Differential expression of Ran-binding protein 2 (RanBP2) and Targeting Protein for Xklp2 (TPX2) in head and neck squamous cell carcinoma cells
DOI:
https://doi.org/10.14456/gag.2019.7Keywords:
Head and neck cancer; RanGTPase; RanBP2; TPX2Abstract
RanGTPase is overexpressed in various human cancer cell lines. RanGTPase-related genes are essential for viability of head and neck cancer cell lines. However, it is currently unclear whether RanGTPase may play a role in metastasis of head and neck cancer. This study aimed to examine mRNA and protein expression of RanGTPase in primary and metastatic head and neck cancer cell lines using real-time PCR and Western blot. Three pairs of head and neck cancer cell lines were used. HN18 HN30, HN4 were derived from primary lesions whereas HN17 HN31, HN12 were derived from metastasis lesions which belong to same patients, respectively. They were compared to HaCat which is an immortalized non-tumorigenic human keratinocyte cell line. Cells were cultured and total RNA was extracted and converted to complementary DNA. mRNA expression of Ran, KPNB1, RanBP2, and TPX2 was monitored using real-time PCR. GAPDH was used as an internal control. Western blots were also performed to examine protein expression of Ran, Importin-beta, RanBP2 and TPX2. Realtime PCR data showed that mRNA expression of Ran, KPNB1, RanBP2 and TPX2 in primary and metastatic head and neck cancer cell lines was significantly different from that of HaCat at p value of 0.0079, 0.0005, 0.0074 and 0.0039, respectively. Furthermore, expression of mRNA transcript of Ran and TPX2 in metastatic head and neck cancer cell lines was higher than that of primary cancer cells. However, protein expression of RanBP2 and TPX2 was found to be higher in metastatic cancer cells compared to their primary counterparts. These findings suggest that expression of mRNA transcripts and protein of RanGTPase network was significantly different from that of normal cells. Particularly, RanBP2, and TPX2 expression was elevated in metastatic head and neck cancer cell lines. Further studies are required to understand its functional role in metastatic head and neck cancer.
References
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Caceres-Gorriti, K.Y., E. Carmona, V. Barres, K. Rahimi, I.J. Letourneau, P.N. Tonin, D. Provencher, and A.M. Mes-Masson (2014) RAN nucleo-cytoplasmic transport and mitotic spindle assembly partners XPO7 and TPX2 are new prognostic biomarkers in serous epithelial ovarian cancer. PloS one. 9:e91000.
Cardinali, M., H. Pietraszkiewicz, J.F. Ensley, and K.C. Robbins (1995) Tyrosine phosphorylation as a marker for aberrantly regulated growth-promoting pathways in cell lines derived from head and neck malignancies. International journal of cancer. 61:98-103.
Chang, H., J. Wang, Y. Tian, J. Xu, X. Gou, and J. Cheng (2012) The TPX2 gene is a promising diagnostic and therapeutic target for cervical cancer. Oncology reports. 27:1353-1359.
Clarke, P.R., and C. Zhang (2001) Ran GTPase: a master regulator of nuclear structure and function during the eukaryotic cell division cycle? Trends in cell biology. 11:366-371.
Dawlaty, M.M., L. Malureanu, K.B. Jeganathan, E. Kao, C. Sustmann, S. Tahk, K. Shuai, R. Grosschedl, and J.M. van Deursen (2008) Resolution of sister centromeres requires RanBP2-mediated SUMOylation of topoisomerase IIalpha. Cell. 133:103-115.
Deng, L., Y. Shang, S. Guo, C. Liu, L. Zhou, Y. Sun, Y. Nie, D. Fan, Y. Lu, and X. Guo (2014) Ran GTPase protein promotes metastasis and invasion in pancreatic cancer by deregulating the expression of AR and CXCR4. Cancer biology & therapy. 15:1087-1093.
Fan, H., Y. Lu, H. Qin, Y. Zhou, Y. Gu, J. Zhou, X. Wang, and D. Fan (2013) High Ran level is correlated with poor prognosis in patients with colorectal cancer. International journal of clinical oncology. 18:856-863.
Felix, R.S., G.W. Colleoni, O.L. Caballero, M. Yamamoto, M.S. Almeida, V.C. Andrade, L. Chauffaille Mde, W.A. Silva, Jr., M.D. Begnami, F.A. Soares, A.J. Simpson, M.A. Zago, and A.L. Vettore (2009) SAGE analysis highlights the importance of p53csv, ddx5, mapkapk2 and ranbp2 to multiple myeloma tumorigenesis. Cancer letters. 278:41-48.
