Phenotypic Mutation 'nomoco' (pdf version)
Allelenomoco
Mutation Type missense
Chromosome7
Coordinate141,307,456 bp (GRCm39)
Base Change T ⇒ C (forward strand)
Gene Muc2
Gene Name mucin 2
Synonym(s) 2010015E03Rik
Chromosomal Location 141,276,583-141,308,428 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 mucin protein family. Mucins are high molecular weight glycoproteins produced by many epithelial tissues. The protein encoded by this gene is secreted and forms an insoluble mucous barrier that protects the gut lumen. The protein polymerizes into a gel of which 80% is composed of oligosaccharide side chains by weight. The protein features a central domain containing tandem repeats rich in threonine and proline that varies between 50 and 115 copies in different individuals. Downregulation of this gene has been observed in patients with Crohn disease and ulcerative colitis. [provided by RefSeq, Oct 2016]
PHENOTYPE: Homozygotes for a point mutation have soft feces at weaning and develop diarrhea associated with malapsorption syndrome. Homozygous null mutants pass blood in their feces at 6 months, and 65% of null mutants have intestinal tumors at 1 year. [provided by MGI curators]
Accession Number

NCBI RefSeq: NM_023566, MGI: 1339364

MappedYes 
Amino Acid Change Cysteine changed to Arginine
Institutional SourceBeutler Lab
Gene Model predicted gene model for protein(s): [ENSMUSP00000026590]
AlphaFold no structure available at present
SMART Domains Protein: ENSMUSP00000026590
Gene: ENSMUSG00000025515
AA Change: C365R

DomainStartEndE-ValueType
C8 1 63 1.65e-11 SMART
VWC 120 188 5.48e-2 SMART
VWC 229 293 2.38e-11 SMART
Blast:VWD 299 363 4e-17 BLAST
CT 380 463 3.6e-35 SMART
Predicted Effect probably damaging

PolyPhen 2 Score 0.999 (Sensitivity: 0.14; Specificity: 0.99)
(Using ENSMUST00000026590)
Predicted Effect probably damaging

PolyPhen 2 Score 1.000 (Sensitivity: 0.00; Specificity: 1.00)
(Using ENSMUST00000187945)
Meta Mutation Damage Score 0.6467 question?
Is this an essential gene? Probably nonessential (E-score: 0.099) question?
Phenotypic Category Autosomal Recessive
Candidate Explorer Status loading ...
Single pedigree
Linkage Analysis Data
Penetrance  
Alleles Listed at MGI

All alleles(9) : Chemically induced (ENU)(5) Radiation induced(1) Targeted(3)

