Phosphorylation of Serine 68 of Twist1 by MAPKs Stabilizes Twist1 Protein and Promotes Breast Cancer Cell Invasiveness

Cancer Research - Tập 71 Số 11 - Trang 3980-3990 - 2011
Jun Ki Hong1, Jian Zhou1, Junjiang Fu1, Tao He1, Jun Qin1, Li Wang1, Lan Liao1, Jianming Xu1
1Authors' Affiliations: 1Department of Molecular and Cellular Biology and Dan L. Duncan Cancer Center, Baylor College of Medicine, Houston, Texas; and 2Luzhou Medical College, Luzhou, Sichuan 646000, China

Tóm tắt

Abstract Twist1, a basic helix–loop–helix transcription factor, promotes breast tumor cell epithelial–mesenchymal transition (EMT), invasiveness, and metastasis. However, the mechanisms responsible for regulating Twist1 stability are unknown in these cells. We identified the serine 68 (Ser 68) as a major phosphorylation site of Twist1 by mass spectrometry and with specific antibodies. This Ser 68 is phosphorylated by p38, c-Jun N-terminal kinases (JNK), and extracellular signal-regulated kinases1/2 in vitro, and its phosphorylation levels positively correlate with Twist1 protein levels in human embryonic kidney 293 and breast cancer cells. Prevention of Ser 68 phosphorylation by an alanine (A) mutation (Ser 68A) dramatically accelerates Twist1 ubiquitination and degradation. Furthermore, activation of mitogen-activated protein kinases (MAPK) by an active Ras protein or TGF-β treatment significantly increases Ser 68 phosphorylation and Twist1 protein levels without altering Twist1 mRNA expression, whereas blocking of MAPK activities by either specific inhibitors or dominant negative inhibitory mutants effectively reduces the levels of both induced and uninduced Ser 68 phosphorylation and Twist protein. Accordingly, the mammary epithelial cells expressing Twist1 exhibit much higher degrees of EMT and invasiveness on stimulation with TGF-β or the active Ras and paclitaxel resistance compared with the same cells expressing the Ser 68A-Twist1 mutant. Importantly, the levels of Ser 68 phosphorylation in the invasive human breast ductal carcinomas positively correlate with the levels of Twist1 protein and JNK activity and are significantly higher in progesterone receptor–negative and HER2-positive breast cancers. These findings suggest that activation of MAPKs by tyrosine kinase receptors and Ras signaling pathways may substantially promote breast tumor cell EMT and metastasis via phoshorylation and stabilization of Twist1. Cancer Res; 71(11); 3980–90. ©2011 AACR.

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Tài liệu tham khảo

Pearson, 2001, Mitogen-activated protein (MAP) kinase pathways: regulation and physiological functions, Endocr Rev, 22, 153

Haagenson, 2010, The role of MAP kinases and MAP kinase phosphatase-1 in resistance to breast cancer treatment, Cancer Metastasis Rev, 29, 143, 10.1007/s10555-010-9208-5

Salh, 1999, Investigation of the Mek-MAP kinase-Rsk pathway in human breast cancer, Anticancer Res, 19, 731

Johnston, 1999, Increased activator protein-1 DNA binding and c-Jun NH2-terminal kinase activity in human breast tumors with acquired tamoxifen resistance, Clin Cancer Res, 5, 251

Schiff, 2000, Oxidative stress and AP-1 activity in tamoxifen-resistant breast tumors in vivo, J Natl Cancer Inst, 92, 1926, 10.1093/jnci/92.23.1926

Murtagh, 2004, Organization of mammary epithelial cells into 3D acinar structures requires glucocorticoid and JNK signaling, J Cell Biol, 166, 133, 10.1083/jcb.200403020

Iiizumi, 2008, RhoC promotes metastasis via activation of the Pyk2 pathway in prostate cancer, Cancer Res, 68, 7613, 10.1158/0008-5472.CAN-07-6700

Malaney, 2001, The ras signaling pathway in mammary tumorigenesis and metastasis, J Mammary Gland Biol Neoplasia, 6, 101, 10.1023/A:1009572700317

Webb, 1998, Signaling pathways in Ras-mediated tumorigenicity and metastasis, Proc Natl Acad Sci U S A, 95, 8773, 10.1073/pnas.95.15.8773

Sinn, 1987, Coexpression of MMTV/v-Ha-ras and MMTV/c-myc genes in transgenic mice: synergistic action of oncogenes in vivo, Cell, 49, 465, 10.1016/0092-8674(87)90449-1

