Cellular and Molecular Life Sciences

, Volume 73, Issue 5, pp 1085–1101 | Cite as

TRIM39 negatively regulates the NFκB-mediated signaling pathway through stabilization of Cactin

  • Masanobu Suzuki
  • Masashi Watanabe
  • Yuji Nakamaru
  • Dai Takagi
  • Hidehisa Takahashi
  • Satoshi Fukuda
  • Shigetsugu Hatakeyama
Original Article


NFκB is one of the central regulators of cell survival, immunity, inflammation, carcinogenesis and organogenesis. The activation of NFκB is strictly regulated by several posttranslational modifications including phosphorylation, neddylation and ubiquitination. Several types of ubiquitination play important roles in multi-step regulations of the NFκB pathway. Some of the tripartite motif-containing (TRIM) proteins functioning as E3 ubiquitin ligases are known to regulate various biological processes such as inflammatory signaling pathways. One of the TRIM family proteins, TRIM39, for which the gene has single nucleotide polymorphisms, has been identified as one of the genetic factors in Behcet’s disease. However, the role of TRIM39 in inflammatory signaling had not been fully elucidated. In this study, to elucidate the function of TRIM39 in inflammatory signaling, we performed yeast two-hybrid screening using TRIM39 as a bait and identified Cactin, which has been reported to inhibit NFκB- and TLR-mediated transcriptions. We show that TRIM39 stabilizes Cactin protein and that Cactin is upregulated after TNFα stimulation. TRIM39 knockdown also causes activation of the NFκB signal. These findings suggest that TRIM39 negatively regulates the NFκB signal in collaboration with Cactin induced by inflammatory stimulants such as TNFα.


RelA/p65 IκBα TLR Ubiquitin Ubiquitin ligase E3 



We are grateful to Dr. M. Bohgaki for providing the reagent for this study, Ms. M. Uchiumi for help in preparing the manuscript and Ms. M. Matsuo for technical assistance. We are also grateful to Dr. H. Hatakeyama, Dr. S. Kano, Dr. T. Mizumachi, Dr. T. Sakashita, Dr. K. Mizoguchi, Dr. A. Homma, and Dr. T. Suzuki for technical advice. This work was supported in part by KAKENHI (24112006, 24390065, 15H04690 to S.H.) from the Ministry of Education, Culture, Sports, Science and Technology of Japan and by The Uehara Memorial Foundation and the Japan Rheumatism Foundation (to S.H.).

Compliance with ethical standards

Conflict of interest

The authors declare no competing financial interests.

Supplementary material

18_2015_2040_MOESM1_ESM.pdf (63 kb)
Supplementary material 1 (PDF 63 kb)
18_2015_2040_MOESM2_ESM.pdf (107 kb)
Supplementary material 2 (PDF 106 kb)
18_2015_2040_MOESM3_ESM.pdf (84 kb)
Supplementary material 3 (PDF 83 kb)
18_2015_2040_MOESM4_ESM.pdf (66 kb)
Supplementary material 4 (PDF 66 kb)
18_2015_2040_MOESM5_ESM.pdf (117 kb)
Supplementary material 5 (PDF 117 kb)
18_2015_2040_MOESM6_ESM.pdf (63 kb)
Supplementary material 6 (PDF 62 kb)


