Phenotypic Mutation 'snowdrop' (pdf version)
Allelesnowdrop
Mutation Type critical splice donor site
Chromosome19
Coordinate25,162,305 bp (GRCm39)
Base Change G ⇒ A (forward strand)
Gene Dock8
Gene Name dedicator of cytokinesis 8
Synonym(s) 1200017A24Rik, 5830472H07Rik, A130095G14Rik
Chromosomal Location 24,976,898-25,179,796 bp (+) (GRCm39)
MGI Phenotype FUNCTION: [Summary is not available for the mouse gene. This summary is for the human ortholog.] This gene encodes a member of the DOCK180 family of guanine nucleotide exchange factors. Guanine nucleotide exchange factors interact with Rho GTPases and are components of intracellular signaling networks. Mutations in this gene result in the autosomal recessive form of the hyper-IgE syndrome. Alternatively spliced transcript variants encoding different isoforms have been described.[provided by RefSeq, Jun 2010]
PHENOTYPE: Mice homozygous for inactivating mutations of this gene exhibit loss of marginal zone B cells, decrease in peritoneal B1 cells and peripheral naive T cells, failure of sustained antibody response after immunization, failure of germinal center persistence, and failure of B cell affinity maturation. [provided by MGI curators]
Accession Number

NCBI RefSeq: NM_028785; MGI:1921396

MappedYes 
Limits of the Critical Region 24999529 - 25202432 bp
Amino Acid Change
Institutional SourceBeutler Lab
Gene Model predicted gene model for protein(s): [ENSMUSP00000025831 ]   † probably from a misspliced transcript
AlphaFold Q8C147
PDB Structure Crystal structure of the DHR-2 domain of DOCK8 in complex with Cdc42 (T17N mutant) [X-RAY DIFFRACTION]
SMART Domains Protein: ENSMUSP00000025831
Gene: ENSMUSG00000052085

DomainStartEndE-ValueType
Pfam:DUF3398 71 164 3.9e-25 PFAM
Pfam:DOCK-C2 557 739 6.7e-49 PFAM
low complexity region 786 803 N/A INTRINSIC
low complexity region 1003 1020 N/A INTRINSIC
low complexity region 1123 1138 N/A INTRINSIC
low complexity region 1236 1246 N/A INTRINSIC
low complexity region 1371 1383 N/A INTRINSIC
Pfam:DHR-2 1534 2060 5e-210 PFAM
Predicted Effect probably null
Meta Mutation Damage Score 0.9593 question?
Is this an essential gene? Probably nonessential (E-score: 0.083) question?
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)(4) Gene trapped(4) Targeted(3)

