|Mutation Type||frame shift|
|Coordinate||68,697,987 bp (GRCm38)|
|Base Change||TCAC ⇒ TC (forward strand)|
|Gene Name||interleukin 12a|
|Chromosomal Location||68,690,644-68,698,547 bp (+)|
FUNCTION: [Summary is not available for the mouse gene. This summary is for the human ortholog.] This gene encodes a subunit of a cytokine that acts on T and natural killer cells, and has a broad array of biological activities. The cytokine is a disulfide-linked heterodimer composed of the 35-kD subunit encoded by this gene, and a 40-kD subunit that is a member of the cytokine receptor family. This cytokine is required for the T-cell-independent induction of interferon (IFN)-gamma, and is important for the differentiation of both Th1 and Th2 cells. The responses of lymphocytes to this cytokine are mediated by the activator of transcription protein STAT4. Nitric oxide synthase 2A (NOS2A/NOS2) is found to be required for the signaling process of this cytokine in innate immunity. [provided by RefSeq, Jul 2008]
PHENOTYPE: Null homozygotes have decreased NK cell responses, altered effector T cell differentiation, and increased susceptibility to parasitic infections. [provided by MGI curators]
|Amino Acid Change|
|Institutional Source||Beutler Lab|
|Gene Model||predicted gene model for protein(s): [ENSMUSP00000029345] [ENSMUSP00000103446]|
|Predicted Effect||probably null|
|Predicted Effect||probably null|
|Alleles Listed at MGI|
|Mode of Inheritance||Autosomal Recessive|
|Last Updated||2018-12-20 12:52 PM by Anne Murray|
|Record Created||2015-12-10 1:43 PM by Bruce Beutler|
The bakers_dozen phenotype was identified among G3 mice of the pedigree R2925, some of which showed increased mouse cytomegalovirus (MCMV) proliferation in macrophages of the spleen after MCMV infection, indicating increased MCMV susceptibility (Figure 1).
|Nature of Mutation|
Whole exome HiSeq sequencing of the G1 grandsire identified 48 mutations. The increased susceptibility to MCMV infection phenotype was linked by continuous variable mapping to a mutation in Il12a: a TCAC>TC substitution at base pair 68,697,987 (v38) on chromosome 3, or base pair 7,344 in the GenBank genomic region NC_000069 within exon 8 out of 8 total exons. Linkage was found with a recessive model of inheritance (P = 1.28 x 10-4), wherein seven variant homozygotes departed phenotypically from 14 homozygous reference mice and 21 heterozygous mice (Figure 2).
The effect of the mutation at the cDNA and protein level have not examined, but the mutation is predicted to result in a frame-shift in the encoded protein. The frame-shift is predicted to result in coding of 34 aberrant amino acids after amino acid 216, and is predicted to result in loss of the original stop codon after amino acid 236. The frame-shift results in coding of a stop codon 45-base pairs within the 3’-untranslated region (3’-UTR) of exon 8.
Genomic numbering corresponds to NC_000069. The mutated nucleotides are indicated in red. The highlighted portion indicates nucleotides with the 3’-UTR.
Il12a encodes interleukin 12a (IL-12a; alternatively IL-12p35 or p35), a subunit of the cytokines IL-12 and IL-35. IL-12 and IL-35 are members of the IL-12 family of cytokines, which also includes IL-23 and IL-27. Each member of the IL-12 family forms a heterodimer containing an α-subunit (i.e., IL-12p35, IL-23p19 and IL-27p28) and a β-subunit (i.e., IL-12p40 and Ebi3); both subunits are required for secretion of an active cytokine (1). The IL-12p40 β-subunit can pair with both IL-12p35 and IL-23p19 to form IL-12 and IL-23, respectively, while Ebi3 can dimerize with IL-27p28 or IL-12p35 to form IL-27 or IL-35, respectively.
Il12a has alternative transcriptional start sites that produce p35 transcripts with variable translational efficiencies (4). The p35 mRNA is constitutively synthesized in unstimulated cells, but little protein is secreted due to an inhibitory ATG in the 5’-UTR. After LPS stimulation, the transcription start site changes so the inhibitory region is excluded, and translation occurs (4).