  1. 1.
    Hayden MS, Ghosh S (2004) Signaling to NF-kappaB. Genes Dev 18:2195–2224CrossRefPubMedGoogle Scholar
  2. 2.
    Hayden MS, Ghosh S (2008) Shared principles in NF-kappaB signaling. Cell 132:344–362CrossRefPubMedGoogle Scholar
  3. 3.
    Hatakeyama S, Kitagawa M, Nakayama K, Shirane M, Matsumoto M, Hattori K, Higashi H, Nakano H, Okumura K, Onoe K, Good RA (1999) Ubiquitin-dependent degradation of IkappaBalpha is mediated by a ubiquitin ligase Skp1/Cul 1/F-box protein FWD1. Proc Natl Acad Sci USA 96:3859–3863PubMedCentralCrossRefPubMedGoogle Scholar
  4. 4.
    Li Y, Gazdoiu S, Pan ZQ, Fuchs SY (2004) Stability of homologue of Slimb F-box protein is regulated by availability of its substrate. J Biol Chem 279:11074–11080CrossRefPubMedGoogle Scholar
  5. 5.
    Rahighi S, Ikeda F, Kawasaki M, Akutsu M, Suzuki N, Kato R, Kensche T, Uejima T, Bloor S, Komander D, Randow F, Wakatsuki S, Dikic I (2009) Specific recognition of linear ubiquitin chains by NEMO is important for NF-kappaB activation. Cell 136:1098–1109CrossRefPubMedGoogle Scholar
  6. 6.
    Hershko A, Ciechanover A (1998) The ubiquitin system. Annu Rev Biochem 67:425–479CrossRefPubMedGoogle Scholar
  7. 7.
    Scheffner M, Nuber U, Huibregtse JM (1995) Protein ubiquitination involving an E1–E2–E3 enzyme ubiquitin thioester cascade. Nature 373:81–83CrossRefPubMedGoogle Scholar
  8. 8.
    Hershko A, Heller H, Elias S, Ciechanover A (1983) Components of ubiquitin-protein ligase system. Resolution, affinity purification, and role in protein breakdown. J Biol Chem 258:8206–8214PubMedGoogle Scholar
  9. 9.
    Huibregtse JM, Scheffner M, Beaudenon S, Howley PM (1995) A family of proteins structurally and functionally related to the E6-AP ubiquitin-protein ligase. Proc Natl Acad Sci USA 92:2563–2567PubMedCentralCrossRefPubMedGoogle Scholar
  10. 10.
    Lorick KL, Jensen JP, Fang S, Ong AM, Hatakeyama S, Weissman AM (1999) RING fingers mediate ubiquitin-conjugating enzyme (E2)-dependent ubiquitination. Proc Natl Acad Sci USA 96:11364–11369PubMedCentralCrossRefPubMedGoogle Scholar
  11. 11.
    Hatakeyama S, Yada M, Matsumoto M, Ishida N, Nakayama KI (2001) U box proteins as a new family of ubiquitin-protein ligases. J Biol Chem 276:33111–33120CrossRefPubMedGoogle Scholar
  12. 12.
    Meroni G, Diez-Roux G (2005) TRIM/RBCC, a novel class of ‘single protein RING finger’ E3 ubiquitin ligases. Bioessays 27:1147–1157CrossRefPubMedGoogle Scholar
  13. 13.
    Kano S, Miyajima N, Fukuda S, Hatakeyama S (2008) Tripartite motif protein 32 facilitates cell growth and migration via degradation of Abl-interactor 2. Cancer Res 68:5572–5580CrossRefPubMedGoogle Scholar
  14. 14.
    Hatakeyama S (2011) TRIM proteins and cancer. Nat Rev Cancer 11:792–804CrossRefPubMedGoogle Scholar
  15. 15.
    Zhang L, Huang NJ, Chen C, Tang W, Kornbluth S (2012) Ubiquitylation of p53 by the APC/C inhibitor Trim39. Proc Natl Acad Sci USA 109:20931–20936PubMedCentralCrossRefPubMedGoogle Scholar
  16. 16.
    Lee SS, Fu NY, Sukumaran SK, Wan KF, Wan Q, Yu VC (2009) TRIM39 is a MOAP-1-binding protein that stabilizes MOAP-1 through inhibition of its poly-ubiquitination process. Exp Cell Res 315:1313–1325CrossRefPubMedGoogle Scholar
  17. 17.
    Nisole S, Stoye JP, Saib A (2005) TRIM family proteins: retroviral restriction and antiviral defence. Nat Rev Microbiol 3:799–808CrossRefPubMedGoogle Scholar
  18. 18.
    Reymond A, Meroni G, Fantozzi A, Merla G, Cairo S, Luzi L, Riganelli D, Zanaria E, Messali S, Cainarca S, Guffanti A, Minucci S, Pelicci PG, Ballabio A (2001) The tripartite motif family identifies cell compartments. EMBO J 20:2140–2151PubMedCentralCrossRefPubMedGoogle Scholar
  19. 19.
