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. 1999 Mar 1;18(5):1280–1291. doi: 10.1093/emboj/18.5.1280

Targeted disruption of SMAD3 results in impaired mucosal immunity and diminished T cell responsiveness to TGF-beta.

X Yang 1, J J Letterio 1, R J Lechleider 1, L Chen 1, R Hayman 1, H Gu 1, A B Roberts 1, C Deng 1
PMCID: PMC1171218  PMID: 10064594

Abstract

SMAD3 is one of the intracellular mediators that transduces signals from transforming growth factor-beta (TGF-beta) and activin receptors. We show that SMAD3 mutant mice generated by gene targeting die between 1 and 8 months due to a primary defect in immune function. Symptomatic mice exhibit thymic involution, enlarged lymph nodes, and formation of bacterial abscesses adjacent to mucosal surfaces. Mutant T cells exhibit an activated phenotype in vivo, and are not inhibited by TGF-beta1 in vitro. Mutant neutrophils are also impaired in their chemotactic response toward TGF-beta. Chronic intestinal inflammation is infrequently associated with colonic adenocarcinoma in mice older than 6 months of age. These data suggest that SMAD3 has an important role in TGF-beta-mediated regulation of T cell activation and mucosal immunity, and that the loss of these functions is responsible for chronic infection and the lethality of Smad3-null mice.

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Selected References

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  1. Abdollah S., Macías-Silva M., Tsukazaki T., Hayashi H., Attisano L., Wrana J. L. TbetaRI phosphorylation of Smad2 on Ser465 and Ser467 is required for Smad2-Smad4 complex formation and signaling. J Biol Chem. 1997 Oct 31;272(44):27678–27685. doi: 10.1074/jbc.272.44.27678. [DOI] [PubMed] [Google Scholar]
  2. Arai T., Akiyama Y., Okabe S., Ando M., Endo M., Yuasa Y. Genomic structure of the human Smad3 gene and its infrequent alterations in colorectal cancers. Cancer Lett. 1998 Jan 9;122(1-2):157–163. doi: 10.1016/s0304-3835(97)00384-4. [DOI] [PubMed] [Google Scholar]
  3. Baker J. C., Harland R. M. A novel mesoderm inducer, Madr2, functions in the activin signal transduction pathway. Genes Dev. 1996 Aug 1;10(15):1880–1889. doi: 10.1101/gad.10.15.1880. [DOI] [PubMed] [Google Scholar]
  4. Brandes M. E., Mai U. E., Ohura K., Wahl S. M. Type I transforming growth factor-beta receptors on neutrophils mediate chemotaxis to transforming growth factor-beta. J Immunol. 1991 Sep 1;147(5):1600–1606. [PubMed] [Google Scholar]
  5. Chen X., Rubock M. J., Whitman M. A transcriptional partner for MAD proteins in TGF-beta signalling. Nature. 1996 Oct 24;383(6602):691–696. doi: 10.1038/383691a0. [DOI] [PubMed] [Google Scholar]
  6. Chen X., Weisberg E., Fridmacher V., Watanabe M., Naco G., Whitman M. Smad4 and FAST-1 in the assembly of activin-responsive factor. Nature. 1997 Sep 4;389(6646):85–89. doi: 10.1038/38008. [DOI] [PubMed] [Google Scholar]
  7. Chen Y., Inobe J., Marks R., Gonnella P., Kuchroo V. K., Weiner H. L. Peripheral deletion of antigen-reactive T cells in oral tolerance. Nature. 1995 Jul 13;376(6536):177–180. doi: 10.1038/376177a0. [DOI] [PubMed] [Google Scholar]
  8. Coffman R. L., Lebman D. A., Shrader B. Transforming growth factor beta specifically enhances IgA production by lipopolysaccharide-stimulated murine B lymphocytes. J Exp Med. 1989 Sep 1;170(3):1039–1044. doi: 10.1084/jem.170.3.1039. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Deng C. X., Wynshaw-Boris A., Shen M. M., Daugherty C., Ornitz D. M., Leder P. Murine FGFR-1 is required for early postimplantation growth and axial organization. Genes Dev. 1994 Dec 15;8(24):3045–3057. doi: 10.1101/gad.8.24.3045. [DOI] [PubMed] [Google Scholar]
