Phenotypic Mutation 'nibbler' (pdf version)
Allele | nibbler |
Mutation Type |
critical splice donor site
|
Chromosome | 10 |
Coordinate | 77,397,050 bp (GRCm39) |
Base Change | G ⇒ A (forward strand) |
Gene |
Itgb2
|
Gene Name | integrin beta 2 |
Synonym(s) | Mac-1 beta, 2E6, Cd18 |
Chromosomal Location |
77,366,164-77,401,542 bp (+) (GRCm39)
|
MGI Phenotype |
FUNCTION: [Summary is not available for the mouse gene. This summary is for the human ortholog.] This gene encodes an integrin beta chain, which combines with multiple different alpha chains to form different integrin heterodimers. Integrins are integral cell-surface proteins that participate in cell adhesion as well as cell-surface mediated signalling. The encoded protein plays an important role in immune response and defects in this gene cause leukocyte adhesion deficiency. Alternative splicing results in multiple transcript variants. [provided by RefSeq, Dec 2014] PHENOTYPE: Homozygotes for targeted null and hypomorphic mutations are subject to granulocytosis, impaired inflammatory and immune responses, and chronic dermatitis. [provided by MGI curators]
|
Accession Number | NCBI RefSeq: NM_008404; MGI: 96611
|
Mapped | Yes |
Limits of the Critical Region |
77530252 - 77565708 bp |
Amino Acid Change |
|
Institutional Source | Beutler Lab |
Gene Model |
predicted gene model for protein(s):
[ENSMUSP00000000299 †]
[ENSMUSP00000137734 †]
[ENSMUSP00000137865 †]
† probably from a misspliced transcript
|
AlphaFold |
no structure available at present |
SMART Domains |
Protein: ENSMUSP00000000299 Gene: ENSMUSG00000000290
Domain | Start | End | E-Value | Type |
signal peptide
|
1 |
22 |
N/A |
INTRINSIC |
PSI
|
24 |
74 |
6.91e-7 |
SMART |
INB
|
32 |
447 |
1.98e-268 |
SMART |
VWA
|
126 |
357 |
1.25e-1 |
SMART |
internal_repeat_1
|
459 |
509 |
7.99e-5 |
PROSPERO |
EGF_like
|
535 |
574 |
6.81e1 |
SMART |
Integrin_B_tail
|
622 |
701 |
5.53e-22 |
SMART |
transmembrane domain
|
702 |
724 |
N/A |
INTRINSIC |
Integrin_b_cyt
|
725 |
770 |
1.58e-17 |
SMART |
|
Predicted Effect |
probably null
|
SMART Domains |
Protein: ENSMUSP00000137734 Gene: ENSMUSG00000000290
Domain | Start | End | E-Value | Type |
signal peptide
|
1 |
22 |
N/A |
INTRINSIC |
PSI
|
24 |
74 |
6.91e-7 |
SMART |
INB
|
32 |
447 |
1.98e-268 |
SMART |
VWA
|
126 |
357 |
1.25e-1 |
SMART |
|
Predicted Effect |
probably benign
|
SMART Domains |
Protein: ENSMUSP00000137865 Gene: ENSMUSG00000000290
Domain | Start | End | E-Value | Type |
signal peptide
|
1 |
22 |
N/A |
INTRINSIC |
PDB:2P28|A
|
23 |
49 |
9e-12 |
PDB |
Blast:PSI
|
24 |
49 |
2e-11 |
BLAST |
|
Predicted Effect |
probably benign
|
Meta Mutation Damage Score |
0.9576 |
Is this an essential gene? |
Possibly nonessential (E-score: 0.477) |
Phenotypic Category |
Autosomal Recessive |
Candidate Explorer Status |
loading ... |
Single pedigree Linkage Analysis Data
|
|
Penetrance | |
Alleles Listed at MGI | All Mutations and Alleles(11) : Chemically induced (ENU)(1) Chemically induced (other)(1) Targeted(9)
|
Lab Alleles |
Allele | Source | Chr | Coord | Type | Predicted Effect | PPH Score |
IGL00332:Itgb2
|
APN |
10 |
77393240 |
missense |
probably damaging |
1.00 |
IGL00427:Itgb2
|
APN |
10 |
77393790 |
missense |
probably benign |
0.13 |
IGL00500:Itgb2
|
APN |
10 |
77400558 |
missense |
probably damaging |
1.00 |
IGL01019:Itgb2
|
APN |
10 |
77378237 |
missense |
possibly damaging |
0.94 |
IGL01104:Itgb2
|
APN |
10 |
77383028 |
splice site |
probably null |
|
IGL01111:Itgb2
|
APN |
10 |
77377834 |
missense |
probably damaging |
0.98 |
IGL01574:Itgb2
|
APN |
10 |
77393798 |
missense |
possibly damaging |
0.82 |
IGL02087:Itgb2
|
APN |
10 |
77395530 |
missense |
possibly damaging |
0.94 |
IGL02132:Itgb2
|
APN |
10 |
77385895 |
missense |
probably damaging |
1.00 |
IGL02325:Itgb2
|
APN |
10 |
77383026 |
missense |
probably damaging |
1.00 |
IGL02505:Itgb2
|
APN |
10 |
77383052 |
missense |
probably damaging |
1.00 |
IGL02590:Itgb2
|
APN |
10 |
77395347 |
missense |
probably damaging |
1.00 |