Behbod, 2009, An intraductal human-in-mouse transplantation model mimics the subtypes of ductal carcinoma in situ, Breast Cancer Res, 11, R66, 10.1186/bcr2358

Arnold, 1995, Phosphorylation of the human estrogen receptor by mitogen-activated protein kinase and casein kinase II: consequence on DNA binding, J Steroid Biochem Mol Biol, 55, 163, 10.1016/0960-0760(95)00177-2

Derijard, 1994, JNK1: a protein kinase stimulated by UV light and Ha-Ras that binds and phosphorylates the c-Jun activation domain, Cell, 76, 1025, 10.1016/0092-8674(94)90380-8

Viatour, 2005, Phosphorylation of NF-kappaB and IkappaB proteins: implications in cancer and inflammation, Trends Biochem Sci, 30, 43, 10.1016/j.tibs.2004.11.009

Foulds, 2004, Ras/mitogen-activated protein kinase signaling activates Ets-1 and Ets-2 by CBP/p300 recruitment, Mol Cell Biol, 24, 10954, 10.1128/MCB.24.24.10954-10964.2004

Shin, 2010, ERK2 but not ERK1 induces epithelial-to-mesenchymal transformation via DEF motif-dependent signaling events, Mol Cell, 38, 114, 10.1016/j.molcel.2010.02.020

Yang, 2004, Twist, a master regulator of morphogenesis, plays an essential role in tumor metastasis, Cell, 117, 927, 10.1016/j.cell.2004.06.006

Lee, 2006, Twist overexpression correlates with hepatocellular carcinoma metastasis through induction of epithelial-mesenchymal transition, Clin Cancer Res, 12, 5369, 10.1158/1078-0432.CCR-05-2722

Vesuna, 2008, Twist is a transcriptional repressor of E-cadherin gene expression in breast cancer, Biochem Biophys Res Commun, 367, 235, 10.1016/j.bbrc.2007.11.151

Qin, 2009, The steroid receptor coactivator-1 regulates twist expression and promotes breast cancer metastasis, Cancer Res, 69, 3819, 10.1158/0008-5472.CAN-08-4389

Fu, The TWIST/Mi2/NuRD protein complex and its essential role in cancer metastasis, Cell Res, 21, 2012

Mani, 2008, The epithelial-mesenchymal transition generates cells with properties of stem cells, Cell, 133, 704, 10.1016/j.cell.2008.03.027

Ansieau, 2008, Induction of EMT by twist proteins as a collateral effect of tumor-promoting inactivation of premature senescence, Cancer Cell, 14, 79, 10.1016/j.ccr.2008.06.005

Zhang, 1998, Identification of phosphorylation sites in proteins separated by polyacrylamide gel electrophoresis, Anal Chem, 70, 2050, 10.1021/ac971207m

El Ghouzzi, 2000, Saethre-Chotzen mutations cause TWIST protein degradation or impaired nuclear location, Hum Mol Genet, 9, 813, 10.1093/hmg/9.5.813

Widmann, 1999, Mitogen-activated protein kinase: conservation of a three-kinase module from yeast to human, Physiol Rev, 79, 143, 10.1152/physrev.1999.79.1.143

Li, 1996, The Ras-JNK pathway is involved in shear-induced gene expression, Mol Cell Biol, 16, 5947, 10.1128/MCB.16.11.5947

Matsumoto, 1999, Platelet-derived growth factor activates p38 mitogen-activated protein kinase through a Ras-dependent pathway that is important for actin reorganization and cell migration, J Biol Chem, 274, 13954, 10.1074/jbc.274.20.13954

Tibbles, 1999, The stress-activated protein kinase pathways, Cell Mol Life Sci, 55, 1230, 10.1007/s000180050369

Raingeaud, 1995, Pro-inflammatory cytokines and environmental stress cause p38 mitogen-activated protein kinase activation by dual phosphorylation on tyrosine and threonine, J Biol Chem, 270, 7420, 10.1074/jbc.270.13.7420

Dickens, 1997, A cytoplasmic inhibitor of the JNK signal transduction pathway, Science, 277, 693, 10.1126/science.277.5326.693

Pages, 1994, Constitutive mutant and putative regulatory serine phosphorylation site of mammalian MAP kinase kinase (MEK1), Embo J, 13, 3003, 10.1002/j.1460-2075.1994.tb06599.x