  10. Deng C., Wynshaw-Boris A., Zhou F., Kuo A., Leder P. Fibroblast growth factor receptor 3 is a negative regulator of bone growth. Cell. 1996 Mar 22;84(6):911–921. doi: 10.1016/s0092-8674(00)81069-7. [DOI] [PubMed] [Google Scholar]
  11. Dennler S., Itoh S., Vivien D., ten Dijke P., Huet S., Gauthier J. M. Direct binding of Smad3 and Smad4 to critical TGF beta-inducible elements in the promoter of human plasminogen activator inhibitor-type 1 gene. EMBO J. 1998 Jun 1;17(11):3091–3100. doi: 10.1093/emboj/17.11.3091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Derynck R., Feng X. H. TGF-beta receptor signaling. Biochim Biophys Acta. 1997 Oct 24;1333(2):F105–F150. doi: 10.1016/s0304-419x(97)00017-6. [DOI] [PubMed] [Google Scholar]
  13. Eppert K., Scherer S. W., Ozcelik H., Pirone R., Hoodless P., Kim H., Tsui L. C., Bapat B., Gallinger S., Andrulis I. L. MADR2 maps to 18q21 and encodes a TGFbeta-regulated MAD-related protein that is functionally mutated in colorectal carcinoma. Cell. 1996 Aug 23;86(4):543–552. doi: 10.1016/s0092-8674(00)80128-2. [DOI] [PubMed] [Google Scholar]
  14. Fukaura H., Kent S. C., Pietrusewicz M. J., Khoury S. J., Weiner H. L., Hafler D. A. Antigen-specific TGF-beta1 secretion with bovine myelin oral tolerization in multiple sclerosis. Ann N Y Acad Sci. 1996 Feb 13;778:251–257. doi: 10.1111/j.1749-6632.1996.tb21133.x. [DOI] [PubMed] [Google Scholar]
  15. Hayashi H., Abdollah S., Qiu Y., Cai J., Xu Y. Y., Grinnell B. W., Richardson M. A., Topper J. N., Gimbrone M. A., Jr, Wrana J. L. The MAD-related protein Smad7 associates with the TGFbeta receptor and functions as an antagonist of TGFbeta signaling. Cell. 1997 Jun 27;89(7):1165–1173. doi: 10.1016/s0092-8674(00)80303-7. [DOI] [PubMed] [Google Scholar]
  16. Heldin C. H., Miyazono K., ten Dijke P. TGF-beta signalling from cell membrane to nucleus through SMAD proteins. Nature. 1997 Dec 4;390(6659):465–471. doi: 10.1038/37284. [DOI] [PubMed] [Google Scholar]
  17. Horwitz D. A., Gray J. D., Ohtsuka K., Hirokawa M., Takahashi T. The immunoregulatory effects of NK cells: the role of TGF-beta and implications for autoimmunity. Immunol Today. 1997 Nov;18(11):538–542. doi: 10.1016/s0167-5699(97)01149-3. [DOI] [PubMed] [Google Scholar]
  18. Imai S., Okuno M., Moriwaki H., Muto Y., Murakami K., Shudo K., Suzuki Y., Kojima S. 9,13-di-cis-Retinoic acid induces the production of tPA and activation of latent TGF-beta via RAR alpha in a human liver stellate cell line, LI90. FEBS Lett. 1997 Jul 7;411(1):102–106. doi: 10.1016/s0014-5793(97)00673-x. [DOI] [PubMed] [Google Scholar]
  19. Kingsley D. M. The TGF-beta superfamily: new members, new receptors, and new genetic tests of function in different organisms. Genes Dev. 1994 Jan;8(2):133–146. doi: 10.1101/gad.8.2.133. [DOI] [PubMed] [Google Scholar]
  20. Kühn R., Löhler J., Rennick D., Rajewsky K., Müller W. Interleukin-10-deficient mice develop chronic enterocolitis. Cell. 1993 Oct 22;75(2):263–274. doi: 10.1016/0092-8674(93)80068-p. [DOI] [PubMed] [Google Scholar]
  21. Lagna G., Hata A., Hemmati-Brivanlou A., Massagué J. Partnership between DPC4 and SMAD proteins in TGF-beta signalling pathways. Nature. 1996 Oct 31;383(6603):832–836. doi: 10.1038/383832a0. [DOI] [PubMed] [Google Scholar]
