Phenotypic Mutation 'dan' (pdf version)
Alleledan
Mutation Type nonsense
Chromosome15
Coordinate100,933,505 bp (GRCm39)
Base Change A ⇒ T (forward strand)
Gene Scn8a
Gene Name sodium channel, voltage-gated, type VIII, alpha
Synonym(s) nmf335, nmf58, NMF335, C630029C19Rik, nur14, mnd2, seal, mnd-2, nmf2, med, ataxia 3, NaCh6, Nav1.6, motor end-plate disease
Chromosomal Location 100,767,739-100,943,819 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 sodium channel alpha subunit gene family. The encoded protein forms the ion pore region of the voltage-gated sodium channel. This protein is essential for the rapid membrane depolarization that occurs during the formation of the action potential in excitable neurons. Mutations in this gene are associated with mental retardation, pancerebellar atrophy and ataxia. Alternate splicing results in multiple transcript variants.[provided by RefSeq, May 2010]
PHENOTYPE: Spontaneous mutant homozygotes have ataxia, dystonia, muscular atrophy, progressive paralysis, Purkinje cell loss, in some cases severe head-tossing and for severe alleles, juvenile lethality. A mild, semidominant ENU allele causes deafness of variable penetrance and severity and mild tremor. [provided by MGI curators]
Accession Number

NCBI RefSeq: NM_001077499, NM_011323; MGI:103169

MappedYes 
Amino Acid Change Lysine changed to Stop codon
Institutional SourceBeutler Lab
Gene Model predicted gene model for protein(s): [ENSMUSP00000080842] [ENSMUSP00000104536] [ENSMUSP00000104537] [ENSMUSP00000104538] [ENSMUSP00000144371] [ENSMUSP00000144013]
AlphaFold no structure available at present
SMART Domains Protein: ENSMUSP00000080842
Gene: ENSMUSG00000023033
AA Change: K1570*

DomainStartEndE-ValueType
Pfam:Ion_trans 131 422 7.4e-82 PFAM
low complexity region 423 452 N/A INTRINSIC
Pfam:Na_trans_cytopl 499 700 3.5e-72 PFAM
low complexity region 701 712 N/A INTRINSIC
Pfam:Ion_trans 750 985 2.2e-57 PFAM
Pfam:Na_trans_assoc 989 1191 2e-59 PFAM
Pfam:Ion_trans 1195 1472 6.2e-69 PFAM
Pfam:Ion_trans 1519 1775 1.2e-56 PFAM
IQ 1892 1914 1.2e-4 SMART
low complexity region 1953 1972 N/A INTRINSIC
Predicted Effect probably null
SMART Domains Protein: ENSMUSP00000104536
Gene: ENSMUSG00000023033
AA Change: K1570*

DomainStartEndE-ValueType
Pfam:Ion_trans 72 322 1.9e-76 PFAM
low complexity region 367 378 N/A INTRINSIC
Pfam:Ion_trans 451 640 1.1e-47 PFAM
Pfam:Na_trans_assoc 655 872 1.9e-71 PFAM
Pfam:Ion_trans 898 1127 4.4e-59 PFAM
PDB:1BYY|A 1129 1181 7e-30 PDB
Pfam:Ion_trans 1220 1429 1.9e-51 PFAM
Pfam:PKD_channel 1281 1436 5.6e-7 PFAM
IQ 1558 1580 1.2e-4 SMART
low complexity region 1619 1638 N/A INTRINSIC
Predicted Effect probably null
SMART Domains Protein: ENSMUSP00000104537
Gene: ENSMUSG00000023033
AA Change: K1580*

DomainStartEndE-ValueType
Pfam:Ion_trans 72 322 2.2e-76 PFAM
low complexity region 335 364 N/A INTRINSIC
Pfam:DUF3451 390 616 8.7e-70 PFAM
Pfam:Ion_trans 697 886 1.3e-47 PFAM
Pfam:Na_trans_assoc 901 1118 2.3e-71 PFAM
Pfam:Ion_trans 1144 1186 9.7e-10 PFAM
Pfam:Ion_trans 1182 1332 1.7e-31 PFAM
PDB:1BYY|A 1334 1386 2e-29 PDB
Pfam:Ion_trans 1425 1634 2.3e-51 PFAM
Pfam:PKD_channel 1486 1641 6.6e-7 PFAM
IQ 1763 1785 1.2e-4 SMART
low complexity region 1824 1843 N/A INTRINSIC
Predicted Effect probably null
SMART Domains Protein: ENSMUSP00000104538
Gene: ENSMUSG00000023033
AA Change: K1570*

