Phenotypic Mutation 'newhome' (pdf version)
Allelenewhome
Mutation Type missense
Chromosome10
Coordinate77,395,515 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

MappedYes 
Amino Acid Change Valine changed to Isoleucine
Institutional SourceBeutler Lab
Gene Model predicted gene model for protein(s): [ENSMUSP00000000299] [ENSMUSP00000137734] [ENSMUSP00000137865]
AlphaFold no structure available at present
SMART Domains Protein: ENSMUSP00000000299
Gene: ENSMUSG00000000290
AA Change: V539I

DomainStartEndE-ValueType
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 benign

PolyPhen 2 Score 0.001 (Sensitivity: 0.99; Specificity: 0.15)
(Using ENSMUST00000000299)
SMART Domains Protein: ENSMUSP00000137734
Gene: ENSMUSG00000000290

DomainStartEndE-ValueType
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

DomainStartEndE-ValueType
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 0.0898 question?
Is this an essential gene? Possibly nonessential (E-score: 0.477) question?
Phenotypic Category Autosomal Recessive
Candidate Explorer Status loading ...
Single pedigree
Linkage Analysis Data
Penetrance  
Alleles Listed at MGI

All mutations/alleles(10) : Chemically induced (ENU)(1) Targeted(9)

Lab Alleles
AlleleSourceChrCoordTypePredicted EffectPPH 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
nibbler UTSW 10 77397050 critical splice donor site probably null
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 9:44 PM by Diantha La Vine
Record Created 2015-08-04 10:24 AM by Doan Dao
Record Posted 2018-06-13
Phenotypic Description

Figure 1. Newhome mice exhibited increased IFN-β secretion from macrophages in response to dsDNA treatment. IFN-β levels were determined by ELISA. Normalized data are shown. Abbreviations: WT, wild-type; REF, homozygous reference mice; HET, heterozygous variant mice; VAR, homozygous variant mice. Mean (μ) and standard deviation (σ) are indicated.

The newhome phenotype was identified among G3 mice of the pedigree R2374, some of which showed increased secretion of IFN-β from isolated peritoneal macrophages upon stimulation with double-stranded DNA (dsDNA) (Figure 1).

Nature of Mutation

Figure 2. Linkage mapping of the increased IFN-β secretion after dsDNA stimulation using a recessive model of inheritance. Manhattan plot shows -log10 P values (Y-axis) plotted against the chromosome positions of 34 mutations (X-axis) identified in the G1 male of pedigree R2374.  Normalized phenotype data are shown for single locus linkage analysis without consideration of G2 dam identity.  Horizontal pink and red lines represent thresholds of P = 0.05, and the threshold for P = 0.05 after applying Bonferroni correction, respectively.

Whole exome HiSeq sequencing of the G1 grandsire identified 34 mutations. The cytoplasmic dsDNA sensing-associated phenotype was linked by continuous variable mapping to a mutation in Itgb2:  a G to A transition at base pair 77,559,681 (v38) on chromosome 10, or base pair 29,354 in the GenBank genomic region NC_000076.  Linkage was found with a recessive model of inheritance (P = 1.939 x 10-10), wherein five variant homozygotes departed phenotypically from 20 homozygous reference mice and 28 heterozygous mice (Figure 2).  

The mutation corresponds to residue 1,730 in the mRNA sequence NM_008404 within exon 12 of 16 total exons.

1715 TGCGAGTGTGACAATGTCAACTGTGAGAGATAT

534  -C--E--C--D--N--V--N--C--E--R--Y-

 
The mutated nucleotide is indicated in red.  The mutation results in a valine (V) to isoleucine (I) substitution at position 539 (V539I) in the integrin β2 protein, and is strongly predicted by PolyPhen-2 to be benign (score = 0.001) (1).
Illustration of Mutations in
Gene & Protein
Protein Prediction

Figure 3. Domain structure of α and β integrins. Some α subunits do not contain an I domain. The β-propeller and I-like domains make numerous contacts in the native integrin dimer. The location of the newhome mutation results in a valine to isoleucine  substitution at position 539. Abbreviations: HD, hybrid domain; TM, transmembrane domain. This image is interactive. Other mutations found in CD18 are noted in red. Click on the mutations for more specific information.

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 3) [reviewed in (2)]. 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 newhome mutation results in a valine (V) to isoleucine (I) substitution at position 539 (V539I) within the second I-EGF domain.

For more information about Itgb2, please see the record for Joker.

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 (3). 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) (4), 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.

Newhome mice display a phenotype distinct from that of the CD18 null or hypomorphic mutant, and from humans with LAD. In particular, newhome mice do not develop the dermatitis or facial/submandibular inflammation observed in CD18-/- mice. No spontaneous infections have been observed in newhome mice, as they are in the CD18 null mutant and in LAD patients.

The protein encoded by Itgb2newhome may be hypermorphic, causing changes in the rigid 3D structure formed by the four I-EGF domains.

Primers PCR Primer
newhome_pcr_F: CTCCCACATCATACCAGGTGTG
newhome_pcr_R: GGACATGACTATTCCCCTCCAC

Sequencing Primer
newhome_seq_F: CACATCATACCAGGTGTGAGTCTG
newhome_seq_R: ATGACTATTCCCCTCCACAACCTTC
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 417 nucleotides is amplified (chromosome 10, + strand):


1   ctcccacatc ataccaggtg tgagtctggc tacattggga aaaactgtga gtgccagact
61  cagggtcgga gcagccagga gctggagaga aactgtcgga aggacaatag ttccatcgtg
121 tgctcagggc ttggggactg catctgtggg cagtgtgtat gccataccag tgacgtcccc
181 aacaaagaga tctttgggca atactgcgag tgtgacaatg tcaactgtga gagatataac
241 agccaagtct gcggtggctc aggttagtgt tgggggcttc cttgtccatt tgtaccagga
301 gatcttaagt gggacatggg ggggggagca gacagacaag gctagcaatg acctgtatct
361 gtgtctgtgt cagtcagcgt tgtcctggga aggttgtgga ggggaatagt catgtcc


Primer binding sites are underlined and the sequencing primers are highlighted; the mutated nucleotide is shown in red.

References
Science Writers Anne Murray
Illustrators Diantha La Vine
AuthorsDoan Dao, Hexin Shi, Zhao Zhang, Ying Wang, Lei Sun and Bruce A. Beutler