Basolo, 1991, Transformation of human breast epithelial cells by c-Ha-ras oncogene, Mol Carcinog, 4, 25, 10.1002/mc.2940040106

Padua, 2008, TGFbeta primes breast tumors for lung metastasis seeding through angiopoietin-like 4, Cell, 133, 66, 10.1016/j.cell.2008.01.046

Yue, 2000, Requirement of Ras/MAPK pathway activation by transforming growth factor beta for transforming growth factor beta 1 production in a Smad-dependent pathway, J Biol Chem, 275, 35656, 10.1016/S0021-9258(20)88877-2

Yu, 2002, TGF-beta receptor-activated p38 MAP kinase mediates Smad-independent TGF-beta responses, EMBO J, 21, 3749, 10.1093/emboj/cdf366

Wang, 2004, Identification of a novel function of TWIST, a bHLH protein, in the development of acquired taxol resistance in human cancer cells, Oncogene, 23, 474, 10.1038/sj.onc.1207128

Lluis, 2005, E47 phosphorylation by p38 MAPK promotes MyoD/E47 association and muscle-specific gene transcription, EMBO J, 24, 974, 10.1038/sj.emboj.7600528

Forrest, 2004, Phosphorylation regulates Id3 function in vascular smooth muscle cells, Circ Res, 95, 557, 10.1161/01.RES.0000142735.67542.5a

Firulli, 2005, Altered Twist1 and Hand2 dimerization is associated with Saethre-Chotzen syndrome and limb abnormalities, Nat Genet, 37, 373, 10.1038/ng1525

Firulli, 2007, Mutations within helix I of Twist1 result in distinct limb defects and variation of DNA binding affinities, J Biol Chem, 282, 27536, 10.1074/jbc.M702613200

Firulli, 2008, Phosphoregulation of Twist1 provides a mechanism of cell fate control, Curr Med Chem, 15, 2641, 10.2174/092986708785908987

Vichalkovski, 2010, PKB/AKT phosphorylation of the transcription factor Twist-1 at Ser42 inhibits p53 activity in response to DNA damage, Oncogene, 29, 3554, 10.1038/onc.2010.115

Buschmann, 2001, Jun NH2-terminal kinase phosphorylation of p53 on Thr-81 is important for p53 stabilization and transcriptional activities in response to stress, Mol Cell Biol, 21, 2743, 10.1128/MCB.21.8.2743-2754.2001

Yamashita, 2005, Ras-ERK MAPK cascade regulates GATA3 stability and Th2 differentiation through ubiquitin-proteasome pathway, J Biol Chem, 280, 29409, 10.1074/jbc.M502333200

Sears, 1999, Ras enhances Myc protein stability, Mol Cell, 3, 169, 10.1016/S1097-2765(00)80308-1

Hayashi, 2007, Comparative roles of Twist-1 and Id1 in transcriptional regulation by BMP signaling, J Cell Sci, 120, 1350, 10.1242/jcs.000067

Demontis, 2006, Twist is substrate for caspase cleavage and proteasome-mediated degradation, Cell Death Differ, 13, 335, 10.1038/sj.cdd.4401744

Llense, 2008, JNK signaling controls border cell cluster integrity and collective cell migration, Curr Biol, 18, 538, 10.1016/j.cub.2008.03.029

Pastor-Pareja, 2004, Invasive cell behavior during Drosophila imaginal disc eversion is mediated by the JNK signaling cascade, Dev Cell, 7, 387, 10.1016/j.devcel.2004.07.022

Santibanez, 2006, JNK mediates TGF-beta1-induced epithelial mesenchymal transdifferentiation of mouse transformed keratinocytes, FEBS Lett, 580, 5385, 10.1016/j.febslet.2006.09.003

Hagemann, 2005, Macrophages induce invasiveness of epithelial cancer cells via NF-kappa B and JNK, J Immunol, 175, 1197, 10.4049/jimmunol.175.2.1197

Osborne, 2005, Endocrine responsiveness: understanding how progesterone receptor can be used to select endocrine therapy, Breast, 14, 458, 10.1016/j.breast.2005.08.024

Wei, 2008, Bone metastasis is strongly associated with estrogen receptor-positive/progesterone receptor-negative breast carcinomas, Hum Pathol, 39, 1809, 10.1016/j.humpath.2008.05.010

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Cancer Research

Tập 71 Số 11

3980-3990

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