DomainStartEndE-ValueType
Pfam:Ion_trans 160 410 2.5e-76 PFAM
low complexity region 423 452 N/A INTRINSIC
Pfam:DUF3451 478 704 9.6e-70 PFAM
Pfam:Ion_trans 785 974 1.4e-47 PFAM
Pfam:Na_trans_assoc 989 1206 2.5e-71 PFAM
Pfam:Ion_trans 1232 1461 5.7e-59 PFAM
PDB:1BYY|A 1463 1515 4e-29 PDB
Pfam:Ion_trans 1554 1763 2.5e-51 PFAM
Pfam:PKD_channel 1615 1770 7.1e-7 PFAM
IQ 1892 1914 1.2e-4 SMART
low complexity region 1953 1972 N/A INTRINSIC
Predicted Effect probably null
SMART Domains Protein: ENSMUSP00000144371
Gene: ENSMUSG00000023033
AA Change: K1529*

DomainStartEndE-ValueType
Pfam:Ion_trans 131 422 4.1e-80 PFAM
low complexity region 423 452 N/A INTRINSIC
Pfam:Na_trans_cytopl 499 700 2.5e-69 PFAM
low complexity region 701 712 N/A INTRINSIC
Pfam:Ion_trans 750 985 1.2e-55 PFAM
Pfam:Na_trans_assoc 989 1191 9.1e-57 PFAM
Pfam:Ion_trans 1195 1274 7.6e-16 PFAM
Pfam:Ion_trans 1270 1431 2.6e-33 PFAM
Pfam:Ion_trans 1478 1734 6.5e-55 PFAM
IQ 1851 1873 6e-7 SMART
low complexity region 1912 1931 N/A INTRINSIC
Predicted Effect probably null
SMART Domains Protein: ENSMUSP00000144013
Gene: ENSMUSG00000023033
AA Change: K1570*

DomainStartEndE-ValueType
Pfam:Ion_trans 131 422 7.4e-82 PFAM
low complexity region 423 452 N/A INTRINSIC
Pfam:Na_trans_cytopl 499 700 3.5e-72 PFAM
low complexity region 701 712 N/A INTRINSIC
Pfam:Ion_trans 750 985 2.2e-57 PFAM
Pfam:Na_trans_assoc 989 1191 2e-59 PFAM
Pfam:Ion_trans 1195 1472 6.2e-69 PFAM
Pfam:Ion_trans 1519 1775 1.2e-56 PFAM
IQ 1892 1914 1.2e-4 SMART
low complexity region 1953 1972 N/A INTRINSIC
Predicted Effect probably null
Meta Mutation Damage Score 0.9755 question?
Is this an essential gene? Probably essential (E-score: 0.790) question?
Phenotypic Category Autosomal Recessive
Candidate Explorer Status loading ...
Single pedigree
Linkage Analysis Data
Penetrance  
Alleles Listed at MGI

All alleles(22) : Targeted(4) Gene trapped(3) Transgenic(1) Spontaneous(6) Chemically induced(8)