  22. Lauffenburger D., Rothman C., Zigmond S. H. Measurement of leukocyte motility and chemotaxis parameters with a linear under-agarose migration assay. J Immunol. 1983 Aug;131(2):940–947. [PubMed] [Google Scholar]
  23. Letterio J. J., Roberts A. B. Regulation of immune responses by TGF-beta. Annu Rev Immunol. 1998;16:137–161. doi: 10.1146/annurev.immunol.16.1.137. [DOI] [PubMed] [Google Scholar]
  24. Liu F., Hata A., Baker J. C., Doody J., Cárcamo J., Harland R. M., Massagué J. A human Mad protein acting as a BMP-regulated transcriptional activator. Nature. 1996 Jun 13;381(6583):620–623. doi: 10.1038/381620a0. [DOI] [PubMed] [Google Scholar]
  25. Liu F., Pouponnot C., Massagué J. Dual role of the Smad4/DPC4 tumor suppressor in TGFbeta-inducible transcriptional complexes. Genes Dev. 1997 Dec 1;11(23):3157–3167. doi: 10.1101/gad.11.23.3157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Liu X., Sun Y., Constantinescu S. N., Karam E., Weinberg R. A., Lodish H. F. Transforming growth factor beta-induced phosphorylation of Smad3 is required for growth inhibition and transcriptional induction in epithelial cells. Proc Natl Acad Sci U S A. 1997 Sep 30;94(20):10669–10674. doi: 10.1073/pnas.94.20.10669. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lo R. S., Chen Y. G., Shi Y., Pavletich N. P., Massagué J. The L3 loop: a structural motif determining specific interactions between SMAD proteins and TGF-beta receptors. EMBO J. 1998 Feb 16;17(4):996–1005. doi: 10.1093/emboj/17.4.996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Macías-Silva M., Abdollah S., Hoodless P. A., Pirone R., Attisano L., Wrana J. L. MADR2 is a substrate of the TGFbeta receptor and its phosphorylation is required for nuclear accumulation and signaling. Cell. 1996 Dec 27;87(7):1215–1224. doi: 10.1016/s0092-8674(00)81817-6. [DOI] [PubMed] [Google Scholar]
  29. Massagué J. TGF-beta signal transduction. Annu Rev Biochem. 1998;67:753–791. doi: 10.1146/annurev.biochem.67.1.753. [DOI] [PubMed] [Google Scholar]
  30. Miller A., Lider O., Roberts A. B., Sporn M. B., Weiner H. L. Suppressor T cells generated by oral tolerization to myelin basic protein suppress both in vitro and in vivo immune responses by the release of transforming growth factor beta after antigen-specific triggering. Proc Natl Acad Sci U S A. 1992 Jan 1;89(1):421–425. doi: 10.1073/pnas.89.1.421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Mizuguchi T., Kosaka M., Saito S. Activin A suppresses proliferation of interleukin-3-responsive granulocyte-macrophage colony-forming progenitors and stimulates proliferation and differentiation of interleukin-3-responsive erythroid burst-forming progenitors in the peripheral blood. Blood. 1993 Jun 1;81(11):2891–2897. [PubMed] [Google Scholar]
  32. Mombaerts P., Mizoguchi E., Grusby M. J., Glimcher L. H., Bhan A. K., Tonegawa S. Spontaneous development of inflammatory bowel disease in T cell receptor mutant mice. Cell. 1993 Oct 22;75(2):274–282. doi: 10.1016/0092-8674(93)80069-q. [DOI] [PubMed] [Google Scholar]
  33. Nakao A., Imamura T., Souchelnytskyi S., Kawabata M., Ishisaki A., Oeda E., Tamaki K., Hanai J., Heldin C. H., Miyazono K. TGF-beta receptor-mediated signalling through Smad2, Smad3 and Smad4. EMBO J. 1997 Sep 1;16(17):5353–5362. doi: 10.1093/emboj/16.17.5353. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Nakao A., Röijer E., Imamura T., Souchelnytskyi S., Stenman G., Heldin C. H., ten Dijke P. Identification of Smad2, a human Mad-related protein in the transforming growth factor beta signaling pathway. J Biol Chem. 1997 Jan 31;272(5):2896–2900. doi: 10.1074/jbc.272.5.2896. [DOI] [PubMed] [Google Scholar]