Lab Alleles
AlleleSourceChrCoordTypePredicted EffectPPH Score
captain_morgan APN 19 25105076 critical splice donor site probably benign
primurus APN 19 25160973 missense probably damaging 1.00
IGL00737:Dock8 APN 19 25160340 missense probably benign 0.00
IGL00755:Dock8 APN 19 25028873 missense probably benign 0.09
IGL00822:Dock8 APN 19 25165773 nonsense probably null
IGL00838:Dock8 APN 19 25152823 nonsense probably null
IGL01419:Dock8 APN 19 25096816 missense probably benign 0.08
IGL01456:Dock8 APN 19 25096863 missense possibly damaging 0.95
IGL01532:Dock8 APN 19 25146805 missense probably damaging 0.99
IGL01602:Dock8 APN 19 25067252 splice site probably benign
IGL01605:Dock8 APN 19 25067252 splice site probably benign
IGL01753:Dock8 APN 19 25038656 splice site probably benign
IGL01843:Dock8 APN 19 25067292 missense probably benign 0.02
IGL02032:Dock8 APN 19 25107769 missense probably damaging 0.99
IGL02073:Dock8 APN 19 25178350 critical splice acceptor site probably null
IGL02192:Dock8 APN 19 25055569 critical splice donor site probably null
IGL02402:Dock8 APN 19 25055509 missense probably benign 0.25
IGL02529:Dock8 APN 19 25078290 nonsense probably null
IGL02728:Dock8 APN 19 25109584 missense probably benign
IGL02739:Dock8 APN 19 25165852 missense probably damaging 1.00
IGL03037:Dock8 APN 19 25063545 missense probably benign 0.02
IGL03104:Dock8 APN 19 25178384 nonsense probably null
IGL03137:Dock8 APN 19 25133312 missense probably benign 0.19
IGL03365:Dock8 APN 19 25077048 missense possibly damaging 0.70
Defenseless UTSW 19 25028927 missense probably benign 0.00
Guardate UTSW 19 25127195 missense probably benign
hillock UTSW 19 25151697 critical splice donor site probably null
Molehill UTSW 19 25107825 missense probably damaging 1.00
Pap UTSW 19 25099805 missense probably benign 0.31
Papilla UTSW 19 25055448 nonsense probably null
warts_and_all UTSW 19 25146865 critical splice donor site probably null
R0021:Dock8 UTSW 19 25140411 missense probably benign 0.01
R0147:Dock8 UTSW 19 25096823 missense probably benign 0.00
R0148:Dock8 UTSW 19 25096823 missense probably benign 0.00
R0294:Dock8 UTSW 19 25165714 missense probably damaging 1.00
R0537:Dock8 UTSW 19 25148941 missense probably benign 0.08
R0630:Dock8 UTSW 19 25038524 missense probably benign 0.10
R1163:Dock8 UTSW 19 25028867 missense probably benign
R1164:Dock8 UTSW 19 25067391 missense probably benign 0.44
R1471:Dock8 UTSW 19 25178400 missense possibly damaging 0.74
R1477:Dock8 UTSW 19 25072914 missense possibly damaging 0.95
R1633:Dock8 UTSW 19 25028927 missense probably benign 0.00
R1803:Dock8 UTSW 19 25109599 missense probably benign 0.00
R1822:Dock8 UTSW 19 25138422 missense probably benign 0.31
R1852:Dock8 UTSW 19 25104492 missense probably benign 0.45
R1916:Dock8 UTSW 19 25038521 missense probably benign 0.02
R1984:Dock8 UTSW 19 25098545 missense probably null
R2311:Dock8 UTSW 19 25160368 missense possibly damaging 0.93
R2341:Dock8 UTSW 19 25177757 missense probably damaging 0.99
R2483:Dock8 UTSW 19 25057241 missense probably benign
R3116:Dock8 UTSW 19 25165858 missense probably benign 0.00
R3157:Dock8 UTSW 19 25127195 missense probably benign
R3623:Dock8 UTSW 19 25057241 missense probably benign
R3624:Dock8 UTSW 19 25057241 missense probably benign
R3800:Dock8 UTSW 19 25141716 missense probably benign 0.08
R3844:Dock8 UTSW 19 25042794 nonsense probably null
R3895:Dock8 UTSW 19 25028865 missense probably benign 0.31
R3901:Dock8 UTSW 19 25078269 missense possibly damaging 0.69
R3959:Dock8 UTSW 19 25162305 critical splice donor site probably null
R4428:Dock8 UTSW 19 25042754 missense probably benign 0.00
R4428:Dock8 UTSW 19 25177863 missense probably damaging 0.98
R4429:Dock8 UTSW 19 25042754 missense probably benign 0.00
R4431:Dock8 UTSW 19 25042754 missense probably benign 0.00