Lab Alleles
AlleleSourceChrCoordTypePredicted EffectPPH Score
IGL00230:Scn8a APN 15 100853413 unclassified probably benign
IGL00979:Scn8a APN 15 100853287 unclassified probably benign
IGL01339:Scn8a APN 15 100930082 missense probably benign
IGL01992:Scn8a APN 15 100866938 missense probably damaging 1.00
IGL02215:Scn8a APN 15 100927453 splice site probably null
IGL02311:Scn8a APN 15 100911164 missense probably damaging 0.97
IGL02404:Scn8a APN 15 100937611 missense probably damaging 1.00
IGL02652:Scn8a APN 15 100911357 missense probably damaging 0.98
IGL02690:Scn8a APN 15 100868135 missense probably damaging 1.00
IGL02704:Scn8a APN 15 100905943 missense possibly damaging 0.94
IGL03084:Scn8a APN 15 100915053 missense probably damaging 1.00
IGL03108:Scn8a APN 15 100872496 missense probably benign
IGL03224:Scn8a APN 15 100933520 missense probably damaging 1.00
nymph UTSW 15 100933527 missense probably damaging 1.00
Tremord UTSW 15 100911385 missense probably damaging 1.00
3-1:Scn8a UTSW 15 100937820 missense probably benign 0.04
PIT4280001:Scn8a UTSW 15 100855370 missense probably damaging 1.00
PIT4508001:Scn8a UTSW 15 100927573 missense probably damaging 0.98
R0010:Scn8a UTSW 15 100911454 missense probably damaging 1.00
R0010:Scn8a UTSW 15 100911454 missense probably damaging 1.00
R0254:Scn8a UTSW 15 100916245 missense probably damaging 1.00
R0412:Scn8a UTSW 15 100906187 splice site probably benign
R0538:Scn8a UTSW 15 100933505 nonsense probably null
R0539:Scn8a UTSW 15 100914449 missense probably damaging 1.00
R0631:Scn8a UTSW 15 100933418 missense probably damaging 1.00
R0726:Scn8a UTSW 15 100870711 missense probably damaging 1.00
R0945:Scn8a UTSW 15 100913668 missense possibly damaging 0.54
R0967:Scn8a UTSW 15 100933527 missense probably damaging 1.00
R1164:Scn8a UTSW 15 100938043 missense probably benign 0.06
R1283:Scn8a UTSW 15 100867052 missense possibly damaging 0.82
R1368:Scn8a UTSW 15 100933422 missense probably damaging 1.00
R1633:Scn8a UTSW 15 100927696 missense probably benign 0.01
R1669:Scn8a UTSW 15 100909001 missense probably damaging 1.00
R1694:Scn8a UTSW 15 100853409 nonsense probably null
R1735:Scn8a UTSW 15 100913742 missense possibly damaging 0.94
R1773:Scn8a UTSW 15 100937496 missense probably damaging 0.97
R1940:Scn8a UTSW 15 100868085 missense probably benign 0.22
R1996:Scn8a UTSW 15 100922260 missense probably damaging 1.00
R2107:Scn8a UTSW 15 100916244 missense probably damaging 0.99
R2251:Scn8a UTSW 15 100914987 missense probably benign 0.02
R2516:Scn8a UTSW 15 100867043 missense probably benign 0.05
R2917:Scn8a UTSW 15 100937613 missense probably damaging 1.00
R3417:Scn8a UTSW 15 100869549 splice site probably benign
R3896:Scn8a UTSW 15 100933379 missense probably benign
R4024:Scn8a UTSW 15 100937674 missense probably damaging 1.00
R4050:Scn8a UTSW 15 100911294 nonsense probably null
R4193:Scn8a UTSW 15 100869484 missense probably damaging 1.00
R4212:Scn8a UTSW 15 100854954 missense possibly damaging 0.88
R4358:Scn8a UTSW 15 100838014 missense probably benign 0.00
R4396:Scn8a UTSW 15 100870711 missense probably damaging 1.00
R4428:Scn8a UTSW 15 100881784 missense probably damaging 1.00
R4452:Scn8a UTSW 15 100854972 missense possibly damaging 0.95
R4631:Scn8a UTSW 15 100914384 nonsense probably null
R4693:Scn8a UTSW 15 100913572 missense probably damaging 1.00
R4765:Scn8a UTSW 15 100938352 missense probably benign 0.07
R4777:Scn8a UTSW 15 100913832 missense probably damaging 1.00
R4949:Scn8a UTSW 15 100927663 missense probably damaging 1.00
R4997:Scn8a UTSW 15 100854935 missense probably damaging 1.00
R5246:Scn8a UTSW 15 100908938 missense probably damaging 1.00
R5566:Scn8a UTSW 15 100872415 missense probably damaging 1.00
R5875:Scn8a UTSW 15 100870703 nonsense probably null
R6031:Scn8a UTSW 15 100881865 missense probably damaging 1.00
R6031:Scn8a UTSW 15 100881865 missense probably damaging 1.00
R6057:Scn8a UTSW 15 100872548 missense possibly damaging 0.94
R6114:Scn8a UTSW 15 100938477 missense probably damaging 0.99
R6362:Scn8a UTSW 15 100837996 splice site probably null
R6535:Scn8a UTSW 15 100857588 intron probably benign
R6677:Scn8a UTSW 15 100866953 missense probably damaging 1.00
R6687:Scn8a UTSW 15 100872508 missense probably benign 0.12
R6701:Scn8a UTSW 15 100937977 missense probably damaging 1.00
R6719:Scn8a UTSW 15 100908896 critical splice acceptor site probably null
R6739:Scn8a UTSW 15 100913836 missense possibly damaging 0.82
R6769:Scn8a UTSW 15 100933445 missense probably benign
R6786:Scn8a UTSW 15 100930096 missense probably benign 0.00
R6849:Scn8a UTSW 15 100853468 splice site probably null
R7108:Scn8a UTSW 15 100937659 missense probably benign 0.01
R7215:Scn8a UTSW 15 100927711 missense possibly damaging 0.80
R7217:Scn8a UTSW 15 100868108 missense probably benign 0.00
R7219:Scn8a UTSW 15 100866984 missense probably damaging 1.00
R7356:Scn8a UTSW 15 100855460 missense probably damaging 1.00
R7479:Scn8a UTSW 15 100853358 missense probably damaging 0.99
R7816:Scn8a UTSW 15 100908917 missense possibly damaging 0.63
R7985:Scn8a UTSW 15 100914843 splice site probably null
R8112:Scn8a UTSW 15 100927718 missense probably benign 0.27
R8263:Scn8a UTSW 15 100881736 missense probably damaging 1.00
R8305:Scn8a UTSW 15 100938387 missense probably benign 0.01
R8489:Scn8a UTSW 15 100867014 missense probably damaging 1.00
R8983:Scn8a UTSW 15 100900030 missense possibly damaging 0.81
R9034:Scn8a UTSW 15 100927642 missense probably damaging 0.98
R9050:Scn8a UTSW 15 100906161 missense possibly damaging 0.80
R9240:Scn8a UTSW 15 100915068 nonsense probably null
R9249:Scn8a UTSW 15 100914456 missense probably benign 0.00
R9462:Scn8a UTSW 15 100930159 missense
R9599:Scn8a UTSW 15 100911172 missense probably damaging 1.00
R9609:Scn8a UTSW 15 100834407 missense possibly damaging 0.91
R9653:Scn8a UTSW 15 100937947 missense probably damaging 1.00
R9794:Scn8a UTSW 15 100933332 missense probably benign 0.00
X0066:Scn8a UTSW 15 100937962 missense probably damaging 1.00
X0066:Scn8a UTSW 15 100937961 missense probably damaging 1.00
Z1176:Scn8a UTSW 15 100931399 missense probably damaging 1.00
Z1177:Scn8a UTSW 15 100938103 missense probably benign 0.00
Mode of Inheritance Autosomal Recessive
Local Stock Live Mice
MMRRC Submission 038240-MU
Last Updated 2018-04-25 1:58 PM by Anne Murray
Record Created 2013-05-19 9:56 AM by Jennifer Weatherly
Record Posted 2014-03-28
Phenotypic Description
Figure 1. Video of the dan phenotype.