The bakers_dozen mutation is predicted to result in a frame-shift in the encoded protein. The frame-shift is predicted to result in coding of 34 aberrant amino acids after amino acid 216, and is predicted to result in loss of the original stop codon after amino acid 236 and coding of a new stop codon 45-base pairs within the 3’-UTR of exon 8.
p35 is expressed ubiquitously at low levels (4;5). IL-12 is secreted primarily by monocytes, dendritic cells, B cells, and macrophages in response to pathogens (6;7). IL-12 (primarily IL-12p40) is positively regulated by IFNγ (8) and negatively regulated by IL-4, IL-10, IL-11, IL-13, and type 1 IFNs (9). IL-12 expression in monocytes/macrophages and dendritic cells is also induced by TLR2, -4, -5, and -9 agonists (10). IL-35 is exclusively produced by regulatory T cells (11;12).
IL-12 cytokines mediate their functions by the Janus activating kinase (JAK)-signal transducer and activator of transcription (STAT) signaling pathway. Only the p35-associated cytokines (IL-12 and IL-35) will be discussed further. IL-12 binds the IL-12 receptor (IL-12Rβ1/IL-12Rβ2) on activated T cells, natural killer cells and dendritic cells, subsequently signaling through the JAK2/tyrosine kinase 2 (TYK2)-STAT4 signaling pathway [Figure 4; reviewed by (2)]. IL-35 binds a receptor that is composed of a IL-12Rβ2/gp130 heterodimer, and signals through both JAK2/STAT4 and JAK1/STAT1 (13). IL-35 can also bind receptors that contain gp130-gp130 and IL12Rβ2-lL12Rβ2 homodimers to signal via JAK1/STAT1 and JAK2/STAT4, respectively.
The canonical JAK-STAT signaling pathway begins with the binding of one or more cytokines to their cognate cell-surface receptors. These receptors are associated with JAK tyrosine kinases (see the record mount_tai for information about JAK3), which are normally dephosphorylated and inactive. Receptor stimulation results in dimerization/oligomerization and subsequent apposition of JAK proteins, which are now capable of trans-phosphorylation as they are brought in close proximity. This activates JAKs to phosphorylate the receptor cytoplasmic domains, creating phosphotyrosine ligands for the SH2 domains of STAT proteins (see the record domino for information about STAT1 and numb for information about STAT2). Once recruited to the receptor, STAT proteins are also tyrosine phosphorylated by JAKs, a phosphorylation event which occurs on a single tyrosine residue that is found at around residue 700 of all STATs. Tyrosine phosphorylation of STATs may allow formation and/or conformational reorganization of the activated STAT dimer, involving reciprocal SH2 domain-phosphotyrosine interactions between STAT monomers. Phosphorylated, activated STATs enter the nucleus and accumulate there to promote transcription. For more information about JAK-STAT signaling, see the record domino.
The IL-12 family members prime naïve CD4 T cells, promote the differentiation of CD4 T cells to cytokine-producing T-helper subsets and memory T cells, and promote the activation of pro-inflammatory responses that subsequently protect against infection and prevent autoimmune diseases (12;14-16). IL-12 induces T-bet and promotes naïve CD4+ T cell differentiation into Th1 cells, while inhibiting IL-4 production and antagonizing Th2 responses [reviewed in (3)]. IL-12 also induces IFNg production by natural killer cells and innate lymphoid cells. IL-12 is required for bacterial and intracellular parasite infection resistance (17). CD8 T cells use IL-12, T-cell receptor, and/or type I interferon signals to promote differentiation and effector functions during an infection to aid in pathogen clearance and the generation of pathogen-specific memory T cells (18).
IL-35 suppresses T cell proliferation and Th17 cell development by inducing cell cycle arrest in G1 (19-21). IL-35 blocks Th2 development by repressing GATA3 and IL-4 expression and limiting Th2 proliferation [reviewed in (3)]. IL-35 also induces the development in the periphery of an induced regulatory T cell (iTreg) population from CD4+Foxp3- conventional T cells (19-21). iTreg cells, together with “natural” Tregs, are required for immune tolerance. Il12a-deficient T cells showed reduced regulatory activity as well as a failure to control homeostatic proliferation (20).
IL12A mutations are associated with cases of primary biliary cirrhosis (22), celiac disease (23;24), multiple sclerosis (25), Graves’ disease (26), and asthma (27). Patients with autoimmune diseases (e.g., rheumatoid arthritis, psoriasis, and inflammatory bowel disease) show increased levels of IL-12 in affected tissues (28-30). IL-12 promotes inflammation and the development of chronic inflammatory diseases, while IL-35 suppresses inflammation and mitigate autoimmune diseases.