  35. Newfeld S. J., Chartoff E. H., Graff J. M., Melton D. A., Gelbart W. M. Mothers against dpp encodes a conserved cytoplasmic protein required in DPP/TGF-beta responsive cells. Development. 1996 Jul;122(7):2099–2108. doi: 10.1242/dev.122.7.2099. [DOI] [PubMed] [Google Scholar]
  36. Nomura M., Li E. Smad2 role in mesoderm formation, left-right patterning and craniofacial development. Nature. 1998 Jun 25;393(6687):786–790. doi: 10.1038/31693. [DOI] [PubMed] [Google Scholar]
  37. Page R. C., Sims T. J., Geissler F., Altman L. C., Baab D. A. Defective neutrophil and monocyte motility in patients with early onset periodontitis. Infect Immun. 1985 Jan;47(1):169–175. doi: 10.1128/iai.47.1.169-175.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Parekh T., Saxena B., Reibman J., Cronstein B. N., Gold L. I. Neutrophil chemotaxis in response to TGF-beta isoforms (TGF-beta 1, TGF-beta 2, TGF-beta 3) is mediated by fibronectin. J Immunol. 1994 Mar 1;152(5):2456–2466. [PubMed] [Google Scholar]
  39. Powrie F., Carlino J., Leach M. W., Mauze S., Coffman R. L. A critical role for transforming growth factor-beta but not interleukin 4 in the suppression of T helper type 1-mediated colitis by CD45RB(low) CD4+ T cells. J Exp Med. 1996 Jun 1;183(6):2669–2674. doi: 10.1084/jem.183.6.2669. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Riggins G. J., Thiagalingam S., Rozenblum E., Weinstein C. L., Kern S. E., Hamilton S. R., Willson J. K., Markowitz S. D., Kinzler K. W., Vogelstein B. Mad-related genes in the human. Nat Genet. 1996 Jul;13(3):347–349. doi: 10.1038/ng0796-347. [DOI] [PubMed] [Google Scholar]
  41. Sadlack B., Merz H., Schorle H., Schimpl A., Feller A. C., Horak I. Ulcerative colitis-like disease in mice with a disrupted interleukin-2 gene. Cell. 1993 Oct 22;75(2):253–261. doi: 10.1016/0092-8674(93)80067-o. [DOI] [PubMed] [Google Scholar]
  42. Savage C., Das P., Finelli A. L., Townsend S. R., Sun C. Y., Baird S. E., Padgett R. W. Caenorhabditis elegans genes sma-2, sma-3, and sma-4 define a conserved family of transforming growth factor beta pathway components. Proc Natl Acad Sci U S A. 1996 Jan 23;93(2):790–794. doi: 10.1073/pnas.93.2.790. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Seder R. A., Marth T., Sieve M. C., Strober W., Letterio J. J., Roberts A. B., Kelsall B. Factors involved in the differentiation of TGF-beta-producing cells from naive CD4+ T cells: IL-4 and IFN-gamma have opposing effects, while TGF-beta positively regulates its own production. J Immunol. 1998 Jun 15;160(12):5719–5728. [PubMed] [Google Scholar]
  44. Sirard C., de la Pompa J. L., Elia A., Itie A., Mirtsos C., Cheung A., Hahn S., Wakeham A., Schwartz L., Kern S. E. The tumor suppressor gene Smad4/Dpc4 is required for gastrulation and later for anterior development of the mouse embryo. Genes Dev. 1998 Jan 1;12(1):107–119. doi: 10.1101/gad.12.1.107. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Souchelnytskyi S., Tamaki K., Engström U., Wernstedt C., ten Dijke P., Heldin C. H. Phosphorylation of Ser465 and Ser467 in the C terminus of Smad2 mediates interaction with Smad4 and is required for transforming growth factor-beta signaling. J Biol Chem. 1997 Oct 31;272(44):28107–28115. doi: 10.1074/jbc.272.44.28107. [DOI] [PubMed] [Google Scholar]
  46. Strober W., Kelsall B., Fuss I., Marth T., Ludviksson B., Ehrhardt R., Neurath M. Reciprocal IFN-gamma and TGF-beta responses regulate the occurrence of mucosal inflammation. Immunol Today. 1997 Feb;18(2):61–64. doi: 10.1016/s0167-5699(97)01000-1. [DOI] [PubMed] [Google Scholar]