Dan was identified as a visible mutant amongst N-ethyl-N-nitrosourea (ENU)-mutagenized G3 mice; the phenotype is characterized by the loss of the use of the hind legs (Figure 1).

Nature of Mutation

Whole exome HiSeq sequencing of the G1 grandsire identified 129 mutations. Four G3 mice with the dan phenotype and 22 unaffected mice from a single pedigree were genotyped at all mutation sites. Three mutations (in Rapgef3, Slc11a2, and Scn8a) on chromosome 15 were homozygous in all four of the dan mice and either heterozygous (n = 12) or wild-type (n = 10) in the unaffected mice (-log10[p(non-linkage)] = 5.922). The dan phenotype resembles that of other Scn8a mutant mice (described in the “Putative Mechanism” section, below), indicating that the mutation in Scn8a is causative. The Scn8a mutation is an A to T transversion at base pair 101,035,624 (v38) on Chromosome 15, or base pair 165,802 in the GenBank genomic region NC_000081 encoding Scn8a. The mutation corresponds to residue 4861 in the mRNA sequence NM_001077499 (isoform 1) within exon 26 of 27 total exons and residue 4762 in the mRNA sequence NM_011323 (isoform 2) within exon 25 of 26 total exons.

165787 TGCGAGTGTGTGCTCAAAATGTTTGCCTTGAGA

1565   -C--E--C--V--L--K--M--F--A--L--R-

Genomic numbering is shown, corresponding to NC_000081. The mutated nucleotide is indicated in red.  The mutation results in substitution of a lysine (K) for a premature stop codon (*) at amino acid 1570 in both isoforms of the SCN8A sodium channel.

