|Coordinate||91,499,537 bp (GRCm38)|
|Base Change||C ⇒ T (forward strand)|
|Gene Name||interferon (alpha and beta) receptor 1|
|Synonym(s)||Ifar, Ifrc, IFN-alpha/betaR|
|Chromosomal Location||91,485,238-91,507,441 bp (+)|
|MGI Phenotype||Homozygotes for targeted null mutations exhibit increased susceptibility to viral infection, elevated levels of myeloid lineage cells in the peripheral blood and bone marrow, and reduced immune response to immunostimulatory DNA.|
|Amino Acid Change||Glutamine changed to Stop codon|
|Institutional Source||Beutler Lab|
|Gene Model||predicted gene model for protein(s): [ENSMUSP00000023689]|
AA Change: Q309*
|Predicted Effect||probably null|
|Phenotypic Category||impaired response to dsDNA- type I IFN production by macrophages|
|Alleles Listed at MGI|
|Mode of Inheritance||Autosomal Recessive|
|Local Stock||Live Mice|
|Last Updated||05/13/2016 3:09 PM by Anne Murray|
|Record Created||02/08/2013 12:08 AM by Hexin Shi|
Shook was initially identified among N-ethyl-N-nitrosourea (ENU)-induced G3 animals in the Double-stranded DNA Macrophage Screen. Shook peritoneal macrophages were defective in producing type I interferon (IFN) in response to dsDNA (Figure 1, red dots).
|Nature of Mutation|
Whole exome HiSeq sequencing of the G1 grandsire identified 59 mutations. Four G3 mice with the shook phenotype were genotyped at all 59 mutation sites and two mutations on chromosome 16 were homozygous in all four of the shook mice. Subsequent analysis of five additional affected mice and 21 unaffected mice supported a causal relationship between the mutation and the phenotype (LOD =5.49). Sequencing of the mutated gene identified a C to T transition at base pair 91499537 (v38) on Chromosome 16 in the GenBank genomic region NC_000082 encoding Ifnar1. The mutation corresponds to residue 1054 in the mRNA sequence (NM_010508.2) within exon 7 of 11 total exons.
The mutated nucleotide is indicated in red. The mutation results in a glutamine (Q) to premature stop codon (*) substitution at residue 309.
The shook mutation (Q309*) is within the fibronectin type-III subdomain 3 (SD3) of the IFNAR1 protein (Figure 2). It is unknown whether this protein is expressed and localized normally.
Please see the record macro-1 for information about Ifnar1.
Shook mice produce significantly reduced amounts of type I IFN in response to dsDNA stimulation of macrophages ex vivo. This finding suggests that the shook mutation impairs signaling through the IFNAR receptor, thus preventing activation of the positive feedback loop for type I IFN production.
shook(F):5'- GGTCCACAGTGCTTACTACAGTGTC -3'
shook(R):5'- CGCCCTACATGAGGAATTACACTCC -3'
shook_seq(F):5'- tgtcttaggtattatagagcctcag -3'
shook_seq(R):5'- gaccacacccacaacctc -3'
Shook 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. The same primers are used for PCR amplification and for sequencing.
Shook(F): 5’- GGTCCACAGTGCTTACTACAGTGTC-3’
Shook(R): 5’- CGCCCTACATGAGGAATTACACTCC-3’
Shook_seq(F): 5’- TGTCTTAGGTATTATAGAGCCTCAG-3’
1) 94°C 2:00
2) 94°C 0:30
3) 55°C 0:30
4) 72°C 1:00
5) repeat steps (2-4) 40X
6) 72°C 10:00
7) 4°C ∞
The following sequence of 496 nucleotides (from Genbank genomic region NC_000082 for linear DNA sequence of Ifnar1) is amplified:
14085 ggtcca cagtgcttac
14101 tacagtgtct taggtattat agagcctcag tacatagtga attaattcat gattggattg
14161 gattttgttt ttgctttctg ttttcaaagt ggctattcaa aaagcagttc tggaagccgt
14221 tcagataaat ggaaaccaat accaacctgt gcaaatgtcc agactacgca ctgtgtcttt
14281 tctcaagata ctgtctacac aggaacgttc tttctccatg tacaagcctc agagggaaat
14341 cacacatcct tttggtctga agagaagttt attgattctc aaaaacacag taagccgagt
14401 tttctttgag acagtctgac actgtagccc aggctggcct ggaactcacg gtgtagccca
14461 gggtagcttc aaacttatgg cagtcctcct gcctgagctc ctgagagctg aggttgtggg
14521 tgtggtccat gcctgctgta tagcaagcgc tttctggagt gtaattcctc atgtagggcg
Primer binding sites are underlined and the sequencing primer is highlighted; the mutated C is shown in red text.
|Science Writers||Anne Murray|
|Authors||Hexin Shi, Ying Wang, Bruce Beutler|