CUL4A encodes a core component of a cullin-based E3 ubiquitin ligase complex that regulates many critical processes such as cell cycle progression, DNA replication, DNA repair and chromatin remodeling by targeting a variety of proteins for ubiquitination and degradation. In the research described in this report we aimed to clarify whether CUL4A participates in multiple drug resistance (MDR) in breast cancer cells. We first transfected vectors carrying CUL4A and specific shCUL4A into breast cancer cells and corresponding Adr cells respectively. Using reverse transcription polymerase chain reactions and western blots, we found that overexpression of CUL4A in MCF7 and MDA-MB-468 cells up-regulated MDR1 /P-gp expression on both the transcription and protein levels, which conferred multidrug resistance to P-gp substrate drugs, as determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays. On the other hand, silencing CUL4A in MCF7/Adr and MDA-MB-468/Adr cells led to the opposite effect. Moreover, ERK1/2 in CUL4A-overexpressing cells was highly activated and after treatment with PD98059, an ERK1/2-specific inhibitor, CUL4A-induced expression of MDR1 /P-gp was decreased significantly. Lastly, immunohistochemistry in breast cancer tissues showed that P-gp expression had a positive correlation with the expression of CUL4A and ERK1/2. Thus, these results implied that CUL4A and ERK1/2 participated in multi-drug resistance in breast cancer through regulation of MDR1/P-gp expression.
References
[1]
Forouzanfar, M.H.; Foreman, K.J.; Delossantos, A.M.; Lozano, R.; Lopez, A.D.; Murray, C.J.; Naghavi, M. Breast and cervical cancer in 187 countries between 1980 and 2010: A systematic analysis. Lancet 2011, 378, 1461–1484, doi:10.1016/S0140-6736(11)61351-2.
[2]
Siegel, R.; Naishadham, D.; Jemal, A. Cancer statistics, 2012. CA Cancer J. Clin. 2012, 62, 10–29, doi:10.3322/caac.20138.
[3]
Kuo, M.T. Roles of multidrug resistance genes in breast cancer chemoresistance. Adv. Exp. Med. Biol. 2007, 608, 23–30, doi:10.1007/978-0-387-74039-3_2.
[4]
Taheri, M.; Mahjoubi, F.; Omranipour, R. Effect of MDR1 polymorphism on multidrug resistance expression in breast cancer patients. Genet. Mol. Res. 2010, 9, 34–40, doi:10.4238/vol9-1gmr669.
[5]
Wind, N.S.; Holen, I. Multidrug resistance in breast cancer: From in vitro models to clinical studies. Int. J. Breast Cancer. 2011, 2011, doi:10.4061/2011/967419.
[6]
Patwardhan, G.; Gupta, V.; Huang, J.; Gu, X.; Liu, Y.Y. Direct assessment of P-glycoprotein efflux to determine tumor response to chemotherapy. Biochem. Pharmacol. 2010, 80, 72–79, doi:10.1016/j.bcp.2010.03.010.
[7]
Donmez, Y.; Gunduz, U. Reversal of multidrug resistance by small interfering RNA (siRNA) in doxorubicin-resistant MCF-7 breast cancer cells. Biomed. Pharmacother. 2011, 65, 85–89, doi:10.1016/j.biopha.2010.12.007.
[8]
Haber, M.; Bordow, S.B.; Haber, P.S.; Marshall, G.M.; Stewart, B.W.; Norris, M.D. The prognostic value of MDR1 gene expression in primary untreated neuroblastoma. Eur. J. Cancer 1997, 33, 2031–2036, doi:10.1016/S0959-8049(97)00229-3.
[9]
Norris, M.D.; Bordow, S.B.; Haber, P.S.; Marshall, G.M.; Kavallaris, M.; Madafiglio, J.; Cohn, S.L.; Salwen, H.; Schmidt, M.L.; Hipfner, D.R.; et al. Evidence that the MYCN oncogene regulates MRP gene expression in neuroblastoma. Eur. J. Cancer 1997, 33, 1911–1916, doi:10.1016/S0959-8049(97)00284-0.
[10]
Keppler, D. Multidrug resistance proteins (MRPs, ABCCs): Importance for pathophysiology and drug therapy. Handb. Exp. Pharmacol. 2011, 201, 299–323, doi:10.1007/978-3-642-14541-4_8.
[11]
Kimura, Y.; Morita, S.Y.; Matsuo, M.; Ueda, K. Mechanism of multidrug recognition by MDR1/ABCB1. Cancer Sci. 2007, 98, 1303–1310, doi:10.1111/j.1349-7006.2007.00538.x.
[12]
Ludwig, J.A.; Szakacs, G.; Martin, S.E.; Chu, B.F.; Cardarelli, C.; Sauna, Z.E.; Caplen, N.J.; Fales, H.M.; Ambudkar, S.V.; Weinstein, J.N.; et al. Selective toxicity of NSC73306 in MDR1-positive cells as a new strategy to circumvent multidrug resistance in cancer. Cancer Res. 2006, 66, 4808–4815, doi:10.1158/0008-5472.CAN-05-3322.
