Abstract
We have previously identified mouse DDA3 as a p53-inducible gene. To explore the functional role of DDA3, we screened a mouse brain cDNA library by the yeast two-hybrid assay, and identified the microtubule plus-end binding protein EB3 as a DDA3-interacting protein. Binding of DDA3 to EB3 was verified by glutathione S-transferase (GST) pull-down assay and subcellular colocalization; co-immunoprecipitation further indicated that interaction of these two proteins within cells required intact microtubules. Domains of DDA3-EB3 interaction were mapped by GST pull-down assay to amino acids 118–241 and 242–329 of DDA3 and the N- and C-termini of EB3. Immunofluorescence analysis revealed colocalization of DDA3 with microtubules in various cell phases, and regions encompassing aa 118–241 and 242–329 contained microtubule-interacting and bundling activities. In vitro microtubule-binding assay showed that DDA3 and EB3 associated directly with microtubules, and cooperated with each other for microtubule binding. In addition, DDA3 bound to the EB3 interacting partner adenomatous polyposis coli 2 (APC2), a homolog of the tumor suppressor APC, which is a component of the β-catenin destruction complex. Ectopic expression of DDA3 and EB3 enhanced β-catenin-dependent transactivation and cyclin D1 production, whereas knockdown of endogenous DDA3 or EB3 inhibited β-catenin-mediated transactivation and the ability of cells to form colonies. Together, our results identify DDA3 as a novel microtubule-associated protein that binds to EB3, and implicate DDA3 and EB3 in the β-catenin-mediated growth signaling.
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References
Behrens J . (2000). Control of beta-catenin signaling in tumor development. Ann NY Acad Sci 910: 21–33.
Bensaad K, Tsuruta A, Selak MA, Vidal MN, Nakano K, Bartrons R et al. (2006). TIGAR, a p53-inducible regulator of glycolysis and apoptosis. Cell 126: 107–120.
Bienz M . (2002). The subcellular destinations of APC proteins. Nat Rev Mol Cell Biol 3: 328–338.
Brummelkamp TR, Bernards R, Agami R . (2002). A system for stable expression of short interfering RNAs in mammalian cells. Science 296: 550–553.
Bu W, Su LK . (2001). Regulation of microtubule assembly by human EB1 family proteins. Oncogene 20: 3185–3192.
Bu W, Su LK . (2003). Characterization of functional domains of human EB1 family proteins. J Biol Chem 278: 49721–49731.
Fang L, Li G, Liu G, Lee SW, Aaronson SA . (2001). p53 induction of heparin-binding EGF-like growth factor counteracts p53 growth suppression through activation of MAPK and PI3K/Akt signaling cascades. EMBO J 20: 1931–1939.
Hainaut P, Hernandez T, Robinson A, Rodriguez-Tome P, Flores T, Hollstein M et al. (1998). IARC Database of p53 gene mutations in human tumors and cell lines: updated compilation, revised formats and new visualisation tools. Nucleic Acids Res 26: 205–213.
Han JA, Kim JI, Ongusaha PP, Hwang DH, Ballou LR, Mahale A et al. (2002). P53-mediated induction of Cox-2 counteracts p53- or genotoxic stress-induced apoptosis. EMBO J 21: 5635–5644.
Hanson CA, Miller JR . (2005). Non-traditional roles for the adenomatous polyposis coli (APC) tumor suppressor protein. Gene 361: 1–12.
He TC, Sparks AB, Rago C, Hermeking H, Zawel L, da Costa LT et al. (1998). Identification of c-MYC as a target of the APC pathway. Science 281: 1509–1512.
Hoffman WH, Biade S, Zilfou JT, Chen J, Murphy M . (2002). Transcriptional repression of the anti-apoptotic survivin gene by wild type p53. J Biol Chem 277: 3247–3257.
Hofseth LJ, Hussain SP, Harris CC . (2004). p53: 25 years after its discovery. Trends Pharmacol Sci 25: 177–181.
Honnappa S, John CM, Kostrewa D, Winkler FK, Steinmetz MO . (2005). Structural insights into the EB1–APC interaction. EMBO J 24: 261–269.
Hsieh SC, Lo PK, Wang FF . (2002). Mouse DDA3 gene is a direct transcriptional target of p53 and p73. Oncogene 21: 3050–3057.
Iwai A, Marusawa H, Matsuzawa S, Fukushima T, Hijikata M, Reed JC et al. (2004). Siah-1L, a novel transcript variant belonging to the human Siah family of proteins, regulates beta-catenin activity in a p53-dependent manner. Oncogene 23: 7593–7600.