R4545:Dock8 UTSW 19 25165722 missense probably damaging 1.00
R4839:Dock8 UTSW 19 25146858 missense probably benign 0.00
R4897:Dock8 UTSW 19 25159001 missense probably benign 0.00
R4939:Dock8 UTSW 19 25099764 missense probably damaging 1.00
R4995:Dock8 UTSW 19 25135747 missense probably benign 0.02
R5035:Dock8 UTSW 19 25063571 missense probably damaging 0.99
R5294:Dock8 UTSW 19 25038517 missense probably benign 0.01
R5324:Dock8 UTSW 19 25140458 missense probably benign 0.17
R5478:Dock8 UTSW 19 25057186 missense probably benign
R5704:Dock8 UTSW 19 25151586 missense probably damaging 1.00
R5724:Dock8 UTSW 19 25099785 missense probably damaging 1.00
R5745:Dock8 UTSW 19 25107761 missense probably benign 0.02
R5864:Dock8 UTSW 19 25038584 missense probably damaging 0.99
R5870:Dock8 UTSW 19 25109490 missense probably benign
R5893:Dock8 UTSW 19 25099811 missense probably damaging 1.00
R5954:Dock8 UTSW 19 25148983 missense probably damaging 1.00
R6087:Dock8 UTSW 19 25138438 missense probably benign 0.00
R6223:Dock8 UTSW 19 25138416 missense probably benign 0.00
R6391:Dock8 UTSW 19 25072914 missense possibly damaging 0.95
R6759:Dock8 UTSW 19 25104848 missense probably damaging 0.99
R6786:Dock8 UTSW 19 25160386 missense possibly damaging 0.49
R6794:Dock8 UTSW 19 25099805 missense probably benign 0.31
R6818:Dock8 UTSW 19 25146865 critical splice donor site probably null
R6885:Dock8 UTSW 19 25124742 missense possibly damaging 0.95
R6908:Dock8 UTSW 19 25165746 missense probably damaging 1.00
R6923:Dock8 UTSW 19 25072970 missense probably benign
R7001:Dock8 UTSW 19 25077041 missense probably benign
R7141:Dock8 UTSW 19 25158984 missense probably null 0.75
R7203:Dock8 UTSW 19 25158927 missense probably damaging 1.00
R7257:Dock8 UTSW 19 25104449 missense probably benign 0.08
R7296:Dock8 UTSW 19 25162245 missense probably benign 0.00
R7538:Dock8 UTSW 19 25135782 missense probably damaging 1.00
R7555:Dock8 UTSW 19 25152764 missense probably damaging 0.99
R7641:Dock8 UTSW 19 25151697 critical splice donor site probably null
R7764:Dock8 UTSW 19 25074899 missense probably benign
R7859:Dock8 UTSW 19 25160934 missense probably damaging 1.00
R7864:Dock8 UTSW 19 25140864 missense possibly damaging 0.95
R8090:Dock8 UTSW 19 25131606 missense probably damaging 1.00
R8160:Dock8 UTSW 19 25124711 missense probably damaging 1.00
R8287:Dock8 UTSW 19 25107825 missense probably damaging 1.00
R8295:Dock8 UTSW 19 25100600 missense probably benign 0.04
R8443:Dock8 UTSW 19 25133281 missense probably benign 0.04
R8537:Dock8 UTSW 19 25107870 missense probably benign 0.00
R8673:Dock8 UTSW 19 25160867 missense probably damaging 0.96
R8709:Dock8 UTSW 19 25055448 nonsense probably null
R8834:Dock8 UTSW 19 25140834 missense probably benign 0.16
R8991:Dock8 UTSW 19 25165731 missense possibly damaging 0.82
R9292:Dock8 UTSW 19 25160995 splice site probably benign
R9509:Dock8 UTSW 19 25072985 missense probably benign 0.00
R9526:Dock8 UTSW 19 25165739 missense probably benign 0.10
R9622:Dock8 UTSW 19 25098545 missense probably null
R9634:Dock8 UTSW 19 25169585 missense probably damaging 1.00
R9654:Dock8 UTSW 19 25124710 missense probably damaging 1.00
R9670:Dock8 UTSW 19 25148926 missense probably null 0.01
R9699:Dock8 UTSW 19 25133388 critical splice donor site probably null
R9726:Dock8 UTSW 19 25154374 missense probably damaging 0.97
R9765:Dock8 UTSW 19 25146832 missense possibly damaging 0.94
X0027:Dock8 UTSW 19 25138493 missense probably benign
Z1177:Dock8 UTSW 19 25133336 missense probably benign 0.16
Z1177:Dock8 UTSW 19 25109487 missense probably benign 0.05
Mode of Inheritance Autosomal Recessive
Local Stock
Repository
Last Updated 2019-09-04 9:44 PM by Anne Murray
Record Created 2015-12-11 7:49 PM by Jin Huk Choi
Record Posted 2016-01-11
Phenotypic Description