[13]
Chen, L.C.; Manjeshwar, S.; Lu, Y.; Moore, D.; Ljung, B.M.; Kuo, W.L.; Dairkee, S.H.; Wernick, M.; Collins, C.; Smith, H.S. The human homologue for the Caenorhabditis elegans cul-4 gene is amplified and overexpressed in primary breast cancers. Cancer Res. 1998, 58, 3677–3683.
[14]
Shiyanov, P.; Nag, A.; Raychaudhuri, P. Cullin 4A associates with the UV-damaged DNA-binding protein DDB. J. Biol. Chem. 1999, 274, 35309–35312, doi:10.1074/jbc.274.50.35309.
[15]
Gupta, A.; Yang, L.X.; Chen, L. Study of the G2/M cell cycle checkpoint in irradiated mammary epithelial cells overexpressing Cul-4A gene. Int. J. Radiat. Oncol. Biol. Phys. 2002, 52, 822–830, doi:10.1016/S0360-3016(01)02739-0.
[16]
Li, B.; Yang, F.C.; Clapp, D.W.; Chun, K.T. Enforced expression of CUL-4A interferes with granulocytic differentiation and exit from the cell cycle. Blood 2003, 101, 1769–1776, doi:10.1182/blood-2002-05-1517.
[17]
Sugasawa, K. DNA repair pathways involving Cul4A ubiquitin ligases. Tanpakushitsu Kakusan Koso. 2006, 51, 1339–1344.
[18]
Schindl, M.; Gnant, M.; Schoppmann, S.F.; Horvat, R.; Birner, P. Overexpression of the human homologue for Caenorhabditis elegans cul-4 gene is associated with poor outcome in node-negative breast cancer. Anticancer Res. 2007, 27, 949–952.
[19]
Ren, S.; Xu, C.; Cui, Z.; Yu, Y.; Xu, W.; Wang, F.; Lu, J.; Wei, M.; Lu, X.; Gao, X.; et al. Oncogenic CUL4A determines the response to thalidomide treatment in prostate cancer. J. Mol. Med. (Berl) 2012, 90, 1121–1132, doi:10.1007/s00109-012-0885-0.
[20]
Panka, D.J.; Atkins, M.B.; Mier, J.W. Targeting the mitogen-activated protein kinase pathway in the treatment of malignant melanoma. Clin. Cancer Res. 2006, 12, 2371S–2375S.
[21]
Johnson, G.L.; Lapadat, R. Mitogen-activated protein kinase pathways mediated by ERK, JNK, and p38 protein kinases. Science 2002, 298, 1911–1912, doi:10.1126/science.1072682.
Kohno, M.; Tanimura, S.; Ozaki, K. Targeting the extracellular signal-regulated kinase pathway in cancer therapy. Biol. Pharm. Bull. 2011, 34, 1781–1784, doi:10.1248/bpb.34.1781.
[24]
Wei, F.; Yan, J.; Tang, D. Extracellular signal-regulated kinases modulate DNA damage response—A contributing factor to using MEK inhibitors in cancer therapy. Curr. Med. Chem. 2011, 18, 5476–5482, doi:10.2174/092986711798194388.
[25]
Katayama, K.; Yoshioka, S.; Tsukahara, S.; Mitsuhashi, J.; Sugimoto, Y. Inhibition of the mitogen-activated protein kinase pathway results in the down-regulation of P-glycoprotein. Mol. Cancer Ther. 2007, 6, 2092–2102, doi:10.1158/1535-7163.MCT-07-0148.
[26]
Kisucka, J.; Barancik, M.; Bohacova, V.; Breier, A. Reversal effect of specific inhibitors of extracellular-signal regulated protein kinase pathway on P-glycoprotein mediated vincristine resistance of L1210 cells. Gen. Physiol. Biophys. 2001, 20, 439–444.
[27]
Lin, J.C.; Chang, S.Y.; Hsieh, D.S.; Lee, C.F.; Yu, D.S. Modulation of mitogen-activated protein kinase cascades by differentiation-1 protein: Acquired drug resistance of hormone independent prostate cancer cells. J. Urol. 2005, 174, 2022–2026, doi:10.1097/01.ju.0000176476.14572.39.
[28]
Li, Y.; Li, S.; Han, Y.; Liu, J.; Zhang, J.; Li, F.; Wang, Y.; Liu, X.; Yao, L. Calebin-A induces apoptosis and modulates MAPK family activity in drug resistant human gastric cancer cells. Eur. J. Pharmacol. 2008, 591, 252–258.
[29]
McCubrey, J.A.; Steelman, L.S.; Chappell, W.H.; Abrams, S.L.; Wong, E.W.; Chang, F.; Lehmann, B.; Terrian, D.M.; Milella, M.; Tafuri, A.; et al. Roles of the Raf/MEK/ERK pathway in cell growth, malignant transformation and drug resistance. Biochim. Biophys. Acta 2007, 1773, 1263–1284.