Il12a-deficient (Il12a-/-) mice lack both IL-12 and IL-35. Il12a-/- mice exhibited reduced numbers of natural killer cells producing IFNγ after MCMV, Leishmania major, or Leishmania infantum infections or in response to the TLR9 agonist CpG (31-34). After MCMV or LCMV infections, the levels of IFNγ was lower than that in controls (31;35). The levels of circulating IL-12 levels after LCMV infection or LPS injection were absent (35). Il12a-/- mice exhibited increased sensitivity to N-methyl-N-nitrosourea (MNU) compared to similarly treated wild-type controls (36) as well as increased incidence of tumors by chemical induction (37). Il12a-/- mice showed increased susceptibility to induced rheumatoid arthritis (38).
The phenotype of the bakers_dozen mice mimics that of Il12a-/- mice, indicating loss of p35-associated function.
bakers_dozen(F):5'- GGACTTTGCATTGACTGTCTCC -3'
bakers_dozen(R):5'- AGGTAGCTGTGCCACCTTTG -3'
bakers_dozen_seq(F):5'- GACTGTCTCCCATTTTGCAGACAAAC -3'
bakers_dozen_seq(R):5'- CCACCTTTGGGGAGATGAG -3'
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14. Trinchieri, G., Pflanz, S., and Kastelein, R. A. (2003) The IL-12 Family of Heterodimeric Cytokines: New Players in the Regulation of T Cell Responses. Immunity. 19, 641-644.
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19. Collison, L. W., Chaturvedi, V., Henderson, A. L., Giacomin, P. R., Guy, C., Bankoti, J., Finkelstein, D., Forbes, K., Workman, C. J., Brown, S. A., Rehg, J. E., Jones, M. L., Ni, H. T., Artis, D., Turk, M. J., and Vignali, D. A. (2010) IL-35-Mediated Induction of a Potent Regulatory T Cell Population. Nat Immunol. 11, 1093-1101.
20. Collison, L. W., Workman, C. J., Kuo, T. T., Boyd, K., Wang, Y., Vignali, K. M., Cross, R., Sehy, D., Blumberg, R. S., and Vignali, D. A. (2007) The Inhibitory Cytokine IL-35 Contributes to Regulatory T-Cell Function. Nature. 450, 566-569.
21. Niedbala, W., Wei, X. Q., Cai, B., Hueber, A. J., Leung, B. P., McInnes, I. B., and Liew, F. Y. (2007) IL-35 is a Novel Cytokine with Therapeutic Effects Against Collagen-Induced Arthritis through the Expansion of Regulatory T Cells and Suppression of Th17 Cells. Eur J Immunol. 37, 3021-3029.
22. Hirschfield, G. M., Liu, X., Xu, C., Lu, Y., Xie, G., Lu, Y., Gu, X., Walker, E. J., Jing, K., Juran, B. D., Mason, A. L., Myers, R. P., Peltekian, K. M., Ghent, C. N., Coltescu, C., Atkinson, E. J., Heathcote, E. J., Lazaridis, K. N., Amos, C. I., and Siminovitch, K. A. (2009) Primary Biliary Cirrhosis Associated with HLA, IL12A, and IL12RB2 Variants. N Engl J Med. 360, 2544-2555.
23. Dubois, P. C., Trynka, G., Franke, L., Hunt, K. A., Romanos, J., Curtotti, A., Zhernakova, A., Heap, G. A., Adany, R., Aromaa, A., Bardella, M. T., van den Berg, L. H., Bockett, N. A., de la Concha, E. G., Dema, B., Fehrmann, R. S., Fernandez-Arquero, M., Fiatal, S., Grandone, E., Green, P. M., Groen, H. J., Gwilliam, R., Houwen, R. H., Hunt, S. E., Kaukinen, K., Kelleher, D., Korponay-Szabo, I., Kurppa, K., MacMathuna, P., Maki, M., Mazzilli, M. C., McCann, O. T., Mearin, M. L., Mein, C. A., Mirza, M. M., Mistry, V., Mora, B., Morley, K. I., Mulder, C. J., Murray, J. A., Nunez, C., Oosterom, E., Ophoff, R. A., Polanco, I., Peltonen, L., Platteel, M., Rybak, A., Salomaa, V., Schweizer, J. J., Sperandeo, M. P., Tack, G. J., Turner, G., Veldink, J. H., Verbeek, W. H., Weersma, R. K., Wolters, V. M., Urcelay, E., Cukrowska, B., Greco, L., Neuhausen, S. L., McManus, R., Barisani, D., Deloukas, P., Barrett, J. C., Saavalainen, P., Wijmenga, C., and van Heel, D. A. (2010) Multiple Common Variants for Celiac Disease Influencing Immune Gene Expression. Nat Genet. 42, 295-302.