Illustration of Mutations in
Gene & Protein
Protein Prediction

Figure 2. A, Domain structure of SCN8A. B, Transmembrane organization of sodium channel subunits. Cylinders represent probable α-helical transmembrane segments (numbered 1-6 for each domain) with green cylinders indicating the pore-lining segments and yellow cylinders indicating the S4 voltage sensors (positively charged residues are represented by the + signs). The extracellular immunoglobulin-like domains of the β1 and β2 subunits are shown and a site of interaction between the  α and β1 subunit is indicated by the wavy, gray lines. Consensus phosphorylation sites are indicated by the orange circles. Purple circles represent the outer (EEDD) and inner (DEKA) rings of amino acid residues forming the ion selectivity filter and the TTX binding site. White circles indicate residues critical for fast inactivation. Binding sites for α- and β-scorpion toxins and anesthetic drugs are also shown. The dan mutation (K1570*) is within the S2 helix of domain IV. Image is interactive; click to see another Scn8a mutation.

Scn8a encodes the voltage-gated sodium channel SCN8A (alternatively, Nav1.6). Nav1.6 contains four homologous domains (I-IV) that each consist of six transmembrane α-helices (S1-S6) (1;2). The S4 helices constitute the voltage sensors of the channel. A membrane reentrant extracellular loop (i.e., the pore loop) is located between S5 and S6 in each of the four domains and lines the extracellular, narrow entrance to the pore (3;4).  The S5 and S6 helices line the intracellular, wider exit of the pore, while amino acids (F1752 and Y1759) in in the S6 helix contribute to drug binding (5;6). The dan mutation (K1570*) is within the S2 helix of domain IV (Figure 2). Protein and mRNA expression have not been examined.

For more information about Scn8a, please see the record for TremorD.

Putative Mechanism

Mutations in Scn8a (e.g., Scn8amed, MGI:1856078; Scn8amed-tg, MGI:1856414; Scn8amed-J, MGI:1856079) lead to a range of neurological disorders in mice such as tremor, ataxia, early-onset progressive paralysis of the hind limbs, severe muscle atrophy, degeneration of Purkinje cells, and juvenile lethality (7). Loss of Scn8a expression in Scn8amed homozygous mice resulted in reduced conduction velocity of the sciatic nerve between postnatal day 17 and 23 and subsequent failure of transmission at the neuromuscular junction and muscle denervation (8-10); these defects may underlie the hindlimb paralysis observed in these and dan homozygous mice.

The dan mutation may result in degradation of the Scn8a mRNA by nonsense-mediated decay; translated protein may be truncated and lack helices S3-S6 and the pore loop of domain IV as well as the cytoplasmic C-terminal tail. 

Primers PCR Primer
dan_pcr_F: TTCGTCACGCAACAAGCCTTCG
dan_pcr_R: GCTCTCAGAAATCTGGTCCTCACAC

Sequencing Primer
dan_seq_F: ACAAGCCTTCGACATCGTG
dan_seq_R: TCCTCACACCTGGGGTC
Genotyping

Dan genotyping is performed by amplifying the region containing the mutation using PCR followed by sequencing of the amplified region to detect the nucleotide change.  The following primers were used for PCR amplification:

Primers for PCR amplification

Dan(F):  5’- TTCGTCACGCAACAAGCCTTCG-3’

Dan(R):  5’- GCTCTCAGAAATCTGGTCCTCACAC-3’

Primers for sequencing

Dan(F):  5’- ACAAGCCTTCGACATCGTG-3’

Dan(R):  5’-TCCTCACACCTGGGGTC-3’

PCR program

1) 94° C        2:00

2) 94° C        0:30

3) 57° C        0:30

4) 72° C        1:00

5) repeat steps (2-4) 29x

6) 72° C        7:00

7) 4° C          hold

The following sequence of 418 nucleotides (from Genbank genomic region: NC_000081 of the linear genomic sequence of Scn8a) is amplified:

165640                                           t tcgtcacgca acaagccttc

165661 gacatcgtga tcatgatgct tatctgcctt aacatggtga ccatgatggt ggagacagac

165721 acacagagca agcagatgga gaacattctc tactggatta atctggtctt cgtcatcttc

165781 ttcacctgcg agtgtgtgct caaaatgttt gccttgagac actactattt caccattggc

165841 tggaacatct ttgactttgt ggtggtcatt ctctccattg tgggtgagtg ggtgcagcca

165901 caggaagggg ggtgagtggg cggggccaca ggaagggtga gtgggcgggg ccacaggaag

165961 agggggtgag tgggcggggc cacgggaaga ggggatgagt gggcggggcc acaggaagga

166021 ccaggacccc aggtgtgagg accagatttc tgagagc

PCR primer binding sites are underlined; Sequencing primer binding sites are highlighted; the mutated nucleotide is in red.

References
Science Writers Anne Murray
Illustrators Diantha La Vine, Peter Jurek
AuthorsJennifer Weatherly Tiana Purrington