IGL02735:Itgb2
|
APN |
10 |
77385833 |
missense |
possibly damaging |
0.81 |
almondine
|
UTSW |
10 |
77384503 |
missense |
probably damaging |
1.00 |
barely
|
UTSW |
10 |
77384370 |
splice site |
probably benign |
|
fresh
|
UTSW |
10 |
77391995 |
missense |
probably damaging |
0.98 |
joker
|
UTSW |
10 |
77549849 |
intron |
probably benign |
|
newhome
|
UTSW |
10 |
77395515 |
missense |
probably benign |
0.00 |
Only_just
|
UTSW |
10 |
77385802 |
missense |
possibly damaging |
0.80 |
salmonid
|
UTSW |
10 |
77396946 |
missense |
probably benign |
|
trout
|
UTSW |
10 |
77401022 |
missense |
probably damaging |
1.00 |
R0217:Itgb2
|
UTSW |
10 |
77384370 |
splice site |
probably benign |
|
R0394:Itgb2
|
UTSW |
10 |
77378309 |
missense |
probably damaging |
1.00 |
R0396:Itgb2
|
UTSW |
10 |
77397023 |
missense |
probably damaging |
0.97 |
R1425:Itgb2
|
UTSW |
10 |
77383130 |
missense |
probably null |
1.00 |
R1499:Itgb2
|
UTSW |
10 |
77381987 |
missense |
possibly damaging |
0.62 |
R1542:Itgb2
|
UTSW |
10 |
77395320 |
missense |
probably benign |
|
R1803:Itgb2
|
UTSW |
10 |
77400624 |
missense |
probably benign |
0.15 |
R1889:Itgb2
|
UTSW |
10 |
77384457 |
missense |
possibly damaging |
0.74 |
R2035:Itgb2
|
UTSW |
10 |
77383033 |
missense |
probably damaging |
1.00 |
R2156:Itgb2
|
UTSW |
10 |
77396082 |
missense |
probably benign |
0.01 |
R2374:Itgb2
|
UTSW |
10 |
77395515 |
missense |
probably benign |
0.00 |
R3769:Itgb2
|
UTSW |
10 |
77385802 |
missense |
possibly damaging |
0.80 |
R3942:Itgb2
|
UTSW |
10 |
77393867 |
missense |
probably benign |
0.31 |
R4352:Itgb2
|
UTSW |
10 |
77392001 |
missense |
probably benign |
0.10 |
R4537:Itgb2
|
UTSW |
10 |
77397050 |
critical splice donor site |
probably null |
|
R4600:Itgb2
|
UTSW |
10 |
77381949 |
missense |
probably benign |
|
R4611:Itgb2
|
UTSW |
10 |
77385884 |
missense |
probably damaging |
1.00 |
R4685:Itgb2
|
UTSW |
10 |
77385937 |
critical splice donor site |
probably null |
|
R4717:Itgb2
|
UTSW |
10 |
77381878 |
nonsense |
probably null |
|
R5068:Itgb2
|
UTSW |
10 |
77384595 |
missense |
probably damaging |
1.00 |
R5297:Itgb2
|
UTSW |
10 |
77400501 |
missense |
probably damaging |
1.00 |
R5355:Itgb2
|
UTSW |
10 |
77393886 |
missense |
probably benign |
|
R5927:Itgb2
|
UTSW |
10 |
77381868 |
missense |
probably damaging |
1.00 |
R6371:Itgb2
|
UTSW |
10 |
77384431 |
missense |
probably damaging |
1.00 |
R6505:Itgb2
|
UTSW |
10 |
77395507 |
missense |
probably damaging |
1.00 |
R7305:Itgb2
|
UTSW |
10 |
77384398 |
missense |
probably damaging |
1.00 |
R7574:Itgb2
|
UTSW |
10 |
77395992 |
missense |
probably benign |
0.18 |
R7606:Itgb2
|
UTSW |
10 |
77391995 |
missense |
probably damaging |
0.98 |
R7772:Itgb2
|
UTSW |
10 |
77396946 |
missense |
probably benign |
|
R7888:Itgb2
|
UTSW |
10 |
77400478 |
missense |
probably benign |
0.00 |
R8716:Itgb2
|
UTSW |
10 |
77393787 |
missense |
probably damaging |
0.99 |
R8933:Itgb2
|
UTSW |
10 |
77401022 |
missense |
probably damaging |
1.00 |
R9082:Itgb2
|
UTSW |
10 |
77384503 |
missense |
probably damaging |
1.00 |
R9479:Itgb2
|
UTSW |
10 |
77396942 |
missense |
probably benign |
0.01 |
Z1176:Itgb2
|
UTSW |
10 |
77393796 |
missense |
probably benign |
0.01 |
|
Mode of Inheritance |
Autosomal Recessive |
Local Stock | |
Repository | |
Last Updated |
2019-09-04 9:42 PM
by Diantha La Vine
|
Record Created |
2016-09-02 3:07 PM
by Evan Nair-Gill
|
Record Posted |
2018-06-13 |
Phenotypic Description |