24. Hunt, K. A., Zhernakova, A., Turner, G., Heap, G. A., Franke, L., Bruinenberg, M., Romanos, J., Dinesen, L. C., Ryan, A. W., Panesar, D., Gwilliam, R., Takeuchi, F., McLaren, W. M., Holmes, G. K., Howdle, P. D., Walters, J. R., Sanders, D. S., Playford, R. J., Trynka, G., Mulder, C. J., Mearin, M. L., Verbeek, W. H., Trimble, V., Stevens, F. M., O'Morain, C., Kennedy, N. P., Kelleher, D., Pennington, D. J., Strachan, D. P., McArdle, W. L., Mein, C. A., Wapenaar, M. C., Deloukas, P., McGinnis, R., McManus, R., Wijmenga, C., and van Heel, D. A. (2008) Newly Identified Genetic Risk Variants for Celiac Disease Related to the Immune Response. Nat Genet. 40, 395-402.
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26. Guo, T., Yang, S., Liu, N., Wang, S., Cui, B., and Ning, G. (2011) Association Study of Interleukin-12A Gene Polymorphisms with Graves' Disease in Two Chinese Populations. Clin Endocrinol (Oxf). 74, 125-129.
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30. Yawalkar, N., Tscharner, G. G., Hunger, R. E., and Hassan, A. S. (2009) Increased Expression of IL-12p70 and IL-23 by Multiple Dendritic Cell and Macrophage Subsets in Plaque Psoriasis. J Dermatol Sci. 54, 99-105.
31. Nguyen, K. B., Salazar-Mather, T. P., Dalod, M. Y., Van Deusen, J. B., Wei, X. Q., Liew, F. Y., Caligiuri, M. A., Durbin, J. E., and Biron, C. A. (2002) Coordinated and Distinct Roles for IFN-Alpha Beta, IL-12, and IL-15 Regulation of NK Cell Responses to Viral Infection. J Immunol. 169, 4279-4287.
32. Schleicher, U., Liese, J., Knippertz, I., Kurzmann, C., Hesse, A., Heit, A., Fischer, J. A., Weiss, S., Kalinke, U., Kunz, S., and Bogdan, C. (2007) NK Cell Activation in Visceral Leishmaniasis Requires TLR9, Myeloid DCs, and IL-12, but is Independent of Plasmacytoid DCs. J Exp Med. 204, 893-906.
33. Hou, B., Reizis, B., and DeFranco, A. L. (2008) Toll-Like Receptors Activate Innate and Adaptive Immunity by using Dendritic Cell-Intrinsic and -Extrinsic Mechanisms. Immunity. 29, 272-282.
34. Mattner, F., Magram, J., Ferrante, J., Launois, P., Di Padova, K., Behin, R., Gately, M. K., Louis, J. A., and Alber, G. (1996) Genetically Resistant Mice Lacking Interleukin-12 are Susceptible to Infection with Leishmania Major and Mount a Polarized Th2 Cell Response. Eur J Immunol. 26, 1553-1559.
35. Cousens, L. P., Peterson, R., Hsu, S., Dorner, A., Altman, J. D., Ahmed, R., and Biron, C. A. (1999) Two Roads Diverged: Interferon alpha/beta- and Interleukin 12-Mediated Pathways in Promoting T Cell Interferon Gamma Responses during Viral Infection. J Exp Med. 189, 1315-1328.
36. Liu, J., Xiang, Z., and Ma, X. (2004) Role of IFN Regulatory Factor-1 and IL-12 in Immunological Resistance to Pathogenesis of N-Methyl-N-Nitrosourea-Induced T Lymphoma. J Immunol. 173, 1184-1193.
37. Langowski, J. L., Zhang, X., Wu, L., Mattson, J. D., Chen, T., Smith, K., Basham, B., McClanahan, T., Kastelein, R. A., and Oft, M. (2006) IL-23 Promotes Tumour Incidence and Growth. Nature. 442, 461-465.
|Science Writers||Anne Murray|
|Illustrators||Diantha La Vine|
|Authors||Duanwu Zhang, Tao Yue, and Bruce Beutler|