1 Department of Urology, The First Affiliated Hospital of Xiamen University, Xiamen, China;
2 Department of Urology, Ningbo Yinzhou Second Hospital, Ningbo, China
Pygopus 2 promotes kidney cancer OS-RC-2 cells proliferation and invasion in vitro and in vivo
1 Department of Urology, The First Affiliated Hospital of Xiamen University, Xiamen, China;
2 Department of Urology, Ningbo Yinzhou Second Hospital, Ningbo, China
摘要 Objective: Human Pygopus 2 (Pygo2) was recently discovered to be a component of the Wnt signaling pathway required for β-catenin/Tcf-mediated transcription. But the role of Pygo2 in malignant cell proliferation and invasion has not yet been determined. Methods: Lentivirus-mediated small interfering RNA (siRNA) and vector-based overexpression were used to study the function of Pygo2 in OS-RC-2 cells. The resulted cells were subject to Western blotting assay, MTT assay, colony formation and cell invasion assays. Furthermore, renal cell carcinoma (RCC) models were established in BALB/c nude mice inoculated with OS-RC-2 cells. Immunohistochemistry (IHC) staining of matrix metalloproteinase-7 (MMP-7), matrix metalloproteinase-9 (MMP-9) and vascular endothelial growth factor (VEGF) was performed in tumor tissue. Results:Pygo2 gene was successful knocked down and overexpressed in RCC OS-RC-2 cells by using an shRNA and overexpressing vector, respectively. Overexpression of Pygo2 effectively promoted cell proliferation, colony formation and invasion in vitro. Knockdown of Pygo2 obviously inhibited xenograft tumor growth in nude mice. In addition, overexpression of Pygo2 increased the levels of MMP-7, MMP-9 and VEGF in the xenograft tumors. Conclusion:Pygo2 has a role in promoting cell proliferation, invasion and metastasis, and may regulate angiogenesis via the Wnt/β-catenin signaling pathway.
Abstract: Objective: Human Pygopus 2 (Pygo2) was recently discovered to be a component of the Wnt signaling pathway required for β-catenin/Tcf-mediated transcription. But the role of Pygo2 in malignant cell proliferation and invasion has not yet been determined. Methods: Lentivirus-mediated small interfering RNA (siRNA) and vector-based overexpression were used to study the function of Pygo2 in OS-RC-2 cells. The resulted cells were subject to Western blotting assay, MTT assay, colony formation and cell invasion assays. Furthermore, renal cell carcinoma (RCC) models were established in BALB/c nude mice inoculated with OS-RC-2 cells. Immunohistochemistry (IHC) staining of matrix metalloproteinase-7 (MMP-7), matrix metalloproteinase-9 (MMP-9) and vascular endothelial growth factor (VEGF) was performed in tumor tissue. Results:Pygo2 gene was successful knocked down and overexpressed in RCC OS-RC-2 cells by using an shRNA and overexpressing vector, respectively. Overexpression of Pygo2 effectively promoted cell proliferation, colony formation and invasion in vitro. Knockdown of Pygo2 obviously inhibited xenograft tumor growth in nude mice. In addition, overexpression of Pygo2 increased the levels of MMP-7, MMP-9 and VEGF in the xenograft tumors. Conclusion:Pygo2 has a role in promoting cell proliferation, invasion and metastasis, and may regulate angiogenesis via the Wnt/β-catenin signaling pathway.
基金资助: We thank Xiaohong Ma for statistical analysis and technical assistance. We also thank Professor Boan Li of college of Life Science of Xiamen University for contributing research reagents. This study was supported by grants (No. 2011J01254) from Natural Science Foundation of Fujian Province.
通讯作者:
Rongfu Liu
E-mail: lliurf@126.com
引用本文:
Rongfu Liu, Xiangcheng Qin, Chengyong Ji, Weixin Zeng, Yufeng Yang, Wei Tan. Pygopus 2 promotes kidney cancer OS-RC-2 cells proliferation and invasion in vitro and in vivo[J]. Asian Journal of Urology, 2015, 2(3): 151-157.
Rongfu Liu, Xiangcheng Qin, Chengyong Ji, Weixin Zeng, Yufeng Yang, Wei Tan. Pygopus 2 promotes kidney cancer OS-RC-2 cells proliferation and invasion in vitro and in vivo. Asian Journal of Urology, 2015, 2(3): 151-157.
