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JunB promotes cell invasion and angiogenesis in VHL-defective renal cell carcinoma

Abstract

Inactivation of the von Hippel–Lindau (VHL) tumor-suppressor gene causes both hereditary and sporadic clear-cell renal-cell carcinoma (ccRCC). Although the best-characterized function of the VHL protein (pVHL) is regulation of hypoxia-inducible factor-α (HIFα), pVHL also controls the development of pheochromocytoma through HIF-independent pathways by regulating JunB. However, it is largely unknown how these pathways contribute to the development and progression of ccRCC. In the present study, we confirmed that JunB was upregulated in VHL-defective ccRCC specimens by immunostaining. Short-hairpin RNA (shRNA)-mediated knockdown of JunB in 786-O and A498 VHL null ccRCC cells suppressed their invasiveness. In addition, JunB knockdown significantly repressed tumor growth and microvessel density in xenograft tumor assays. Conversely, forced expression of wild-type, but not dimerization-defective, JunB in a VHL-restored 786-O subclone promoted invasion in vitro and tumor growth and vessel formation in vivo. Quantitative PCR array analysis revealed that JunB regulated multiple genes relating to tumor invasion and angiogenesis such as matrix metalloproteinase-2 (MMP-2), MMP-9 and chemokine (C-C motif) ligand-2 (CCL2) in 786-O cells. JunB knockdown in these cells reduced the proteolytic activity of both MMPs in gelatin zymography and the amount of CCL2 in the culture supernatant. Moreover, shRNA-mediated knockdown of MMP-2 or inhibition of CCL2 activity with a neutralizing antibody repressed xenograft tumor growth and angiogenesis. Collectively, these results suggest that JunB promotes tumor invasiveness and enhances angiogenesis in VHL-defective ccRCCs.

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Acknowledgements

We thank Dr William G Kaelin, Jr, for critical comments and for providing us with the different RCC cell lines. We appreciate all members of Cancer Research Courses for Integrated Research Training at Kyoto University Graduate School of Medicine for helpful advice and discussion. We also thank Tomoko Matsushita, Megumi Kuraguchi and Yuko Fujieda for technical assistance. Grant support: Grant-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology, Japan.

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Correspondence to E Nakamura.

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Kanno, T., Kamba, T., Yamasaki, T. et al. JunB promotes cell invasion and angiogenesis in VHL-defective renal cell carcinoma. Oncogene 31, 3098–3110 (2012). https://doi.org/10.1038/onc.2011.475

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Keywords

  • renal-cell carcinoma
  • VHL gene
  • JunB
  • invasion
  • angiogenesis

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