Gama-Carvalho, M., and M. Carmo-Fonseca (2001) The rules and roles of nucleocytoplasmic shuttling proteins. FEBS letters. 498:157-163.
Hill, R., B. Cautain, N. de Pedro, and W. Link (2014) Targeting nucleocytoplasmic transport in cancer therapy. Oncotarget. 5:11-28.
Iamaroon, A., K. Pattanaporn, S. Pongsiriwet, S. Wanachantararak, S. Prapayasatok, S. Jittidecharaks, I. Chitapanarux, and V. Lorvidhaya (2004) Analysis of 587 cases of oral squamous cell carcinoma in northern Thailand with a focus on young people. International journal of oral and maxillofacial surgery. 33:84-88.
Kalab, P., and R. Heald (2008) The RanGTP gradient - a GPS for the mitotic spindle. Journal of cell science. 121:1577-1586.
Kodama, M., T. Kodama, J.Y. Newberg, H. Katayama, M. Kobayashi, S.M. Hanash, K. Yoshihara, Z. Wei, J.C. Tien, R. Rangel, K. Hashimoto, S. Mabuchi, K. Sawada, T. Kimura, N.G. Copeland, and N.A. Jenkins (2017) In vivo loss-of-function screens identify KPNB1 as a new druggable oncogene in epithelial ovarian cancer. Proceedings of the National Academy of Sciences of the United States of America. 114:E7301-e7310.
Koontongkaew, S., P. Amornphimoltham, P. Monthanpisut, T. Saensuk, and M. Leelakriangsak (2012) Fibroblasts and extracellular matrix differently modulate MMP activation by primary and metastatic head and neck cancer cells. Medical oncology (Northwood, London, England). 29:690-703.
Leemans, C.R., B.J. Braakhuis, and R.H. Brakenhoff (2011) The molecular biology of head and neck cancer. Nature reviews. Cancer. 11:9-22.
Liu, Q., K. Tu, H. Zhang, X. Zheng, Y. Yao, and Q. Liu (2015) TPX2 as a novel prognostic biomarker for hepatocellular carcinoma. Hepatology research : the official journal of the Japan Society of Hepatology. 45:906-918.
Liu, Q., P. Yang, K. Tu, H. Zhang, X. Zheng, Y. Yao, and Q. Liu (2014) TPX2 knockdown suppressed hepatocellular carcinoma cell invasion via inactivating AKT signaling and inhibiting MMP2 and MMP9 expression. Chinese journal of cancer research = Chung-kuo yen cheng yen chiu. 26:410-417.
Livak, K.J., and T.D. Schmittgen (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods (San Diego, Calif.). 25:402-408.
Lu, T., Z. Bao, Y. Wang, L. Yang, B. Lu, K. Yan, S. Wang, H. Wei, Z. Zhang, and G. Cui (2016) Karyopherinbeta1 regulates proliferation of human glioma cells via Wnt/beta-catenin pathway. Biochemical and biophysical research communications. 478:1189-1197.
Ly, T.K., J. Wang, R. Pereira, K.S. Rojas, X. Peng, Q. Feng, R.A. Cerione, and K.F. Wilson (2010) Activation of the Ran GTPase is subject to growth factor regulation and can give rise to cellular transformation. The Journal of biological chemistry. 285:5815-5826.
Martens-de Kemp, S.R., R. Nagel, M. Stigter-van Walsum, I.H. van der Meulen, V.W. van Beusechem, B.J. Braakhuis, and R.H. Brakenhoff (2013) Functional genetic screens identify genes essential for tumor cell survival in head and neck and lung cancer. Clinical cancer research : an official journal of the American Association for Cancer Research. 19:1994-2003.
Packham, S., D. Warsito, Y. Lin, S. Sadi, R. Karlsson, B. Sehat, and O. Larsson (2015) Nuclear translocation of IGF-1R via p150(Glued) and an importin-beta/RanBP2-dependent pathway in cancer cells. Oncogene. 34:2227-2238.
Rensen, W.M., R. Mangiacasale, M. Ciciarello, and P. Lavia (2008) The GTPase Ran: regulation of cell life and potential roles in cell transformation. Frontiers in bioscience : a journal and virtual library. 13:4097-4121.