    Versteeg GA, Rajsbaum R, Sanchez-Aparicio MT, Maestre AM, Valdiviezo J, Shi M, Inn KS, Fernandez-Sesma A, Jung J, Garcia-Sastre A (2013) The E3-ligase TRIM family of proteins regulates signaling pathways triggered by innate immune pattern-recognition receptors. Immunity 38:384–398PubMedCentralCrossRefPubMedGoogle Scholar
  20. 20.
    Kurata R, Tajima A, Yonezawa T, Inoko H (2013) TRIM39R, but not TRIM39B, regulates type I interferon response. Biochem Biophys Res Commun 436:90–95CrossRefPubMedGoogle Scholar
  21. 21.
    Kurata R, Nakaoka H, Tajima A, Hosomichi K, Shiina T, Meguro A, Mizuki N, Ohono S, Inoue I, Inoko H (2010) TRIM39 and RNF39 are associated with Behcet’s disease independently of HLA-B *51 and -A *26. Biochem Biophys Res Commun 401:533–537CrossRefPubMedGoogle Scholar
  22. 22.
    Nakajima K, Yamanaka Y, Nakae K, Kojima H, Ichiba M, Kiuchi N, Kitaoka T, Fukada T, Hibi M, Hirano T (1996) A central role for Stat3 in IL-6-induced regulation of growth and differentiation in M1 leukemia cells. EMBO J 15:3651–3658PubMedCentralPubMedGoogle Scholar
  23. 23.
    Matsuda M, Tsukiyama T, Bohgaki M, Nonomura K, Hatakeyama S (2007) Establishment of a newly improved detection system for NF-kappaB activity. Immunol Lett 109:175–181CrossRefPubMedGoogle Scholar
  24. 24.
    Kondo T, Watanabe M, Hatakeyama S (2012) TRIM59 interacts with ECSIT and negatively regulates NF-kappaB and IRF-3/7-mediated signal pathways. Biochem Biophys Res Commun 422:501–507CrossRefPubMedGoogle Scholar
  25. 25.
    Noguchi K, Okumura F, Takahashi N, Kataoka A, Kamiyama T, Todo S, Hatakeyama S (2011) TRIM40 promotes neddylation of IKKgamma and is downregulated in gastrointestinal cancers. Carcinogenesis 32:995–1004CrossRefPubMedGoogle Scholar
  26. 26.
    Watanabe M, Tsukiyama T, Hatakeyama S (2009) TRIM31 interacts with p52(Shc) and inhibits Src-induced anchorage-independent growth. Biochem Biophys Res Commun 388:422–427CrossRefPubMedGoogle Scholar
  27. 27.
    Atzei P, Gargan S, Curran N, Moynagh PN (2010) Cactin targets the MHC class III protein IkappaB-like (IkappaBL) and inhibits NF-kappaB and interferon-regulatory factor signaling pathways. J Biol Chem 285:36804–36817PubMedCentralCrossRefPubMedGoogle Scholar
  28. 28.
    Lin P, Huang LH, Steward R (2000) Cactin, a conserved protein that interacts with the Drosophila IkappaB protein cactus and modulates its function. Mech Dev 94:57–65CrossRefPubMedGoogle Scholar
  29. 29.
    Arenzana-Seisdedos F, Thompson J, Rodriguez MS, Bachelerie F, Thomas D, Hay RT (1995) Inducible nuclear expression of newly synthesized I kappa B alpha negatively regulates DNA-binding and transcriptional activities of NF-kappa B. Mol Cell Biol 15:2689–2696PubMedCentralCrossRefPubMedGoogle Scholar
  30. 30.
    Jin J, Samuvel DJ, Zhang X, Li Y, Lu Z, Lopes-Virella MF, Huang Y (2011) Coactivation of TLR4 and TLR2/6 coordinates an additive augmentation on IL-6 gene transcription via p38MAPK pathway in U937 mononuclear cells. Mol Immunol 49:423–432PubMedCentralCrossRefPubMedGoogle Scholar
  31. 31.
    Atzei P, Yang F, Collery R, Kennedy BN, Moynagh PN (2010) Characterisation of expression patterns and functional role of Cactin in early zebrafish development. Gene Expr Patterns 10:199–206CrossRefPubMedGoogle Scholar
  32. 32.
    Tannoury H, Rodriguez V, Kovacevic I, Ibourk M, Lee M, Cram EJ (2010) CACN-1/Cactin interacts genetically with MIG-2 GTPase signaling to control distal tip cell migration in C. elegans. Dev Biol 341:176–185PubMedCentralCrossRefPubMedGoogle Scholar
  33. 33.
    LaBonty M, Szmygiel C, Byrnes LE, Hughes S, Woollard A, Cram EJ (2014) CACN-1/Cactin plays a role in Wnt signaling in C. elegans. PLoS One 9:e101945PubMedCentralCrossRefPubMedGoogle Scholar
  34. 34.
    Lehner B, Semple JI, Brown SE, Counsell D, Campbell RD, Sanderson CM (2004) Analysis of a high-throughput yeast two-hybrid system and its use to predict the function of intracellular proteins encoded within the human MHC class III region. Genomics 83:153–167CrossRefPubMedGoogle Scholar