Lab Alleles
AlleleSourceChrCoordTypePredicted EffectPPH Score
Eeyore APN 7 141693356 missense probably benign 0.35
kenny APN 7 nonsense
Winnie APN 7 141286029 missense probably damaging 1.00
IGL01303:Muc2 APN 7 141306132 missense probably benign
IGL01482:Muc2 APN 7 141307797 missense probably damaging 0.96
IGL01875:Muc2 APN 7 141306477 missense probably damaging 0.99
IGL02088:Muc2 APN 7 141305241 missense probably damaging 1.00
IGL02415:Muc2 APN 7 141305609 nonsense probably null
IGL02548:Muc2 APN 7 141305594 missense probably damaging 1.00
IGL02836:Muc2 APN 7 141300450 unclassified probably benign
IGL03196:Muc2 APN 7 141301367 missense probably damaging 0.97
Muskatenwein UTSW 7 141307176 missense unknown
Schlendrian UTSW 7 141281925 missense probably damaging 1.00
Seco UTSW 7 141284976 missense probably damaging 1.00
BB001:Muc2 UTSW 7 141281631 missense probably damaging 1.00
BB011:Muc2 UTSW 7 141281631 missense probably damaging 1.00
E0370:Muc2 UTSW 7 141282598 missense probably damaging 1.00
R0127:Muc2 UTSW 7 141302691 missense probably benign 0.00
R0179:Muc2 UTSW 7 141302708 missense probably damaging 1.00
R0201:Muc2 UTSW 7 141699185 frame shift probably null
R0299:Muc2 UTSW 7 141306466 missense probably damaging 1.00
R0547:Muc2 UTSW 7 141699185 frame shift probably null
R0699:Muc2 UTSW 7 141306037 missense probably damaging 1.00
R0900:Muc2 UTSW 7 141699185 frame shift probably null
R1348:Muc2 UTSW 7 141699185 frame shift probably null
R1466:Muc2 UTSW 7 141302711 missense probably damaging 1.00
R1466:Muc2 UTSW 7 141302711 missense probably damaging 1.00
R1625:Muc2 UTSW 7 141283405 missense probably damaging 1.00
R2010:Muc2 UTSW 7 141287444 missense probably damaging 0.99
R2149:Muc2 UTSW 7 141699185 frame shift probably null
R2163:Muc2 UTSW 7 141699185 frame shift probably null
R3008:Muc2 UTSW 7 141281347 missense possibly damaging 0.93
R3110:Muc2 UTSW 7 141299225 unclassified probably benign
R3112:Muc2 UTSW 7 141299225 unclassified probably benign
R3424:Muc2 UTSW 7 141279595 missense probably damaging 0.99
R3786:Muc2 UTSW 7 141283590 missense probably benign 0.01
R3854:Muc2 UTSW 7 141308081 missense probably damaging 1.00
R3964:Muc2 UTSW 7 141286233 missense probably benign 0.17
R3965:Muc2 UTSW 7 141286233 missense probably benign 0.17
R3966:Muc2 UTSW 7 141286233 missense probably benign 0.17
R3973:Muc2 UTSW 7 141300541 unclassified probably benign
R3974:Muc2 UTSW 7 141300541 unclassified probably benign
R3976:Muc2 UTSW 7 141300541 unclassified probably benign
R4327:Muc2 UTSW 7 141281577 missense probably damaging 0.96
R4694:Muc2 UTSW 7 141306082 missense probably damaging 1.00
R4764:Muc2 UTSW 7 141299345 missense possibly damaging 0.88
R4769:Muc2 UTSW 7 141286260 critical splice donor site probably null
R4798:Muc2 UTSW 7 141307877 missense probably benign 0.01
R4900:Muc2 UTSW 7 141303280 missense probably benign 0.32
R5383:Muc2 UTSW 7 141307456 missense probably damaging 1.00
R5489:Muc2 UTSW 7 141305169 missense probably benign 0.00
R5615:Muc2 UTSW 7 141277446 missense probably damaging 1.00
R5856:Muc2 UTSW 7 141299381 unclassified probably benign
R5919:Muc2 UTSW 7 141281171 missense probably damaging 0.97
R5953:Muc2 UTSW 7 141287951 missense probably damaging 0.96
R5979:Muc2 UTSW 7 141305143 missense probably damaging 0.99
R5979:Muc2 UTSW 7 141283493 splice site probably null
R6175:Muc2 UTSW 7 141282875 missense probably damaging 1.00
R6213:Muc2 UTSW 7 141305151 missense probably damaging 1.00
R6281:Muc2 UTSW 7 141306140 missense probably damaging 1.00
R6321:Muc2 UTSW 7 141287397 missense probably benign 0.28
R6390:Muc2 UTSW 7 141305883 missense probably damaging 0.97
R6485:Muc2 UTSW 7 141300473 unclassified probably benign
R6582:Muc2 UTSW 7 141282941 missense probably benign 0.00
R6683:Muc2 UTSW 7 141305214 missense probably benign 0.38
R6896:Muc2 UTSW 7 141306432 missense possibly damaging 0.48
R6906:Muc2 UTSW 7 141284976 missense probably damaging 1.00
R6924:Muc2 UTSW 7 141284077 missense possibly damaging 0.87
R7040:Muc2 UTSW 7 141305194 missense unknown
R7222:Muc2 UTSW 7 141290758 missense
R7251:Muc2 UTSW 7 141278965 missense possibly damaging 0.91
R7282:Muc2 UTSW 7 141306481 missense
R7315:Muc2 UTSW 7 141276645 missense probably damaging 0.99
R7421:Muc2 UTSW 7 141301863 missense
R7556:Muc2 UTSW 7 141307439 missense
R7651:Muc2 UTSW 7 141290750 missense
R7710:Muc2 UTSW 7 141287452 missense possibly damaging 0.92
R7776:Muc2 UTSW 7 141290942 missense
R7813:Muc2 UTSW 7 141282543 splice site probably null
R7843:Muc2 UTSW 7 141281662 missense probably benign 0.03
R7869:Muc2 UTSW 7 141303471 missense
R7924:Muc2 UTSW 7 141281631 missense probably damaging 1.00
R7993:Muc2 UTSW 7 141308173 missense
R8053:Muc2 UTSW 7 141284575 missense probably benign 0.01
R8068:Muc2 UTSW 7 141298422 missense
R8099:Muc2 UTSW 7 141299175 splice site probably null
R8192:Muc2 UTSW 7 141305215 missense
R8194:Muc2 UTSW 7 141290801 missense
R8545:Muc2 UTSW 7 141306130 missense unknown
R8701:Muc2 UTSW 7 141281850 missense probably damaging 1.00
R8883:Muc2 UTSW 7 141287469 missense probably damaging 0.98
R8894:Muc2 UTSW 7 141280758 missense probably damaging 1.00
R8905:Muc2 UTSW 7 141279643 missense probably benign 0.00
R9024:Muc2 UTSW 7 141287936 missense probably damaging 0.98
R9032:Muc2 UTSW 7 141287058 missense probably damaging 1.00
R9085:Muc2 UTSW 7 141287058 missense probably damaging 1.00
R9091:Muc2 UTSW 7 141290816 missense
R9104:Muc2 UTSW 7 141286224 missense probably damaging 1.00
R9114:Muc2 UTSW 7 141287983 nonsense probably null
R9270:Muc2 UTSW 7 141290816 missense
R9297:Muc2 UTSW 7 141302759 missense
R9325:Muc2 UTSW 7 141298559 missense
R9354:Muc2 UTSW 7 141307157 missense
R9386:Muc2 UTSW 7 141279389 missense probably damaging 1.00
R9529:Muc2 UTSW 7 141287453 missense possibly damaging 0.55
R9550:Muc2 UTSW 7 141308242 missense probably damaging 1.00
R9583:Muc2 UTSW 7 141300559 missense
R9607:Muc2 UTSW 7 141305190 missense
R9646:Muc2 UTSW 7 141276643 missense probably benign
R9651:Muc2 UTSW 7 141288014 missense probably damaging 0.99
R9774:Muc2 UTSW 7 141285811 missense probably benign
R9784:Muc2 UTSW 7 141280785 nonsense probably null
Z1176:Muc2 UTSW 7 141300451 missense
Z1177:Muc2 UTSW 7 141298531 missense
Mode of Inheritance Autosomal Recessive
Local Stock
Repository
Last Updated 2019-09-04 9:38 PM by Anne Murray
Record Created 2017-09-11 8:28 PM
Record Posted 2017-10-12
Phenotypic Description