Roepman, P., L.F. Wessels, N. Kettelarij, P. Kemmeren, A.J. Miles, P. Lijnzaad, M.G. Tilanus, R. Koole, G.J. Hordijk, P.C. van der Vliet, M.J. Reinders, P.J. Slootweg, and F.C. Holstege (2005) An expression profile for diagnosis of lymph node metastases from primary head and neck squamous cell carcinomas. Nature genetics. 37:182-186.
Rosenthal, E.L., and L.M. Matrisian (2006) Matrix metalloproteases in head and neck cancer. Head & neck. 28:639-648.
Stadler, M.E., M.R. Patel, M.E. Couch, and D.N. Hayes (2008) Molecular biology of head and neck cancer: risks and pathways. Hematology/oncology clinics of North America. 22:1099-1124, vii.
Supokawej, A., P. Kheolamai, K. Nartprayut, U.P. Y, S. Manochantr, M. Chayosumrit, and S. Issaragrisil (2013) Cardiogenic and myogenic gene expression in mesenchymal stem cells after 5-azacytidine treatment. Turkish journal of haematology : official journal of Turkish Society of Haematology. 30:115-121.
van der Watt, P.J., C.P. Maske, D.T. Hendricks, M.I. Parker, L. Denny, D. Govender, M.J. Birrer, and V.D. Leaner (2009) The Karyopherin proteins, Crm1 and Karyopherin beta1, are overexpressed in cervical cancer and are critical for cancer cell survival and proliferation. International journal of cancer. 124:1829-1840.
Warner, S.L., B.J. Stephens, S. Nwokenkwo, G. Hostetter, A. Sugeng, M. Hidalgo, J.M. Trent, H. Han, and D.D. Von Hoff (2009) Validation of TPX2 as a potential therapeutic target in pancreatic cancer cells. Clinical cancer research : an official journal of the American Association for Cancer Research. 15:6519-6528.
Wei, P., N. Zhang, Y. Xu, X. Li, D. Shi, Y. Wang, D. Li, and S. Cai (2013) TPX2 is a novel prognostic marker for the growth and metastasis of colon cancer. Journal of translational medicine. 11:313.
Xia, F., P.M. Canovas, T.M. Guadagno, and D.C. Altieri (2008a) A survivin-ran complex regulates spindle formation in tumor cells. Molecular and cellular biology. 28:5299-5311.
Xia, F., C.W. Lee, and D.C. Altieri (2008b) Tumor cell dependence on Ran-GTP-directed mitosis. Cancer research. 68:1826-1833.
Yang, L., B. Hu, Y. Zhang, S. Qiang, J. Cai, W. Huang, C. Gong, T. Zhang, S. Zhang, P. Xu, X. Wu, and J. Liu (2015) Suppression of the nuclear transporter-KPNbeta1 expression inhibits tumor proliferation in hepatocellular carcinoma. Medical oncology (Northwood, London, England). 32:128.
Yu, T., Y. Wu, J.I. Helman, Y. Wen, C. Wang, and L. Li (2011) CXCR4 promotes oral squamous cell carcinoma migration and invasion through inducing expression of MMP-9 and MMP-13 via the ERK signaling pathway. Molecular cancer research : MCR. 9:161-172.
Albert, S., M.E. Riveiro, C. Halimi, M. Hourseau, A. Couvelard, M. Serova, B. Barry, E. Raymond, and S. Faivre (2013) Focus on the role of the CXCL12/CXCR4 chemokine axis in head and neck squamous cell carcinoma. Head & neck. 35:1819-1828.
Argiris, A., M.V. Karamouzis, D. Raben, and R.L. Ferris (2008) Head and neck cancer. Lancet (London, England). 371:1695-1709.
Caceres-Gorriti, K.Y., E. Carmona, V. Barres, K. Rahimi, I.J. Letourneau, P.N. Tonin, D. Provencher, and A.M. Mes-Masson (2014) RAN nucleo-cytoplasmic transport and mitotic spindle assembly partners XPO7 and TPX2 are new prognostic biomarkers in serous epithelial ovarian cancer. PloS one. 9:e91000.
Cardinali, M., H. Pietraszkiewicz, J.F. Ensley, and K.C. Robbins (1995) Tyrosine phosphorylation as a marker for aberrantly regulated growth-promoting pathways in cell lines derived from head and neck malignancies. International journal of cancer. 61:98-103.
Chang, H., J. Wang, Y. Tian, J. Xu, X. Gou, and J. Cheng (2012) The TPX2 gene is a promising diagnostic and therapeutic target for cervical cancer. Oncology reports. 27:1353-1359.
