Loss of giant obscurins alters breast epithelial cell mechanosensing of matrix stiffness.

Kimberly M Stroka,Bin Sheng Wong,Marey Shriver,Konstantinos Konstantopoulos,Aikaterini Kontrogianni-Konstantopoulos,Denis Wirtz,Jude M Phillip

doi:10.18632/oncotarget.10997

Abstract

Obscurins are a family of RhoGEF-containing proteins with tumor and metastasis suppressing roles in breast epithelium. Downregulation of giant obscurins in normal breast epithelial cells leads to reduced levels of active RhoA and of its downstream effectors. Herein, we elucidate how depletion of giant obscurins affects the response of breast epithelial cells to changes in the mechanical properties of the microenvironment. We find that knockdown of obscurins increases cell morphodynamics, migration speed, and diffusivity on polyacrylamide gels of ≥ 1 kPa, presumably by decreasing focal adhesion area and density as well as cell traction forces. Depletion of obscurins also increases cell mechanosensitivity on soft (0.4–4 kPa) surfaces. Similar to downregulation of obscurins, pharmacological inhibition of Rho kinase in breast epithelial cells increases migration and morphodynamics, suggesting that suppression of Rho kinase activity following obscurin knockdown can account for alterations in morphodynamics and migration. In contrast, inhibition of myosin light chain kinase reduces morphodynamics and migration, suggesting that temporal changes in cell shape are required for efficient migration. Collectively, downregulation of giant obscurins facilitates cell migration through heterogeneous microenvironments of varying stiffness by altering cell mechanobiology.

Highlights

As tumor cells spread during metastasis, they must navigate through complex physical and biochemical microenvironments [1]
We demonstrate that loss of giant obscurins alters cell morphology, increases morphodynamics and mechanosensitivity, and affects focal adhesion morphology and traction forces
Our results indicate that loss of giant obscurins facilitates cell migration through heterogeneous microenvironments of varying stiffness by altering cell mechanobiology via RhoA-mediated effects

Summary

Introduction

As tumor cells spread during metastasis, they must navigate through complex physical and biochemical microenvironments [1]. The stiffness of the cellular microenvironment varies throughout different tissues in vivo [2], and influences cell migration via alterations in cell signaling pathways [3,4,5,6,7]. Tissue stiffness increases in many cancers [2, 8], likely due to the alterations in extracellular matrix (ECM) composition during tumor growth [9]. Metastasizing tumor cells must possess the ability to migrate along substrates and through matrices of a wide range of stiffnesses. It is mostly unknown what genetic changes allow cells to alter their mechanobiology and respond to these varying mechanical properties of the microenvironment during metastasis. Recent findings from our labs have implicated giant obscurins in breast cancer progression and metastasis [10,11,12], and here we explore their role in cell mechanobiology and mechanosensing

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