Figure 1. Nomoco mice exhibited susceptibility to dextran sodium sulfate (DSS)-induced colitis seven days after DSS treatment. 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 nomoco phenotype was identified among N-ethyl-N-nitrosourea (ENU)-mutagenized G3 mice of the pedigree R5383, some of which showed susceptibility to dextran sodium sulfate (DSS)-induced colitis seven days after DSS treatment (Figure 1).

Nature of Mutation

Figure 2. Linkage mapping of the DSS susceptibility phenotype using a recessive model of inheritance. Manhattan plot shows -log10 P values (Y-axis) plotted against the chromosome positions of 63 mutations (X-axis) identified in the G1 male of pedigree R5383. 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 63 mutations. The colitis susceptibility phenotype was linked by continuous variable mapping to a mutation in Muc2:  a T to C transition at base pair 141,753,719 (v38) on chromosome 7, or base pair 63,380 in the GenBank genomic region NC_000073 encoding Muc2.  Linkage was found with a recessive model of inheritance, wherein four variant homozygotes departed phenotypically from 19 homozygous reference mice and 20 heterozygous mice with a P value of 1.9 x 10-8 (Figure 3).  

The mutation corresponds to residue 6,852 in the mRNA sequence NM_023566 within exon 14 of 16 total exons.

6837 ATGCCTAATGGCTGCTGTAAGACATGCATCCCT

360  -M--P--N--G--C--C--K--T--C--I--P-

 

The mutated nucleotide is indicated in red.  The mutation results in a cysteine to arginine substitution at position 365 (C365R) in the MUC2 protein, and is strongly predicted by PolyPhen-2 to be damaging (score = 1.000).

Illustration of Mutations in
Gene & Protein
Protein Prediction
Figure 3.  Domain structure of Mucin2. The nomoco mutation results in a cysteine to arginine substitution at position 365 (C365R).  S=Signal sequence; D=D domains (homology to VWF mediates trimerization); CR=Cystein-rich domain; TR= Tandem repeat domain (heavily O glycosylated); GDPH=GDPH autocatalytic proteolytic site; B=B domain (homology to VWF); C= C domain (homology to VWF); CK= Cysteine-knot domain (homology to VWF mediates dimerization). This image is interactive. Click on the image to view other mutations found in Mucin2 (red). Click on the mutations for more specific information.