  47. Tompkins A. B., Hutchinson P., de Kretser D. M., Hedger M. P. Characterization of lymphocytes in the adult rat testis by flow cytometry: effects of activin and transforming growth factor beta on lymphocyte subsets in vitro. Biol Reprod. 1998 Apr;58(4):943–951. doi: 10.1095/biolreprod58.4.943. [DOI] [PubMed] [Google Scholar]
  48. Tonetti M. S. Molecular factors associated with compartmentalization of gingival immune responses and transepithelial neutrophil migration. J Periodontal Res. 1997 Jan;32(1 Pt 2):104–109. doi: 10.1111/j.1600-0765.1997.tb01389.x. [DOI] [PubMed] [Google Scholar]
  49. Vatopoulos A. C., Kalapothaki V., Legakis N. J. Risk factors for nosocomial infections caused by gram-negative bacilli. The Hellenic Antibiotic Resistance Study Group. J Hosp Infect. 1996 Sep;34(1):11–22. doi: 10.1016/s0195-6701(96)90121-8. [DOI] [PubMed] [Google Scholar]
  50. Wahl S. M., Hunt D. A., Wakefield L. M., McCartney-Francis N., Wahl L. M., Roberts A. B., Sporn M. B. Transforming growth factor type beta induces monocyte chemotaxis and growth factor production. Proc Natl Acad Sci U S A. 1987 Aug;84(16):5788–5792. doi: 10.1073/pnas.84.16.5788. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Waldrip W. R., Bikoff E. K., Hoodless P. A., Wrana J. L., Robertson E. J. Smad2 signaling in extraembryonic tissues determines anterior-posterior polarity of the early mouse embryo. Cell. 1998 Mar 20;92(6):797–808. doi: 10.1016/s0092-8674(00)81407-5. [DOI] [PubMed] [Google Scholar]
  52. Weinstein M., Yang X., Li C., Xu X., Gotay J., Deng C. X. Failure of egg cylinder elongation and mesoderm induction in mouse embryos lacking the tumor suppressor smad2. Proc Natl Acad Sci U S A. 1998 Aug 4;95(16):9378–9383. doi: 10.1073/pnas.95.16.9378. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Wiersdorff V., Lecuit T., Cohen S. M., Mlodzik M. Mad acts downstream of Dpp receptors, revealing a differential requirement for dpp signaling in initiation and propagation of morphogenesis in the Drosophila eye. Development. 1996 Jul;122(7):2153–2162. doi: 10.1242/dev.122.7.2153. [DOI] [PubMed] [Google Scholar]
  54. Yang X., Li C., Xu X., Deng C. The tumor suppressor SMAD4/DPC4 is essential for epiblast proliferation and mesoderm induction in mice. Proc Natl Acad Sci U S A. 1998 Mar 31;95(7):3667–3672. doi: 10.1073/pnas.95.7.3667. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Yingling J. M., Datto M. B., Wong C., Frederick J. P., Liberati N. T., Wang X. F. Tumor suppressor Smad4 is a transforming growth factor beta-inducible DNA binding protein. Mol Cell Biol. 1997 Dec;17(12):7019–7028. doi: 10.1128/mcb.17.12.7019. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Zawel L., Dai J. L., Buckhaults P., Zhou S., Kinzler K. W., Vogelstein B., Kern S. E. Human Smad3 and Smad4 are sequence-specific transcription activators. Mol Cell. 1998 Mar;1(4):611–617. doi: 10.1016/s1097-2765(00)80061-1. [DOI] [PubMed] [Google Scholar]
  57. Zhang Y., Feng X., We R., Derynck R. Receptor-associated Mad homologues synergize as effectors of the TGF-beta response. Nature. 1996 Sep 12;383(6596):168–172. doi: 10.1038/383168a0. [DOI] [PubMed] [Google Scholar]
  58. Zhu Y., Richardson J. A., Parada L. F., Graff J. M. Smad3 mutant mice develop metastatic colorectal cancer. Cell. 1998 Sep 18;94(6):703–714. doi: 10.1016/s0092-8674(00)81730-4. [DOI] [PubMed] [Google Scholar]

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