Phenotypic Mutation 'joplin'

• Allele: joplin
• Mutation Type: missense
• Chromosome: 17
• Coordinate: 34,412,232 bp (GRCm39)
• Base Change: T ⇒ A (forward strand)
• Gene: Tap1
• Gene Name: transporter 1, ATP-binding cassette, sub-family B (MDR/TAP)
• Synonym(s): TAP, Ham1, RING4, MTP1, Tap-1, Ham-1, Abcb2, PSF-1
• Chromosomal Location: 34,406,530-34,416,199 bp (+) (GRCm39)
• MGI Phenotype: FUNCTION: The membrane-associated protein encoded by this gene is a member of the superfamily of ATP-binding cassette (ABC) transporters. ABC proteins transport various molecules across extra- and intra-cellular membranes. ABC genes are divided into seven distinct subfamilies (ABC1, MDR/TAP, MRP, ALD, OABP, GCN20, White). This protein is a member of the MDR/TAP subfamily. Members of the MDR/TAP subfamily are involved in multidrug resistance. This protein forms a heterodimer with Tap2 that transports short peptides from the cytosol into the endoplasmic reticulum lumen. Mutations in the human gene may be associated with ankylosing spondylitis, insulin-dependent diabetes mellitus, and celiac disease. Two transcript variants encoding different isoforms have been found for this gene. [provided by RefSeq, Jun 2009] ; PHENOTYPE: Mice homozygous for targeted mutations that inactivate the gene are deficient in antigen presentation, surface class I antigens, and CD4-8+ T cells. [provided by MGI curators]
• Mapped: Yes
• Amino Acid Change: Valine changed to Glutamic Acid
• Institutional Source: Beutler Lab
• Gene Model: predicted gene model for protein(s): [ENSMUSP00000039264] [ENSMUSP00000128401]
• AlphaFold: P21958
• SMART Domains: Protein: ENSMUSP00000039264; Gene: ENSMUSG00000037321; AA Change: V451E

Domain	Start	End	E-Value	Type
transmembrane domain	5	27	N/A	INTRINSIC
transmembrane domain	37	59	N/A	INTRINSIC
transmembrane domain	66	88	N/A	INTRINSIC
transmembrane domain	116	138	N/A	INTRINSIC
Pfam:ABC_membrane	163	420	9.1e-55	PFAM
AAA	478	666	2.21e-18	SMART

• Predicted Effect: probably damaging ; PolyPhen 2 Score 1.000 (Sensitivity: 0.00; Specificity: 1.00)
  (Using ENSMUST00000041633)
• SMART Domains: Protein: ENSMUSP00000128401; Gene: ENSMUSG00000037321; AA Change: V479E

Domain	Start	End	E-Value	Type
transmembrane domain	5	27	N/A	INTRINSIC
transmembrane domain	37	59	N/A	INTRINSIC
transmembrane domain	66	88	N/A	INTRINSIC
transmembrane domain	116	138	N/A	INTRINSIC
Pfam:ABC_membrane	163	434	5.8e-70	PFAM
AAA	506	694	2.21e-18	SMART

• Predicted Effect: probably damaging ; PolyPhen 2 Score 1.000 (Sensitivity: 0.00; Specificity: 1.00)
  (Using ENSMUST00000170086)
• SMART Domains: Protein: ENSMUSP00000130189; Gene: ENSMUSG00000037321; AA Change: V148E

Domain	Start	End	E-Value	Type
Pfam:ABC_membrane	1	114	1.5e-24	PFAM
Pfam:ABC_tran	167	196	1e-7	PFAM

• Predicted Effect: unknown
• Meta Mutation Damage Score: 0.6467
• Is this an essential gene?: Non Essential (E-score: 0.000)
• Phenotypic Category: Autosomal Recessive
• Mode of Inheritance: Autosomal Recessive
• Last Updated: 2019-09-04 9:39 PM by Anne Murray
• Record Created: 2017-08-28 1:47 PM by Bruce Beutler
• Record Posted: 2017-09-15
• Science Writers: Anne Murray
• Illustrators: Diantha La Vine
• Authors: Xue Zhong, Jin Huk Choi, and Bruce Beutler

Accession Number

NCBI RefSeq: NM_013683, NM_001161730; MGI: 98483

Alleles Listed at MGI

All mutations/alleles(6) : Chemically induced (ENU)(1) Targeted(5)

Phenotypic Description

Figure 1. Joplin mice exhibit increased CD4 to CD8 T cell ratios. Flow cytometric analysis of peripheral blood was utilized to determine T cell frequency. Raw 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.

Figure 2. Joplin mice exhibit decreased frequencies of peripheral CD8+ T cells. Flow cytometric analysis of peripheral blood was utilized to determine T cell frequency. Raw 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.