Jarrett CR, Blancato J, Cao T, Bressette DS, Cepeda M, Young PE et al. (2001). Human APC2 localization and allelic imbalance. Cancer Res 61: 7978–7984.
Johnsen JI, Aurelio ON, Kwaja Z, Jorgensen GE, Pellegata NS, Plattner R et al. (2000). p53-mediated negative regulation of stathmin/Op18 expression is associated with G(2)/M cell-cycle arrest. Int J Cancer 88: 685–691.
Levine AJ, Finlay CA, Hinds PW . (2004). P53 is a tumor suppressor gene. Cell 116: S67–S69, 1.
Lo PK, Chen JY, Lo WC, Chen BF, Hsin JP, Tang PP et al. (1999). Identification of a novel mouse p53 target gene DDA3. Oncogene 18: 7765–7774.
Matsuzawa SI, Reed JC . (2001). Siah-1, SIP, and Ebi collaborate in a novel pathway for beta-catenin degradation linked to p53 responses. Mol Cell 7: 915–926.
Miller SJ, Rangwala F, Williams J, Ackerman P, Kong S, Jegga AG et al. (2006). Large-scale molecular comparison of human schwann cells to malignant peripheral nerve sheath tumor cell lines and tissues. Cancer Res 66: 2584–2591.
Moon RT, Kohn AD, De Ferrari GV, Kaykas A . (2004). WNT and beta-catenin signalling: diseases and therapies. Nat Rev Genet 5: 691–701.
Nakagawa H, Koyama K, Murata Y, Morito M, Akiyama T, Nakamura Y . (2000). EB3, a novel member of the EB1 family preferentially expressed in the central nervous system, binds to a CNS-specific APC homologue. Oncogene 19: 210–216.
Nakagawa H, Murata Y, Koyama K, Fujiyama A, Miyoshi Y, Monden M et al. (1998). Identification of a brain-specific APC homologue, APCL, and its interaction with beta-catenin. Cancer Res 58: 5176–5181.
Nakamura M, Zhou XZ, Lu KP . (2001). Critical role for the EB1 and APC interaction in the regulation of microtubule polymerization. Curr Biol 11: 1062–1067.
Ongusaha PP, Kim JI, Fang L, Wong TW, Yancopoulos GD, Aaronson SA et al. (2003). p53 induction and activation of DDR1 kinase counteract p53-mediated apoptosis and influence p53 regulation through a positive feedback loop. EMBO J 22: 1289–1301.
Park WR, Nakamura Y . (2005). p53CSV, a novel p53-inducible gene involved in the p53-dependent cell-survival pathway. Cancer Res 65: 1197–1206.
Senda T, Shimomura A, Iizuka-Kogo A . (2005). Adenomatous polyposis coli (APC) tumor suppressor gene as a multifunctional gene. Anat Sci Int 80: 121–131.
Su LK, Burrell M, Hill DE, Gyuris J, Brent R, Wiltshire R et al. (1995). APC binds to the novel protein EB1. Cancer Res 55: 2972–2977.
Tetsu O, McCormick F . (1999). Beta-catenin regulates expression of cyclin D1 in colon carcinoma cells. Nature 398: 422–426.
van Es JH, Kirkpatrick C, van de WM, Molenaar M, Miles A, Kuipers J et al. (1999). Identification of APC2, a homologue of the adenomatous polyposis coli tumour suppressor. Curr Biol 9: 105–108.
Vogelstein B, Lane D, Levine AJ . (2000). Surfing the p53 network. Nature 408: 307–310.
Wang Y, Zhou X, Zhu H, Liu S, Zhou C, Zhang G et al. (2005). Overexpression of EB1 in human esophageal squamous cell carcinoma (ESCC) may promote cellular growth by activating beta-catenin/TCF pathway. Oncogene 24: 6637–6645.
Acknowledgements
This work was supported by Grants (NHRI-EX92-9124BI and NHRI-EX93-9124BI) from the National Health Research Institute, Taiwan.
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Supplementary Information accompanies the paper on the Oncogene website (http://www.nature.com/onc).
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Hsieh, PC., Chang, JC., Sun, WT. et al. p53 downstream target DDA3 is a novel microtubule-associated protein that interacts with end-binding protein EB3 and activates β-catenin pathway. Oncogene 26, 4928–4940 (2007). https://doi.org/10.1038/sj.onc.1210304
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DOI: https://doi.org/10.1038/sj.onc.1210304
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