Vinculin Force Sensor Detects Tumor-Osteocyte Interactions

Fangjia Li,Andrew Reeser,Bai-Yan Li,Yue Wang,Xinyu Zhao,Andy Chen,Hiroki Yokota,Yao Fan,Jing Liu,Shengzhi Liu,Divya Kota,Rahul Prakash

doi:10.1038/s41598-019-42132-x

Abstract

This study utilized a Förster resonance energy transfer (FRET)-based molecular tension sensor and live cell imaging to evaluate the effect of osteocytes, a mechanosensitive bone cell, on the migratory behavior of tumor cells. Two cell lines derived from MDA-MB-231 breast cancer cells were transfected with the vinculin tension sensor to quantitatively evaluate the force in focal adhesions of the tumor cell. Tumor cells treated with MLO-A5 osteocyte-conditioned media (CM) decreased the tensile forces in their focal adhesions and decreased their migratory potential. Tumor cells treated with media derived from MLO-A5 cells exposed to fluid flow-driven shear stress (FFCM) increased the tensile forces and increased migratory potential. Focal adhesion tension in tumor cells was also affected by distance from MLO-A5 cells when the two cells were co-cultured, where tumor cells close to MLO-A5 cells exhibited lower tension and decreased cell motility. Overall, this study demonstrates that focal adhesion tension is involved in altered migratory potential of tumor cells, and tumor-osteocyte interactions decrease the tension and motility of tumor cells.

Highlights

Breast cancer is one of the most common cancers among women, and almost 30% of primary breast tumors are reported to metastasize to other organs[1]
The results indicate that migratory behaviors of tumor cells are closely linked to tensile forces at focal adhesions, and in response to mechanical stimulation osteocytes toggle their regulation of migration in tumor cells
The vinculin tension sensor (VinTS) is a tension sensitive sensor that consists of the head and tail domains of vinculin with the elastic Förster resonance energy transfer (FRET) module inserted between them (Fig. 1A), and the sensor exhibits low(high) FRET efficiency when it undergoes high tensions (Fig. 1A,B); the FRET map of the TMD cell transfected with the VinTS probe has shown low and varying FRET efficiency for individual focal adhesion sites (Fig. 1B)

Summary

Introduction

Breast cancer is one of the most common cancers among women, and almost 30% of primary breast tumors are reported to metastasize to other organs[1]. We hypothesized that in the presence and absence of mechanical stimulation, osteocytes interact differently with breast cancer cells via molecular machinery at focal adhesions To test this hypothesis, we employed a molecular tension sensor as well as live cell imaging to investigate the force dynamics of the focal adhesion during the regulatory migration behaviors. The TSMod includes an elastic peptide linker of 40 amino acids and two fluorophores (mTFP1 and Venus) on each end, in which extension of the linker changes FRET between fluorescent proteins This vinculin tension senor is recruited to focal adhesions (FAs) which connect integrin and the actin filaments[11,12] and are sensitive to mechanical stimulation and induce signaling for cell migration through extracellular matrices[13]. This study suggests a critical role for mechanotransduction of bone in tumor metastasis and a novel target of interactions in bone metastatic tumor cells

Methods

Results

Conclusion