The invadopodia scaffold protein Tks5 is required for the growth of human breast cancer cells in vitro and in vivo.

Barbara Blouw,Jian-Liang Li,Begoña Diaz,Christopher Abdullah,Sara A Courtneidge,Manishha Patel,Weon Kyoo You,Shinji Iizuka,William B Stallcup,Xiayu Huang,Ming Tan

doi:10.1371/journal.pone.0121003

Abstract

The ability of cancer cells to invade underlies metastatic progression. One mechanism by which cancer cells can become invasive is through the formation of structures called invadopodia, which are dynamic, actin-rich membrane protrusions that are sites of focal extracellular matrix degradation. While there is a growing consensus that invadopodia are instrumental in tumor metastasis, less is known about whether they are involved in tumor growth, particularly in vivo. The adaptor protein Tks5 is an obligate component of invadopodia, and is linked molecularly to both actin-remodeling proteins and pericellular proteases. Tks5 appears to localize exclusively to invadopodia in cancer cells, and in vitro studies have demonstrated its critical requirement for the invasive nature of these cells, making it an ideal surrogate to investigate the role of invadopodia in vivo. In this study, we examined how Tks5 contributes to human breast cancer progression. We used immunohistochemistry and RNA sequencing data to evaluate Tks5 expression in clinical samples, and we characterized the role of Tks5 in breast cancer progression using RNA interference and orthotopic implantation in SCID-Beige mice. We found that Tks5 is expressed to high levels in approximately 50% of primary invasive breast cancers. Furthermore, high expression was correlated with poor outcome, particularly in those patients with late relapse of stage I/II disease. Knockdown of Tks5 expression in breast cancer cells resulted in decreased growth, both in 3D in vitro cultures and in vivo. Moreover, our data also suggest that Tks5 is important for the integrity and permeability of the tumor vasculature. Together, this work establishes an important role for Tks5 in tumor growth in vivo, and suggests that invadopodia may play broad roles in tumor progression.

Highlights

Improvements in the early detection and the treatment of breast cancer have greatly reduced the mortality of the disease
We find that the high expression of Tks5α, in patients with stage I/II disease, is associated with a worse outcome, regardless of tumor subtype, for those patients with late recurrence
RNA interference experiments showed that loss of Tks5 was associated with reduced tumor cell growth, both in vitro in 3D culture, and in vivo in orthotopic sites

Summary

Introduction

Improvements in the early detection and the treatment of breast cancer have greatly reduced the mortality of the disease. Our goal is to identify the molecular mediators of invasion in breast cancer cells that may warrant efficient and targeted drug design in the future. Invadopodia were first noted in Src-transformed fibroblasts [6,7], and subsequently have been well described in many invasive human cancer cells, those derived from breast cancer, melanoma and glioblastoma. A host of proteins are localized to invadopodia, including proteases, actin regulatory proteins and signaling molecules. If any, of these molecules are selectively localized to invadopodia, . This makes it challenging to separate the functions of invadopodia from those of other actin-based structures and protrusions. Many studies define invadopodia by the simultaneous presence of ECM degradative capacity, and key actin regulators such as cofilin or cortactin. It is not currently feasible to use these criteria to study the functions of invadopodia in vivo

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