Figure 1. Homozygous snowball mice exhibit diminished T-dependent IgG responses to ovalbumin administered with aluminum hydroxide. IgG 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 snowdrop phenotype was identified among N-ethyl-N-nitrosourea (ENU)-mutagenized G3 mice of the pedigree R3959, some of which  exhibited a diminished T-dependent antibody (IgG) response to ovalbumin administered with aluminum hydroxide (OVA/alum) (Figure 1).

Nature of Mutation

Figure 2. Linkage mapping of the reduced T-dependent antibody response to OVA/alum using a recessive model of inheritance. Manhattan plot shows -log10 P values (Y-axis) plotted against the chromosome positions of 38 mutations (X-axis) identified in the G1 male of pedigree R3959. 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 38 mutations. The reduced T-dependent antibody response to OVA/alum was linked by continuous variable mapping to a mutation in Dock8: a G to A transition at base pair 25,184,941 (v38) on chromosome 19, or base pair 185,431 in the GenBank genomic region NC_000085 encoding the Dock8 gene. Linkage was found with a recessive model of inheritance, wherein five variant homozygotes departed phenotypically from eight homozygous reference mice and 11 heterozygous mice with a P value of 7.041 x 10-6 (Figure 2). A substantial semidominant effect was observed, but the mutation is preponderantly recessive. 

The effect of the mutation at the cDNA and protein levels have not examined, but the mutation is predicted to result in skipping of the 90-base pair exon 43 (out of 48 total exons), which begins after amino acid 1831 (out of 2100 total amino acids) of the protein.

              <--exon 42        <--exon 43 intron 43-->     exon 44-->            <--exon 48

184099 ……TCACATAGACTAGAG ……TTGGATCCTAACAAG gtatggaaaaaata…… GCCTACATTCAG…… ……TCACAGGGCAGCTGA
1827   ……-S--H--R--L--E- ……-L--D--P--N--K-                  -A--Y--I--Q-…… ……-S--Q--G--S--*-
            correct           deleted                                  correct

Genomic numbering corresponds to NC_000085. The donor splice site of intron 43, which is destroyed by the snowdrop mutation, is indicated in blue lettering and the mutated nucleotide is indicated in red.

Illustration of Mutations in
Gene & Protein
Protein Prediction
Figure 3. Domain structure of DOCK8, a member of the DOCKC subfamily. DHR-1 domain shares a weak homology to the C2 domain. The large DHR-2 domain interacts with the nucleotide-free form of Rac and/or Cdc42. The snowball mutation results in abnormal splicing of Dock8 causing deletion of exon 44, which encodes amino acids 1862-1940 C-terminal to the DHR-2 domain. Click on the image to view other mutations found in DOCK8. Click on the mutation for more specific information.

DOCK8 belongs to the DOCK180 superfamily of guanine nucleotide exchange factors (GEFs) that have been shown to activate members of the Rho family of small GTPases (1-4). The DOCK C subfamily (which includes DOCK8 and DOCK7; see the record for moonlight) has dual specificity for Rac and Cdc42 (1;3;5;6). Two domains are shared amongst all DOCK proteins, the catalytic DHR-2 (DOCK homology region 2) or CZH-2 (CDM-zizimin homology 2) domain and the DHR-1 or CZH-1 domain (Figure 3). The DHR-1 domain is located N-terminal to the DHR-2 domain (2;4).

The snowdrop mutation results in abnormal splicing of Dock8 causing deletion of exon 44, which encodes amino acids 1862-1940 C-terminal to the DHR-2 domain.

For more information on Dock8, please see the record for captain morgan.