Clarke, P.R., and C. Zhang (2001) Ran GTPase: a master regulator of nuclear structure and function during the eukaryotic cell division cycle? Trends in cell biology. 11:366-371.
Dawlaty, M.M., L. Malureanu, K.B. Jeganathan, E. Kao, C. Sustmann, S. Tahk, K. Shuai, R. Grosschedl, and J.M. van Deursen (2008) Resolution of sister centromeres requires RanBP2-mediated SUMOylation of topoisomerase IIalpha. Cell. 133:103-115.
Deng, L., Y. Shang, S. Guo, C. Liu, L. Zhou, Y. Sun, Y. Nie, D. Fan, Y. Lu, and X. Guo (2014) Ran GTPase protein promotes metastasis and invasion in pancreatic cancer by deregulating the expression of AR and CXCR4. Cancer biology & therapy. 15:1087-1093.
Fan, H., Y. Lu, H. Qin, Y. Zhou, Y. Gu, J. Zhou, X. Wang, and D. Fan (2013) High Ran level is correlated with poor prognosis in patients with colorectal cancer. International journal of clinical oncology. 18:856-863.
Felix, R.S., G.W. Colleoni, O.L. Caballero, M. Yamamoto, M.S. Almeida, V.C. Andrade, L. Chauffaille Mde, W.A. Silva, Jr., M.D. Begnami, F.A. Soares, A.J. Simpson, M.A. Zago, and A.L. Vettore (2009) SAGE analysis highlights the importance of p53csv, ddx5, mapkapk2 and ranbp2 to multiple myeloma tumorigenesis. Cancer letters. 278:41-48.
Gama-Carvalho, M., and M. Carmo-Fonseca (2001) The rules and roles of nucleocytoplasmic shuttling proteins. FEBS letters. 498:157-163.
Hill, R., B. Cautain, N. de Pedro, and W. Link (2014) Targeting nucleocytoplasmic transport in cancer therapy. Oncotarget. 5:11-28.
Iamaroon, A., K. Pattanaporn, S. Pongsiriwet, S. Wanachantararak, S. Prapayasatok, S. Jittidecharaks, I. Chitapanarux, and V. Lorvidhaya (2004) Analysis of 587 cases of oral squamous cell carcinoma in northern Thailand with a focus on young people. International journal of oral and maxillofacial surgery. 33:84-88.
Kalab, P., and R. Heald (2008) The RanGTP gradient - a GPS for the mitotic spindle. Journal of cell science. 121:1577-1586.
Kodama, M., T. Kodama, J.Y. Newberg, H. Katayama, M. Kobayashi, S.M. Hanash, K. Yoshihara, Z. Wei, J.C. Tien, R. Rangel, K. Hashimoto, S. Mabuchi, K. Sawada, T. Kimura, N.G. Copeland, and N.A. Jenkins (2017) In vivo loss-of-function screens identify KPNB1 as a new druggable oncogene in epithelial ovarian cancer. Proceedings of the National Academy of Sciences of the United States of America. 114:E7301-e7310.
Koontongkaew, S., P. Amornphimoltham, P. Monthanpisut, T. Saensuk, and M. Leelakriangsak (2012) Fibroblasts and extracellular matrix differently modulate MMP activation by primary and metastatic head and neck cancer cells. Medical oncology (Northwood, London, England). 29:690-703.
Leemans, C.R., B.J. Braakhuis, and R.H. Brakenhoff (2011) The molecular biology of head and neck cancer. Nature reviews. Cancer. 11:9-22.
Liu, Q., K. Tu, H. Zhang, X. Zheng, Y. Yao, and Q. Liu (2015) TPX2 as a novel prognostic biomarker for hepatocellular carcinoma. Hepatology research : the official journal of the Japan Society of Hepatology. 45:906-918.
Liu, Q., P. Yang, K. Tu, H. Zhang, X. Zheng, Y. Yao, and Q. Liu (2014) TPX2 knockdown suppressed hepatocellular carcinoma cell invasion via inactivating AKT signaling and inhibiting MMP2 and MMP9 expression. Chinese journal of cancer research = Chung-kuo yen cheng yen chiu. 26:410-417.
Livak, K.J., and T.D. Schmittgen (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods (San Diego, Calif.). 25:402-408.
Lu, T., Z. Bao, Y. Wang, L. Yang, B. Lu, K. Yan, S. Wang, H. Wei, Z. Zhang, and G. Cui (2016) Karyopherinbeta1 regulates proliferation of human glioma cells via Wnt/beta-catenin pathway. Biochemical and biophysical research communications. 478:1189-1197.