Hoffmans R, Städeli R, Basler K. Pygopus and legless provide essential transcriptional coactivator functions to armadillo/-beta-catenin. Curr Biol 2005;15:1207-71.
[2]
Polakis P. Wnt signaling in cancer. Cold Spring Harb Perspect Biol 2012;4(5). http://dx.doi.org/10.1101/cshperspect.a008052.
[3]
Mieszczanek J, de la Roche M, Bienz M. A role of Pygopus as an anti-repressor in facilitating Wnt-dependent transcription. Proc Natl Acad Sci U S A 2008;105:19324-4.
[4]
Miller TC, Rutherford TJ, Johnson CM, Fiedler M, Bienz M. Allosteric remodelling of the histone H3 binding pocket in the Pygo2 PHD finger triggered by its binding to the B9L/BCL9 cofactor. J Mol Biol 2010;401:969-94.
[5]
Kramps T, Peter O, Brunner E, Nellen D, Froesch B, Chatterjee S, et al. Wnt/wingless signaling requires BCL9/legless-mediated recruitment of pygopus to the nuclear β-catenin-TCF complex. Cell 2002;109:47-70.
[6]
Städeli R, Basler K. Dissecting nuclear wingless signalling:recruitment of the transcriptional co-activator Pygopus by a chain of adaptor proteins. Mech Dev 2005;122:1171-12.
[7]
Nelson AR, Fingleton B, Rothenberg ML, Matrisian LM. Matrix metalloproteinases:biologic activity and clinical implications. J Clin Oncol 2000;18:1135-59.
[8]
Townsley FM, Cliffe A, Bienz M. Pygopus and legless target Armadillo/beta-catenin to the nucleus to enable its transcriptional co-activator function. Nat Cell Biol 2004;6:626-63.
[9]
Belenkaya TY, Han C, Standley HJ, Lin X, Houston DW, Heasman J, et al. Pygopus encodes a nuclear protein essential for wing-less/Wnt signaling. Development 2002;129:4089-901.
[10]
Thompson B, Townsley F, Rosin-Arbesfeld R, Musisi H, Bienz M. A new nuclear component of the Wnt signalling pathway. Nat Cell Biol 2002;4:367-73.
[11]
Andrews PG, Kennedy MW, Popadiuk CM, Kao KR. Oncogenic activation of the human Pygopus2 promoter by E74-like factor-1. Mol Cancer Res 2008;6:259-96.
[12]
Lake BB, Kao KR. Pygopus is required for embryonic brain patterning in Xenopus. Dev Biol 2003;261:132-28.
[13]
Song N, Schwab KR, Patterson LT, Yamaguchi T, Lin X, Potter SS, et al. Pygopus 2 has a crucial, Wnt pathwayindependent function in lens induction. Development 2007; 134:1873-35.
[14]
Schwab KR, Patterson LT, Hartman HA, Song N, Lang RA, Lin X, et al. Pygo1 and Pygo2 roles in Wnt signaling in mammalian kidney development. Dev Cell 2007;5:15.
[15]
Popadiuk CM, Xiong J, Wells MG, Andrews PG, Dankwa K, Hirasawa K, et al. Antisense suppression of pygopus2 results in growth arrest of epithelial ovarian cancer. Clin Cancer Res 2006;12:2216-63.
[16]
Wang ZX, Chen YY, Li BA, Tan GW, Liu XY, Shen SH, et al. Decreased Pygopus 2 expression suppresses glioblastoma U251 cell growth. J Neuroomcol 2010;100:31-11.
[17]
Tycko B, Li CM, Buttyan R. The Wnt/beta-catenin pathway in Wilms tumors and prostate cancers. Curr Mol Med 2007;7:479-99.
[18]
De D, Chen A, Wu Z, Lv S, He G, Qi Y. Overexpression of Pygopus2 protects HeLa cells from vinblastine-induced apoptosis. Biol Chem 2009;390:157-75.
[19]
Folkman J. Angiogenesis in cancer, vascular, rheumatoid and other disease. Nat Med 1995;1:27-71.
[20]
Barnes EA, Kenerson HL, Mak BC, Yeung RS. The loss of tuberin promotes cell invasion through the β-catenin pathway. Am J Respir Cell Mol Biol 2010;43:617-77.
Christopher Hartman, Nikhil Gupta, David Leavitt, David Hoenig, Zeph Okeke, Arthur Smith. Advances in percutaneous stone surgery[J]. Asian Journal of Urology, 2015, 2(1): 26
-32
.