The nibbler phenotype was identified among G3 mice of the pedigree R4537, some of which showed reduced cytotoxic T lymphocyte killing (Figure 1), and reduced natural killer cell killing (Figure 2). The T-dependent antibody response to recombinant Semliki Forest virus (rSFV)-encoded β-galactosidase (rSFV-β-gal) was diminished (Figure 3). Some mice showed reduced B to T cell ratios (Figure 4) and increased CD4 to CD8 T cell ratios (Figure 5). The mice showed reduced frequencies of B cells (Figure 6), IgD+ B cells (Figure 7), IgM+ B cells (Figure 8), CD4+ T cells (Figure 9), CD4+ T cells in CD3+ T cells (Figure 10), CD8+ T cells (Figure 11), CD8+ T cells in CD3+ T cells (Figure 12), naïve CD4 T cells in CD4+ T cells (Figure 13), naïve CD8 T cells in CD8+ T cells (Figure 14), macrophages (Figure 15), neutrophils (Figure 16), and CD11b+ dendritic cells (gated in CD11c+ cells) (Figure 17) with concomitant increased frequencies of CD8a+ dendritic cells (gated in CD11c+ cells) (Figure 18), NK T cells (Figure 19), effector memory CD4 T cells in CD4+ T cells (Figure 20), and effector memory CD8 T cells in CD8+ T cells (Figure 21), all in the peripheral blood. Some mice showed increased expression of CD44 on peripheral blood T cells (Figure 22) and CD4+ T cells (Figure 23). Some mice also showed increased levels of total IgE in the serum (Figure 24).
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Nature of Mutation |
Whole exome HiSeq sequencing of the G1 grandsire identified 43 mutations. All of the above anomalies were linked by continuous variable mapping to a mutation in Itgb2: a G to A transition at base pair 77,561,216 (v38) on chromosome 10, or base pair 30,888 in the GenBank genomic region NC_000076 within the donor splice site of intron 14. The strongest association was found with a recessive model of inheritance to the reduced frequency of CD8+ T cells in CD3+ T cells, wherein two variant homozygotes departed phenotypically from 11 homozygous reference mice and 14 heterozygous mice with a P value of 5.67 x 10-17 (Figure 25). The effect of the mutation at the cDNA and protein level have not examined, but the mutation is predicted to result in the use of a cryptic site in intron 14, resulting in an 80-base pair insertion of intron 14. This predicts a frame shifted protein product beginning after amino acid 694 of the protein, which is normally 770 amino acids in length, and terminating after the inclusion of 9 aberrant amino acids.
C57BL/6J:
<--exon 13 <--exon 14 intron 14--> <--exon 15-->
30034 ……GACAACCACAC ……GAGGACAGTCTAG gtgaggctgggtggt…… AGTGTGTGAAG……GCTGAAAGCTAG 34912
624 ……-D--N--H--T ……-E--D--S--L-- E--C--V--K-……-A--E--S--*- 770
|
Genomic numbering corresponds to NC_000076. The donor splice site of intron 14, which is destroyed by the nibbler mutation, is indicated in blue lettering and the mutated nucleotide is indicated in red.
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Illustration of Mutations in
Gene & Protein |
|
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Protein Prediction |
The Itgb2 gene encodes the integrin β2 protein (also called CD18), which forms noncovalently linked dimers with integrin α subunits to form functional integrin receptors. The extracellular domains of integrin α and β subunits are >940 and >640 residues, respectively, but the intracellular domains are much shorter, about 50 residues. The shape of the integrin receptor extracellular domain, as determined by electron microscopy, is a globular ligand-binding headpiece connected to two long stalk regions, connected to the transmembrane and C-terminal cytoplasmic domains. Integrin β2 is a cysteine-rich single pass transmembrane protein with six N-linked extracellular glycosylation sites (Figure 26) [reviewed in (1)]. The N-terminal cysteine-rich region (residues ~1-50) of the integrin β2 extracellular domain is called a PSI (plexins, semaphorins, integrins) domain, and shares homology with the membrane proteins plexins, semaphorins and the c-met receptor. Residues ~100-340 contain a von Willebrand factor-type A domain of 241 amino acids, which is referred to as the inserted (I) domain in α subunits and I-like domain in β subunits. The C-terminal portion of the extracellular domain is the “stalk” region, which contains four EGF-like domains (integrin-EGF, I-EGF) and a tail domain connecting it to the membrane. The nibbler mutation is predicted to result in the use of a cryptic site in intron 14, resulting in an 80-base pair insertion of intron 14. For more information about Itgb2, please see the record for Joker.
