain (NBD) and the first leucine rich repeat (LRR). 
 
Please see the record for ND1 for more information about Nlrp3.
Putative Mechanism
NLRP3 is able to oligomerize through its NBD domain and assemble into large caspase-1-activating multiprotein complexes termed inflammasomes upon the detection of pathogenic or other danger signals in the cytoplasm. A large variety of agents have been shown to activate the NLRP3 inflammasome, and NLRP3 plays an important role in the innate immune response.
 
A total of 93 disease-associated mutations have been found in humans in the NLRP3 gene according to the Infevers database, an online database for autoinflammatory mutations (1-3). Rather than causing immunodeficiencies, these are activating mutations that cause autoinflammatory disorders including Muckle-Wells syndrome (MWS; #191900), familial cold autoinflammatory syndrome (FCAS1; OMIM #120100), and chronic infantile neurologic cutaneous and articular syndrome (CINCA; #607115), also known as NOMID for neonatal onset multisystem inflammatory disease (4-6). Collectively, these diseases are known as cryopyrin associated periodic syndrome (CAPS). Interestingly, the vast majority of these activating missense mutations are located in exon 3, which encodes the NBD and surrounding sequences. These mutations are thought to either allow the protein the ability to permanently bind to and hydrolyze ATP or unable to bind to the inhibitory LRRs that prevent self-oligomerization (7-8).  However, the ND6 mutation, which is located in the same exon, appears to be an inactivating mutation and renders macrophages unresponsive to NLRP3-activating stimuli. The mechanism by which the ND6 mutation affects NLRP3 function is unknown. 
Primers Primers cannot be located by automatic search.
Genotyping
ND6 genotyping is performed by amplifying the region containing the mutation using PCR, followed by sequencing of the amplified region to detect the single nucleotide transition. 
 
Primers
ND6(F): 5’- AGGCTGATCCAAGAGAATGAGGTCC -3’
ND6(R): 5’- AACACACTGAACCTGGTCCAAGGG -3’
 
PCR program
1) 95°C             2:00
2) 95°C             0:30
3) 56°C             0:30
4) 72°C             1:00
5) repeat steps (2-4) 29X
6) 72°C             7:00
7) 4°C               8
 
Primers for sequencing
ND6 _seq(F): 5'- ATCTTCTGAACCGAGACGTG -3'
ND6 _seq(R): 5’- AGCGTTTTGACCCTATGACAG -3’
 
The following sequence of 939 nucleotides (NCBI Mouse Genome Build 37.1, Chromosome 11, bases 59,362,277 to 59,363,215) is amplified:
 
aggctgatcc aagagaatga ggtcctcttt accatgtgct tcatccccct ggtctgctgg
attgtgtgca cggggctaaa gcaacagatg gagaccggga agagcctggc ccagacctcc
aagaccacta cggccgtcta cgtcttcttc ctttccagcc tgctgcaatc ccgggggggc
attgaggagc atctcttctc tgactaccta caggggctct gttcactggc tgcggatgga
atttggaacc agaaaatcct atttgaggag tgtgatctgc ggaagcacgg cctgcagaag
actgacgtct ccgctttcct gaggatgaac gtgttccaga aggaagtgga ctgcgagaga
ttctacagct tcagccacat gactttccag gagttcttcg ctgctatgta ctatttgctg
gaagaggagg cagaggggga gaccgtgagg aaaggaccag gaggttgttc agatcttctg
aaccgagacg tgaaggtcct actagaaaat tacggcaagt ttgaaaaagg ctatctgatt
tttgttgtcc gattcctctt tggccttgta aaccaggaga gaacctctta tttggagaag
aaactaagtt gcaagatctc tcagcaagtc agactggaac tactgaagtg gattgaagtg
aaagccaagg ccaagaagct gcagtggcag cccagccaac tggaactgtt ctactgcctg
tacgagatgc aggaggaaga ctttgtgcag agtgccatgg accactttcc caaaattgag
atcaacctct ctaccagaat ggaccacgtg gtttcctcct tttgtattaa gaactgtcat
agggtcaaaa cgctttccct gggttttttt cacaactcgc ccaaggagga agaagaagag
aggagaggag gtcga
ccctt ggaccaggtt cagtgtgtt
        
Primer binding sites are underlined; sequencing primer binding sites are highlighted in gray; the mutated A is indicated in red.
References
Science Writers Nora G. Smart
Illustrators Katherine Timer
AuthorsHua Huang, Bruce Beutler
Edit History
07/20/2011 12:27 PM (current)
05/25/2011 10:49 AM
05/24/2011 11:19 AM
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10/19/2010 11:35 AM
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10/18/2010 3:24 PM
07/02/2010 8:14 AM
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07/02/2010 8:13 AM