Activation of Notch1 signaling is required for β-catenin–mediated human primary melanoma progression
Klara Balint, … , Meenhard Herlyn, Zhao-Jun Liu
Klara Balint, … , Meenhard Herlyn, Zhao-Jun Liu
Published November 1, 2005
Citation Information: J Clin Invest. 2005;115(11):3166-3176. https://doi.org/10.1172/JCI25001.
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Research Article Oncology

Activation of Notch1 signaling is required for β-catenin–mediated human primary melanoma progression

Abstract

Notch is a highly conserved transmembrane receptor that determines cell fate. Notch signaling denotes cleavage of the Notch intracellular domain, its translocation to the nucleus, and subsequent activation of target gene transcription. Involvement of Notch signaling in several cancers is well known, but its role in melanoma remains poorly characterized. Here we show that the Notch1 pathway is activated in human melanoma. Blocking Notch signaling suppressed whereas constitutive activation of the Notch1 pathway enhanced primary melanoma cell growth both in vitro and in vivo yet had little effect on metastatic melanoma cells. Activation of Notch1 signaling enabled primary melanoma cells to gain metastatic capability. Furthermore, the oncogenic effect of Notch1 on primary melanoma cells was mediated by β-catenin, which was upregulated following Notch1 activation. Inhibiting β-catenin expression reversed Notch1-enhanced tumor growth and metastasis. Our data therefore suggest a β-catenin–dependent, stage-specific role for Notch1 signaling in promoting the progression of primary melanoma.

Authors

Klara Balint, Min Xiao, Chelsea C. Pinnix, Akinobu Soma, Imre Veres, Istvan Juhasz, Eric J. Brown, Anthony J. Capobianco, Meenhard Herlyn, Zhao-Jun Liu

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Figure 6

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Suppressing β-catenin reverses Notch1-induced tumor growth and metastasi...
Suppressing β-catenin reverses Notch1-induced tumor growth and metastasis. (A and B) β-catenin was examined by immunoblotting in WM35–NIC-GFP cells expressing β-cat–siRNA1, β-cat–siRNA2, β-cat–siRNA3 and control vector (A) and in WM3248–NIC-GFP cells expressing β-cat–siRNA2 (B). β-cat–si–RNA2 efficiently suppressed expression of β-catenin. β-actin was used as loading control. (C and D) MTT assay shows a decreased growth rate induced by β-cat–siRNA1 (*P < 0.01, Student’s t test) and β-cat–siRNA2 (**P < 0.001, Student’s t test) in WM35–NIC-GFP cells and β-cat–siRNA2 in WM3248-NIC-GFP cells when compared with control. Results are mean ± SD from triplicates of 3 independent experiments. (E) Induction of cell apoptosis by β-cat–siRNA2 in WM35–NIC-GFP cells. Round shape indicates cell death. Scale bar: 20 μm. Cell apoptosis was quantitatively measured by ELISA. Results are mean ± SD from triplicates of 3 independent experiments. Similar results were observed in WM3248–NIC-GFP cells. (F) 3[H]-Thymidine incorporation assay shows a decreased growth rate of WM3248 cells transfected with DN β-catenin compared with the control. Results are mean ± SD from 3 independent experiments. Inner panel shows expression of DN β-catenin protein. β-actin was used as loading control. (G) In vivo lung colony–formation assay. β-cat–siRNA-WM3248-NIC-GFP or H1UG-WM3248-NIC-GFP cells were injected intravenously into mice. After 12 weeks, lung samples were harvested and analyzed. Tumor colonies formed on the lung surface were macroscopically counted under a dissection microscope. Data are mean ± SD.