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Putative Mechanism | Integrins are adhesion molecules that mediate cell-cell, cell-extracellular matrix, and cell-pathogen interactions. They regulate cell migration and morphogenesis by coordinating regulatory signals from inside and outside the cell, with the physical machinery for cell movement. Most integrins, including β2-integrins, link to and regulate the actin cytoskeleton. The β2-containing integrins are αLβ2 (CD11aCD18; also called leukocyte function-associated antigen 1, LFA-1), αMβ2 (CD11bCD18; also called MAC-1), αXβ2 (CD11cCD18; also called p150,95) and αDβ2 (CD11dCD18). The CD11/CD18 integrins are referenced collectively as the “leukocyte” integrins, and mediate leukocyte adhesion during inflammatory responses to infections and also during wound repair. Two integrin β2 mutant mice have been developed. One strain contains a hypomorphic allele (Itgb2tm1Bay) of integrin β2 (2). These animals exhibit elevated total numbers of leukocytes, including granulocytes and lymphocytes, as well as an impaired inflammatory response to intraperitoneal injection of thioglycolate medium (slightly fewer numbers of neutrophils migrate into the peritoneal cavity). The second integrin β2 mutant strain contains a null allele (Itgb2tm2Bay) (3), and exhibits a phenotype very similar to that of human patients with leukocyte adhesion deficiency (LAD, OMIM #116920), an autosomal recessive disorder characterized by leukocytosis (especially neutrophilia), failure to recruit leukocytes to sites of infection, recurring bacterial and fungal infections involving the skin and mucosa, impaired wound healing, and lack of pus formation. Nibbler mice display a phenotype distinct from that of the CD18 null or hypomorphic mutant, and from humans with LAD. In particular, nibbler mice do not develop the dermatitis or facial/submandibular inflammation observed in CD18-/- mice. Spontaneous infections have not been observed in nibbler mice, as they are in the CD18 null mutant and in LAD patients.
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Primers |
PCR Primer
nibbler_pcr_F: AGGGCCCTTTTGAGAAGAAC
nibbler_pcr_R: TTCATCTCCTCATACAGGGCCG
Sequencing Primer
nibbler_seq_F: GGCCCTTTTGAGAAGAACTGTAG
nibbler_seq_R: CTCATACAGGGCCGGGTAAG
|
Genotyping | PCR program 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 hold
The following sequence of 431 nucleotides is amplified (chromosome 10, + strand):
1 agggcccttt tgagaagaac tgtagtgttc agtgtgctgg tatgacgctg cagactatcc 61 ctttgaagaa aaagccctgc aaggagaggg actcggaagg ctgttggata acttacactt 121 tgcagcagaa ggacggaagg aacatttaca acatccatgt ggaggacagt ctaggtgagg 181 ctgggtggta cctgggggtg tgatctcatg atctggtctc agtggagcat agttctcagt 241 ggagtactct gaccgtgagg cctttgcaga cagtgagggg gctgtagagg aaaacatctc 301 tcttcagtgg ggcaaagcag gaggtggagc catccgggac acttgggcag ggcagctagc 361 agccatccac aggctctgcc ctgtggccct ctcattccca ctccttaccc ggccctgtat 421 gaggagatga a
Primer binding sites are underlined and the sequencing primers are highlighted; the mutated nucleotide is shown in red. |
References |
2. Wilson, R. W., Ballantyne, C. M., Smith, C. W., Montgomery, C., Bradley, A., O'Brien, W. E., and Beaudet, A. L. (1993) Gene Targeting Yields a CD18-Mutant Mouse for Study of Inflammation. J Immunol. 151, 1571-1578.
3. Scharffetter-Kochanek, K., Lu, H., Norman, K., van, N. N., Munoz, F., Grabbe, S., McArthur, M., Lorenzo, I., Kaplan, S., Ley, K., Smith, C. W., Montgomery, C. A., Rich, S., and Beaudet, A. L. (1998) Spontaneous Skin Ulceration and Defective T Cell Function in CD18 Null Mice. J Exp Med. 188, 119-131.
|
Science Writers | Anne Murray |
Illustrators | Diantha La Vine |
Authors | Evan Nair-Gill, Jin Huk Choi, James Butler, and Bruce Beutler |