  35. 35.
    Kanayama A, Seth RB, Sun L, Ea CK, Hong M, Shaito A, Chiu YH, Deng L, Chen ZJ (2004) TAB 2 and TAB 3 activate the NF-kappaB pathway through binding to polyubiquitin chains. Mol Cell 15:535–548CrossRefPubMedGoogle Scholar
  36. 36.
    Krutzfeldt M, Ellis M, Weekes DB, Bull JJ, Eilers M, Vivanco MD, Sellers WR, Mittnacht S (2005) Selective ablation of retinoblastoma protein function by the RET finger protein. Mol Cell 18:213–224CrossRefPubMedGoogle Scholar
  37. 37.
    Boone DL, Turer EE, Lee EG, Ahmad RC, Wheeler MT, Tsui C, Hurley P, Chien M, Chai S, Hitotsumatsu O, McNally E, Pickart C, Ma A (2004) The ubiquitin-modifying enzyme A20 is required for termination of Toll-like receptor responses. Nat Immunol 5:1052–1060CrossRefPubMedGoogle Scholar
  38. 38.
    Wertz IE, O’Rourke KM, Zhou H, Eby M, Aravind L, Seshagiri S, Wu P, Wiesmann C, Baker R, Boone DL, Ma A, Koonin EV, Dixit VM (2004) De-ubiquitination and ubiquitin ligase domains of A20 downregulate NF-kappaB signalling. Nature 430:694–699CrossRefPubMedGoogle Scholar
  39. 39.
    Burns K, Janssens S, Brissoni B, Olivos N, Beyaert R, Tschopp J (2003) Inhibition of interleukin 1 receptor/Toll-like receptor signaling through the alternatively spliced, short form of MyD88 is due to its failure to recruit IRAK-4. J Exp Med 197:263–268PubMedCentralCrossRefPubMedGoogle Scholar
  40. 40.
    Kimchi-Sarfaty C, Oh JM, Kim IW, Sauna ZE, Calcagno AM, Ambudkar SV, Gottesman MM (2007) A “silent” polymorphism in the MDR1 gene changes substrate specificity. Science 315:525–528CrossRefPubMedGoogle Scholar
  41. 41.
    Okamoto K, Makino S, Yoshikawa Y, Takaki A, Nagatsuka Y, Ota M, Tamiya G, Kimura A, Bahram S, Inoko H (2003) Identification of I kappa BL as the second major histocompatibility complex-linked susceptibility locus for rheumatoid arthritis. Am J Hum Genet 72:303–312PubMedCentralCrossRefPubMedGoogle Scholar
  42. 42.
    Shibata H, Yasunami M, Obuchi N, Takahashi M, Kobayashi Y, Numano F, Kimura A (2006) Direct determination of single nucleotide polymorphism haplotype of NFKBIL1 promoter polymorphism by DNA conformation analysis and its application to association study of chronic inflammatory diseases. Hum Immunol 67:363–373CrossRefPubMedGoogle Scholar
  43. 43.
    de la Concha EG, Fernandez-Arquero M, Lopez-Nava G, Martin E, Allcock RJ, Conejero L, Paredes JG, Diaz-Rubio M (2000) Susceptibility to severe ulcerative colitis is associated with polymorphism in the central MHC gene IKBL. Gastroenterology 119:1491–1495CrossRefPubMedGoogle Scholar
  44. 44.
    Castiblanco J, Anaya JM (2008) The IkappaBL gene polymorphism influences risk of acquiring systemic lupus erythematosus and Sjogren’s syndrome. Hum Immunol 69:45–51CrossRefPubMedGoogle Scholar
  45. 45.
    Yamashita T, Hamaguchi K, Kusuda Y, Kimura A, Sakata T, Yoshimatsu H (2014) IKBL promoter polymorphism is strongly associated with resistance to type 1 diabetes in Japanese. Tissue Antigens 63:223–230CrossRefGoogle Scholar
  46. 46.
    Miterski B, Bohringer S, Klein W, Sindern E, Haupts M, Schimrigk S, Epplen JT (2002) Inhibitors in the NFkappaB cascade comprise prime candidate genes predisposing to multiple sclerosis, especially in selected combinations. Genes Immun 3:211–219CrossRefPubMedGoogle Scholar

Copyright information

© Springer Basel 2015

Authors and Affiliations

  • Masanobu Suzuki
    • 1
    • 2
  • Masashi Watanabe
    • 1
  • Yuji Nakamaru
    • 2
  • Dai Takagi
    • 2
  • Hidehisa Takahashi
    • 1
  • Satoshi Fukuda
    • 2
  • Shigetsugu Hatakeyama
    • 1
  1. 1.Department of BiochemistryHokkaido University Graduate School of MedicineSapporoJapan
  2. 2.Department of Otolaryngology-Head and Neck SurgeryHokkaido University Graduate School of MedicineSapporoJapan

Personalised recommendations