Figure 3. Joplin mice exhibit decreased frequencies of peripheral CD8+ T cells in CD3+ T cells. Flow cytometric analysis of peripheral blood was utilized to determine T cell frequency. Raw 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.

Figure 4. Joplin mice exhibit increased frequencies of peripheral CD4+ T cells in CD3+ T cells. Flow cytometric analysis of peripheral blood was utilized to determine T cell frequency. Raw 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 joplin phenotype was identified among N-ethyl-N-nitrosourea (ENU)-mutagenized G3 mice of the pedigree R3883, some of which exhibited an increase in the CD4 to CD8 T cell ratio (Figure 1) due to reduced frequencies of CD8+ T cells (Figure 2) and CD8+ T cells in CD3+ T cells (Figure 3) with concomitant increased frequencies of CD4+ T cells in CD3+ T cells (Figure 4), all in the peripheral blood.

Nature of Mutation

Figure 5. Linkage mapping of the reduced CD8+ T cells in CD3+ T cells frequency using a recessive model of inheritance. Manhattan plot shows -log10 P values (Y-axis) plotted against the chromosome positions of 50 mutations (X-axis) identified in the G1 male of pedigree R3883. 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 50 mutations. All of the above anomalies were linked by continuous variable mapping to a mutation in Tap1:  a T to A transversion at base pair 34,193,258 (v38) on chromosome 17, or base pair 5,703 in the GenBank genomic region NC_000083. The strongest association was found with a recessive model of inheritance to the reduced CD8+ T cells in CD3+ T cells frequency phenotype, wherein two variant homozygotes departed phenotypically from two homozygous reference mice and seven heterozygous mice with a P value of 2.47 x 10^-8 (Figure 5).  

The mutation corresponds to residue 1,760 in the mRNA sequence NM_013683 (variant 1) within exon 7 of 11 total exons or residue 1,676 in the mRNA sequence NM_001161730 (variant 2) within exon 8 of 12 total exons.

5687 AACATGAAAGGCCTTGTGGAGTTCCAAGATGTC 474  -N--M--K--G--L--V--E--F--Q--D--V- (isoform 1; NP_038711) 446  -N--M--K--G--L--V--E--F--Q--D--V- (isoform 2; NP_001155202)

Genomic numbering corresponds to NC_000083. The mutated nucleotide is indicated in red. The mutation results in a valine (V) to glutamic acid (E) substitution at position 479 (V479E) in isoform 1 and a V to E substitution at position 451 (V451E) in isoform 2 of the TAP1 protein, and is strongly predicted by PolyPhen-2 to cause loss of function (score = 1.00) (1).

Protein Prediction

Figure 6. Predicted membrane topology of TAP. Both TAP1 and TAP2 contain a 6-helix core transmembrane domain (TMD; purple) and a cytosolic nucleotide binding domain (NBD). Tapasin binding N-terminal accessory domains (pink) consist of 4 and 3 transmembrane helices for TAP1 and TAP2, respectively. The Walker A (A), Walker B (B), and signature/C-loop motif (C) sequences, involved in ATP binding and/or hydrolysis, and color coded to indicate to which ATPase site they belong (green = consensus site,; orange = degenerate site). The joplin mutation is indicated. The image is interactive, click to view other Tap1 mutations.

Transporter associated with antigen processing (TAP) is a member of the ATP-binding cassette (ABC) transporter family, ubiquitous proteins that shuttle a variety of substrates, including ions (see record for mayday), sugars, amino acids, peptides, vitamins, lipids, antibiotics, and drugs, across cellular membranes (2;3). All ABC transporters possess a modular architecture, the core of which consists of two hydrophobic transmembrane domains (TMDs) and two cytosolic nucleotide-binding domains (NBDs; also called ABCs) (4). TAP is a heterodimer of the homologous TAP1 and TAP2 (see the record for ganymede) proteins, each of which contains a six-helix TMD and a C-terminal NBD (Figure 6) (5;6). TAP1 and TAP2 also contain N-terminal accessory domains, non-essential for peptide transport, that bind to tapasin, the specialized chaperone that bridges TAP and class I MHC molecules in the peptide loading complex (6;7). The accessory domain of TAP1 consists of four transmembrane helices (5;6).

The joplin mutation occurs in cytoplasmic tail of Tap1.

Please see the record rose for information about Tap1.

