A Twist-Snail Axis Critical for TrkB-Induced Epithelial-Mesenchymal Transition-Like Transformation, Anoikis Resistance, and Metastasis

Abstract

In a genomewide anoikis suppression screen for metastasis genes, we previously identified the neurotrophic receptor tyrosine kinase TrkB. In mouse xenografts, activated TrkB caused highly invasive and metastatic tumors. Here, we describe that TrkB also induces a strong morphological transformation, resembling epithelial-mesenchymal transition (EMT). This required TrkB kinase activity, a functional mitogen-activated protein kinase pathway, suppression of E-cadherin, and induction of Twist, a transcription factor contributing to EMT and metastasis. RNA interference (RNAi)-mediated Twist depletion blocked TrkB-induced EMT-like transformation, anoikis suppression, and growth of tumor xenografts. By searching for essential effectors of TrkB-Twist signaling, we found that Twist induces Snail, another EMT regulator associated with poor cancer prognosis. Snail depletion impaired EMT-like transformation and anoikis suppression induced by TrkB, but in contrast to Twist depletion, it failed to inhibit tumor growth. Instead, Snail RNAi specifically impaired the formation of lung metastases. Epistasis experiments suggested that Twist acts upstream from Snail. Our results demonstrate that TrkB signaling activates a Twist-Snail axis that is critically involved in EMT-like transformation, tumorigenesis, and metastasis. Moreover, our data shed more light on the epistatic relationship between Twist and Snail, two key transcriptional regulators of EMT and metastasis.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://mcb.asm.org/ .

ACKNOWLEDGMENTS

We thank M. Voetel, S. Greven, H. Grimminck, and all animal caretakers for their excellent technical help with the in vivo experiments; L. Brocks and L. Oomen for advice with image analysis; and the experimental animal pathology department and J. Zevenhoven for help with the pathological analysis. We thank M. Heimerikx, M. Nieuwland, J. de Ronde, D. Sie, and R. Kerkhoven for performing microarray analysis; C. Desmet for communication of results prior to publication; N. Armstrong for statistical advice; A. Pfauth and F. van Diepen for help with fluorescence-activated cell sorting; and S. Ellenbroek and S. Mertens for advice on RAC activity assays. We thank all members of the Peeper laboratory for their valuable input and A. Prieur and C. Hömig-Hölzel for critical reading of the manuscript. We thank K. Becker for the Snail antibody, C. Niessen for the E-cadherin plasmid, A. Munoz for the Snail-HA plasmid, and R. Weinberg for the Twist plasmid.

This work was supported by the Dutch Cancer Society (KWF) grant to M.A.S. and D.S.P. and a European Union FP6 grant to T.R.G. and D.S.P. D.S.P. is also supported by the European Molecular Biology Organization (EMBO) Young Investigator program and by a KWF Queen Wilhelmina program grant.