Ly, T.K., J. Wang, R. Pereira, K.S. Rojas, X. Peng, Q. Feng, R.A. Cerione, and K.F. Wilson (2010) Activation of the Ran GTPase is subject to growth factor regulation and can give rise to cellular transformation. The Journal of biological chemistry. 285:5815-5826.
Martens-de Kemp, S.R., R. Nagel, M. Stigter-van Walsum, I.H. van der Meulen, V.W. van Beusechem, B.J. Braakhuis, and R.H. Brakenhoff (2013) Functional genetic screens identify genes essential for tumor cell survival in head and neck and lung cancer. Clinical cancer research : an official journal of the American Association for Cancer Research. 19:1994-2003.
Packham, S., D. Warsito, Y. Lin, S. Sadi, R. Karlsson, B. Sehat, and O. Larsson (2015) Nuclear translocation of IGF-1R via p150(Glued) and an importin-beta/RanBP2-dependent pathway in cancer cells. Oncogene. 34:2227-2238.
Rensen, W.M., R. Mangiacasale, M. Ciciarello, and P. Lavia (2008) The GTPase Ran: regulation of cell life and potential roles in cell transformation. Frontiers in bioscience : a journal and virtual library. 13:4097-4121.
Roepman, P., L.F. Wessels, N. Kettelarij, P. Kemmeren, A.J. Miles, P. Lijnzaad, M.G. Tilanus, R. Koole, G.J. Hordijk, P.C. van der Vliet, M.J. Reinders, P.J. Slootweg, and F.C. Holstege (2005) An expression profile for diagnosis of lymph node metastases from primary head and neck squamous cell carcinomas. Nature genetics. 37:182-186.
Rosenthal, E.L., and L.M. Matrisian (2006) Matrix metalloproteases in head and neck cancer. Head & neck. 28:639-648.
Stadler, M.E., M.R. Patel, M.E. Couch, and D.N. Hayes (2008) Molecular biology of head and neck cancer: risks and pathways. Hematology/oncology clinics of North America. 22:1099-1124, vii.
Supokawej, A., P. Kheolamai, K. Nartprayut, U.P. Y, S. Manochantr, M. Chayosumrit, and S. Issaragrisil (2013) Cardiogenic and myogenic gene expression in mesenchymal stem cells after 5-azacytidine treatment. Turkish journal of haematology : official journal of Turkish Society of Haematology. 30:115-121.
van der Watt, P.J., C.P. Maske, D.T. Hendricks, M.I. Parker, L. Denny, D. Govender, M.J. Birrer, and V.D. Leaner (2009) The Karyopherin proteins, Crm1 and Karyopherin beta1, are overexpressed in cervical cancer and are critical for cancer cell survival and proliferation. International journal of cancer. 124:1829-1840.
Warner, S.L., B.J. Stephens, S. Nwokenkwo, G. Hostetter, A. Sugeng, M. Hidalgo, J.M. Trent, H. Han, and D.D. Von Hoff (2009) Validation of TPX2 as a potential therapeutic target in pancreatic cancer cells. Clinical cancer research : an official journal of the American Association for Cancer Research. 15:6519-6528.
Wei, P., N. Zhang, Y. Xu, X. Li, D. Shi, Y. Wang, D. Li, and S. Cai (2013) TPX2 is a novel prognostic marker for the growth and metastasis of colon cancer. Journal of translational medicine. 11:313.
Xia, F., P.M. Canovas, T.M. Guadagno, and D.C. Altieri (2008a) A survivin-ran complex regulates spindle formation in tumor cells. Molecular and cellular biology. 28:5299-5311.
Xia, F., C.W. Lee, and D.C. Altieri (2008b) Tumor cell dependence on Ran-GTP-directed mitosis. Cancer research. 68:1826-1833.
Yang, L., B. Hu, Y. Zhang, S. Qiang, J. Cai, W. Huang, C. Gong, T. Zhang, S. Zhang, P. Xu, X. Wu, and J. Liu (2015) Suppression of the nuclear transporter-KPNbeta1 expression inhibits tumor proliferation in hepatocellular carcinoma. Medical oncology (Northwood, London, England). 32:128.
Yu, T., Y. Wu, J.I. Helman, Y. Wen, C. Wang, and L. Li (2011) CXCR4 promotes oral squamous cell carcinoma migration and invasion through inducing expression of MMP-9 and MMP-13 via the ERK signaling pathway. Molecular cancer research : MCR. 9:161-172.
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