Putative Mechanism

The TAP complex pumps cytosolic peptides into the endoplasmic reticulum (ER) lumen for loading onto class I major histocompatibility (MHC) molecules and presentation to T lymphocytes. Peptide binding is required to stabilize MHC class I molecules, so mice with disrupted TAP1 or TAP2 genes assemble drastically reduced amounts of MHC class I molecules, and have nearly absent surface expression of MHC class I. Mutations in TAP1 (8;9) cause the rarely occurring bare lymphocyte syndrome type I (type I BLS, OMIM #604571), characterized a reduction in MHC class I surface expression to 1-3% of normal levels. Patients with type I BLS from TAP1 or TAP2 mutations develop chronic bacterial infections of the sinus and bronchi in late childhood and/or granulomatous skin lesions, but no severe combined immunodeficiency, diarrhea, nor persistent systemic infections (10;11). The phenotypes of the joplin mice mimics that of rose and ganymede, indicating loss of TAP1^joplin (and TAP1/TAP2) function.

Primers

PCR Primer
joplin_pcr_F: AAGCAGGTTAGACGGCTCAC
joplin_pcr_R: TTCCAGGATGCAGGGTGAAC

Sequencing Primer
joplin_seq_F: AGACGGCTCACTTGCTTTCG
joplin_seq_R: TGCAGGGTGAACGTCAGC

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 456 nucleotides is amplified (chromosome 17, + strand):

1   aagcaggtta gacggctcac ttgctttcgt ggtccatccc aggctcacac acaccccgct
61  ctgcaatgca ggtcctgctc tccctctacc cctccatgca gaaggctgtg ggctcctcag
121 agaaaatatt cgaatacttg gaccggactc cttgctctcc actcagtggc tcgttggcac
181 cctcaaacat gaaaggcctt gtggagttcc aagatgtctc ttttgcctac ccaaaccagc
241 ccaaagtcca ggtgcttcag gtactgtcta cccatccctt ttatgcctcc ttcttcttct
301 tcttcttctt cttcttcttc ttcttcttct tcttcttctt cttcttcttc ttcttcttct
361 tcttccttct cctcctctcc ctccttctcc tcctcctcct ccttgccacg agcttctctg
421 gcctccaggg gctgacgttc accctgcatc ctggaa

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

References

  1. Adzhubei, I. A., Schmidt, S., Peshkin, L., Ramensky, V. E., Gerasimova, A., Bork, P., Kondrashov, A. S., and Sunyaev, S. R. (2010) A Method and Server for Predicting Damaging Missense Mutations. Nat Methods. 7, 248-249.

  2. Borst, P., and Elferink, R. O. (2002) Mammalian ABC Transporters in Health and Disease. Annu Rev Biochem. 71, 537-592.

  3. Higgins, C. F. (1992) ABC Transporters: From Microorganisms to Man. Annu Rev Cell Biol. 8, 67-113.

  4. Biemans-Oldehinkel, E., Doeven, M. K., and Poolman, B. (2006) ABC Transporter Architecture and Regulatory Roles of Accessory Domains. FEBS Lett. 580, 1023-1035.

  5. Schrodt, S., Koch, J., and Tampe, R. (2006) Membrane Topology of the Transporter Associated with Antigen Processing (TAP1) within an Assembled Functional Peptide-Loading Complex. J Biol Chem. 281, 6455-6462.

  6. Koch, J., Guntrum, R., Heintke, S., Kyritsis, C., and Tampe, R. (2004) Functional Dissection of the Transmembrane Domains of the Transporter Associated with Antigen Processing (TAP). J Biol Chem. 279, 10142-10147.

  7. Procko, E., Raghuraman, G., Wiley, D. C., Raghavan, M., and Gaudet, R. (2005) Identification of Domain Boundaries within the N-Termini of TAP1 and TAP2 and their Importance in Tapasin Binding and Tapasin-Mediated Increase in Peptide Loading of MHC Class I. Immunol Cell Biol. 83, 475-482.

8. de la Salle, H., Zimmer, J., Fricker, D., Angenieux, C., Cazenave, J. P., Okubo, M., Maeda, H., Plebani, A., Tongio, M. M., Dormoy, A., and Hanau, D. (1999) HLA Class I Deficiencies due to Mutations in Subunit 1 of the Peptide Transporter TAP1. J. Clin. Invest. 103, R9-R13.

9. Furukawa, H., Murata, S., Yabe, T., Shimbara, N., Keicho, N., Kashiwase, K., Watanabe, K., Ishikawa, Y., Akaza, T., Tadokoro, K., Tohma, S., Inoue, T., Tokunaga, K., Yamamoto, K., Tanaka, K., and Juji, T. (1999) Splice Acceptor Site Mutation of the Transporter Associated with Antigen Processing-1 Gene in Human Bare Lymphocyte Syndrome. J. Clin. Invest. 103, 755-758.

10. Zimmer, J., Andres, E., Donato, L., Hanau, D., Hentges, F., and de la Salle, H. (2005) Clinical and Immunological Aspects of HLA Class I Deficiency. QJM. 98, 719-727.

11. Lankat-Buttgereit, B., and Tampe, R. (2002) The Transporter Associated with Antigen Processing: Function and Implications in Human Diseases. Physiol. Rev. 82, 187-204.