[30]
Higa, L.A.; Wu, M.; Ye, T.; Kobayashi, R.; Sun, H.; Zhang, H. CUL4-DDB1 ubiquitin ligase interacts with multiple WD40-repeat proteins and regulates histone methylation. Nat. Cell. Biol. 2006, 8, 1277–1283, doi:10.1038/ncb1490.
[31]
Di Cosimo, S.; Baselga, J. Management of breast cancer with targeted agents: Importance of heterogeneity [corrected]. Nat. Rev. Clin. Oncol. 2010, 7, 139–147, doi:10.1038/nrclinonc.2009.234.
[32]
Li, Q.Q.; Wang, W.J.; Xu, J.D.; Cao, X.X.; Chen, Q.; Yang, J.M.; Xu, Z.D. Involvement of CD147 in regulation of multidrug resistance to P-gp substrate drugs and in vitro invasion in breast cancer cells. Cancer Sci. 2007, 98, 1064–1069, doi:10.1111/j.1349-7006.2007.00487.x.
[33]
Xu, J.W.; Li, Q.Q.; Tao, L.L.; Cheng, Y.Y.; Yu, J.; Chen, Q.; Liu, X.P.; Xu, Z.D. Involvement of EGFR in the promotion of malignant properties in multidrug resistant breast cancer cells. Int. J. Oncol. 2011, 39, 1501–1509.
[34]
Amiri-Kordestani, L.; Basseville, A.; Kurdziel, K.; Fojo, A.T.; Bates, S.E. Targeting MDR in breast and lung cancer: Discriminating its potential importance from the failure of drug resistance reversal studies. Drug Resist. Updat. 2012, 15, 50–61, doi:10.1016/j.drup.2012.02.002.
[35]
Borowski, E.; Bontemps-Gracz, M.M.; Piwkowska, A. Strategies for overcoming ABC-transporters-mediated multidrug resistance (MDR) of tumor cells. Acta Biochim. Pol. 2005, 52, 609–627.
[36]
Hung, M.S.; Mao, J.H.; Xu, Z.; Yang, C.T.; Yu, J.S.; Harvard, C.; Lin, Y.C.; Bravo, D.T.; Jablons, D.M.; You, L. Cul4A is an oncogene in malignant pleural mesothelioma. J. Cell. Mol. Med. 2011, 15, 350–358, doi:10.1111/j.1582-4934.2009.00971.x.
[37]
Lee, J.; Zhou, P. Pathogenic role of the CRL4 ubiquitin ligase in human disease. Front. Oncol. 2012, 2, doi:10.3389/fonc.2012.00021.
[38]
Yan, F.; Wang, X.M.; Pan, C.; Ma, Q.M. Down-regulation of extracellular signal-regulated kinase 1/2 activity in P-glycoprotein-mediated multidrug resistant hepatocellular carcinoma cells. World J. Gastroenterol. 2009, 15, 1443–1451, doi:10.3748/wjg.15.1443.
[39]
Nag, A.; Bagchi, S.; Raychaudhuri, P. Cul4A physically associates with MDM2 and participates in the proteolysis of p53. Cancer Res. 2004, 64, 8152–8155, doi:10.1158/0008-5472.CAN-04-2598.
[40]
Bondar, T.; Kalinina, A.; Khair, L.; Kopanja, D.; Nag, A.; Bagchi, S.; Raychaudhuri, P. Cul4A and DDB1 associate with Skp2 to target p27Kip1 for proteolysis involving the COP9 signalosome. Mol. Cell. Biol. 2006, 26, 2531–2539.
[41]
Nishitani, H.; Shiomi, Y.; Iida, H.; Michishita, M.; Takami, T.; Tsurimoto, T. CDK inhibitor p21 is degraded by a proliferating cell nuclear antigen-coupled Cul4-DDB1Cdt2 pathway during S phase and after UV irradiation. J. Biol. Chem. 2008, 283, 29045–29052, doi:10.1074/jbc.M806045200.
[42]
El-Mahdy, M.A.; Zhu, Q.; Wang, Q.E.; Wani, G.; Praetorius-Ibba, M.; Wani, A.A. Cullin 4A-mediated proteolysis of DDB2 protein at DNA damage sites regulates in vivo lesion recognition by XPC. J. Biol. Chem. 2006, 281, 13404–13411.
[43]
Ang, Y.S.; Tsai, S.Y.; Lee, D.F.; Monk, J.; Su, J.; Ratnakumar, K.; Ding, J.; Ge, Y.; Darr, H.; Chang, B.; et al. Wdr5 mediates self-renewal and reprogramming via the embryonic stem cell core transcriptional network. Cell 2011, 145, 183–197, doi:10.1016/j.cell.2011.03.003.