Putative Mechanism

The Rho GTPases are known regulators of the actin cytoskeleton and affect multiple cellular activities including cell morphology, polarity, migration, proliferation and apoptosis, phagocytosis, cytokinesis, adhesion, vesicular transport, transcription, and neurite extension and retraction. Like DOCK2, DOCK8 is likely to regulate the activity of GTPases and thus be involved in cytoskeletal changes associated with various cellular processes. DOCK8 is proposed to serve as an effector downstream of CD19 and PI3K to promote G protein signaling events critical for integrin polarization at the synapse and for the survival of marginal zone B cells and germinal center (GC) B cells. 

During a T cell-dependent humoral immune response, CD4+ T helper cell subsets including TFH, Th1 and Th2 cells migrate to the T cell/B cell borders in secondary lymphoid organs, and interact with cognate antigen-specific B cells through the pairing of T cell and B cell surface ligands and receptors such as CD40 with its ligand (see the record for walla).  This interaction results in the secretion by T helper cells of certain cytokines known to promote B cell survival, proliferation, and antibody production (7;8).

In humans, DOCK8 deficiency results in an autosomal recessive form of hyper-IgE recurrent infection syndrome (HIES; OMIM #243700) (9;10).  Autosomal dominant HIES is characterized by recurrent Staphylococcus aureus skin abscesses, increased serum IgE, and abnormalities of the connective tissue, skeleton, and dentition (11).  The autosomal recessive form shares hyper-IgE, eosinophilia, and recurrent Staphylococcal infections, but is distinguished from autosomal dominant HIES by the lack of connective tissue and skeletal involvement (12). Patients with DOCK8 deficiency are unusually susceptible to viral infections and virally-caused cancers. Reduced numbers of T, B, and NK cells have been reported along with a selective defect in CD8+ T cell activation (9). However, another study suggests most patients have normal numbers of B and NK cells, a greater decrease in the CD4+ T cell population than the CD8+ T cell population, and a more comprehensive T cell activation defect involving both T cell subsets (10). Both autosomal dominant and autosomal recessive HIES are linked to a lack of Th17 cell function including the failure to produce the interleukin 17 (IL-17) cytokine (12). Th17 are a subset of CD4+ T helper cells that play an important role in the development of autoimmune diseases like rheumatoid arthritis, as well as being critical in the clearance of fungal and extracellular bacterial infections (13).    

The relatively normal initiation of antibody production by mice with Dock8 mutations suggests that the extrafollicular B cell clonal expansion, plasma cell formation, and immunoglobulin class switching, which depends on interactions with T helper cells, is intact. However, subsequent antibody responses and affinity maturation occurring in the germinal centers (GCs) is significantly impaired. The humoral deficits are due to a defect in GC B cell survival and selection during the affinity maturation phase of GC responses (14).

Primers PCR Primer
snowdrop_pcr_F: ATCAACAAATCCTGCCTTGTGC
snowdrop_pcr_R: GGTGCTAAACTCCAGCTTGC

Sequencing Primer
snowdrop_seq_F: CTTGTTCCAAAGGAGTGCAAGC
snowdrop_seq_R: TAAACTCCAGCTTGCACATGTGG
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 401 nucleotides is amplified (chromosome 19, + strand):


1   atcaacaaat cctgccttgt gcttgttcca aaggagtgca agctttcaaa ctgaacttta
61  tatactctgg gcctctcttc cctagggatt ttatggccag tgtttcggtg cagagtttgt
121 ggaagtgata aaagactcta ctccagtgga caaaaccaag ttggatccta acaaggtatg
181 gaaaaaatat atactaaaag tggttatcac actctaccct aacagtagaa aatggcagac
241 ccagaggtct ggactccccc agtccttcaa atgtgaccat gttttagttt gttttttagg
301 ggggaggggt gcacacacat gtatgaatgg agtgtatgca tgtgtgtgtg caggctcact
361 cttccataca tgcccacatg tgcaagctgg agtttagcac c


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

References
Science Writers Anne Murray
Illustrators Peter Jurek
AuthorsJin Huk Choi, James Butler, and Bruce Beutler