The Muc2 gene encodes a secreted member of the mucin family of proteins. The structural feature that is common to all mucins is the tandem-repeat domain, which contains tandem repeats of identical or highly similar sequences that are rich in serine, threonine and proline residues. MUC2 contains two tandem repeat domains that are located in the middle of the protein, and shows extensive polymorphism due to the variable number of tandem repeats that can be present (1-5). For human MUC2, the first tandem repeat domain is invariant and consists of 21 repeats of a 23 amino acid sequence, while the second domain contains between 51-115 repeats of a 16 amino acid sequence (1;6). Thus, the size of the MUC2 protein varies widely between individuals and ethnicities with the most common allelic form containing over 5100 amino acids (2). In mouse, the central repetitive region of MUC2 contains two distinct repetitive regions consisting of 8 and 10 amino acid tandem repeats, respectively (7). MUC2 has several highly conserved cysteine-rich domains; many have high homology with von Willebrand platelet aggregating factor (VWF) (8). These include the five D domains known as D1, D2, D’, D3 and D4, as well as the B, C and cysteine-knot (CK) domains. The D1-D3 domains are located in the N-terminus of the protein, and the D4, B, C and CK domains are located in the C-terminus. The nomoco mutation results in a cysteine to arginine substitution at position 365 (C365R) in the MUC2 protein; amino acid 365 is within an undefined region immediately following the VWF domain.

For more information about Muc2, please see the record for Schlendrian.

Putative Mechanism

MUC2 is the major molecular component of the inner and outer mucosal layers of the intestinal tract (9). Because the intestinal mucosal layer plays a critical role in immune defense and protection against commensal bacteria, the disruption of the intestinal mucus layer and epithelium can result in intestinal bowel disease (IBD; OMIM: #266600). MUC2 polymorphisms are associated with the development of CD (10). Moreover, IBD patients often show alterations in mucin composition, including decreases in MUC2 expression, abnormal expression of other mucins, as well as differences in the glycosylation and sulfation of MUC2 (10-13). The excessive inflammation that occurs in IBD may lead to upregulation of MUC2 as MUC2 is regulated in vitro by a variety of inflammatory mediators including NF-κB, an important effector of the immune system (10;14). Targeted inactivation of the murine Muc2 gene resulted in the spontaneous development of intestinal tumors (15), and on certain genetic backgrounds, the development of chronic inflammation and colitis (16)Muc2-/- mice lacked morphologically distinguishable goblet cells, although the goblet cell lineage remained intact. Muc2 knockout mice also exhibited increased proliferation, decreased apoptosis and increased migration of epithelial cells along the intestinal crypts. Two missense mutations in murine Muc2 also resulted in the development of ulcerative colitis in mice, although the mechanism behind the development of disease in these animals appears to be somewhat different than complete lack of the protective mucus layer (17). In mice with missense mutations in Muc2, the development of colitis resulted from the aberrant oligomerization and subsequent accumulation of MUC2 in the ER, which lead to ER stress, triggering of the unfolded protein response (UPR), subsequent inflammation and goblet cell apoptosis (17). We have not demonstrated that the nomoco mutation results in MUC2 accumulation in the ER. It is also possible that secretion of MUC2 in nomoco animals will result in alterations of the intestinal mucus layer. Changes in mucus formation may result in inability to bind to molecules such as trefoil factors or sequester important bioactive molecules. 

Primers PCR Primer
nomoco_pcr_F: CTGTCCATGTATGCGTTGCC
nomoco_pcr_R: CTTTGAGGAACTGAGTCCCAAC

Sequencing Primer
nomoco_seq_F: CCATGTATGCGTTGCCAGGAG
nomoco_seq_R: TGAGTCCCAACATGCAGCTG
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 400 nucleotides is amplified (chromosome 7, + strand):


1   ctgtccatgt atgcgttgcc aggagcaggg aaccttaatg caccagctac ctggtttcca
61  gggggcttgt tccacaccct cctgtaggac agcccttatg catcccatga caagaagggg
121 cactcacttg ctgggggtgc tctggactcc tttcttactc tcctttctct tcagggctcc
181 atcacataca tgcctaatgg ctgctgtaag acatgtgagt acggctgggg aaatgttggg
241 tgatgggtga gttttactca gagccttgag gggaagatgc aggacaggat gcttcaggac
301 tgtgtctcct acactcatgc agatgtatag gaatctgagt ctccttccag ggagaggctg
361 ggtggagccc agctgcatgt tgggactcag ttcctcaaag 


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
AuthorsWilliam McAlpine, Braden Hayse, Kuan-Wen Wang, Emre Turer, and Bruce Beutler