Loss of Myoferlin Redirects Breast Cancer Cell Motility towards Collective Migration

Leonithas I Volakis,Heather M Powell,Douglas A Kniss,Cosmin Mihai,Ruth Li,Meagan Bechel,William E Ackerman,Samir N Ghadiali,Thomas J Rosol,Rachel Zielinski,Christopher S Ahn,Taryn L Summerfield,Jun Li

doi:10.1371/journal.pone.0086110

Abstract

Cell migration plays a central role in the invasion and metastasis of tumors. As cells leave the primary tumor, they undergo an epithelial to mesenchymal transition (EMT) and migrate as single cells. Epithelial tumor cells may also migrate in a highly directional manner as a collective group in some settings. We previously discovered that myoferlin (MYOF) is overexpressed in breast cancer cells and depletion of MYOF results in a mesenchymal to epithelial transition (MET) and reduced invasion through extracellular matrix (ECM). However, the biomechanical mechanisms governing cell motility during MYOF depletion are poorly understood. We first demonstrated that lentivirus-driven shRNA-induced MYOF loss in MDA-MB-231 breast cancer cells (MDA-231MYOF-KD) leads to an epithelial morphology compared to the mesenchymal morphology observed in control (MDA- 231LTVC) and wild-type cells. Knockdown of MYOF led to significant reductions in cell migration velocity and MDA- 231MYOF-KD cells migrated directionally and collectively, while MDA-231LTVC cells exhibited single cell migration. Decreased migration velocity and collective migration were accompanied by significant changes in cell mechanics. MDA-231MYOF-KD cells exhibited a 2-fold decrease in cell stiffness, a 2-fold increase in cell-substrate adhesion and a 1.5-fold decrease in traction force generation. In vivo studies demonstrated that when immunocompromised mice were implanted with MDA- 231MYOF-KD cells, tumors were smaller and demonstrated lower tumor burden. Moreover, MDA- 231MYOF-KD tumors were highly circularized and did not invade locally into the adventia in contrast to MDA- 231LTVC-injected animals. Thus MYOF loss is associated with a change in tumor formation in xenografts and leads to smaller, less invasive tumors. These data indicate that MYOF, a previously unrecognized protein in cancer, is involved in MDA-MB-231 cell migration and contributes to biomechanical alterations. Our results indicate that changes in biomechanical properties following loss of this protein may be an effective way to alter the invasive capacity of cancer cells.

Highlights

Cell migration is an essential biological process involved in inflammation, tissue repair and regeneration, developmental events, cancer, and immune cell surveillance
We recently discovered that silencing a protein involved in membrane dynamics, myoferlin (MYOF), in the highly invasive MDA-MB231 breast cancer cell line induces mesenchymal to epithelial transition (MET) and modulates the invasive capacity of these cells [21]
When MYOF was diminished by .90%, cells reverted to an epithelial shape that was polygonal and typically contained a single broad lamellipodium with longer filopodia (Figure 1C and F)

Summary

Introduction

Cell migration is an essential biological process involved in inflammation, tissue repair and regeneration, developmental events, cancer, and immune cell surveillance. Individual cells migrate within the extracellular matrix (ECM) in a polarized manner, extending forward lamellipodia and actin-rich filopodia [1,2] via either protease-dependent or independent mechanisms [3] In combination with these cellular protrusions, focal adhesion dynamics, actin polymerization, and actomyosin contraction result in internal tension within the cell. The formation of distinct leading and trailing edges coordinate migration activity [5], while collective cell migration is governed by several biophysical factors including the distribution of tensile stress within the monolayer [6], transmission of mechanical force across cell-cell junctions [6,7], and the distribution of cell stiffness within the advancing cell sheet [8] In both cases, the cell motility cycle involves steps that occur in many cell types in response to external stimuli and to intracellular and intercellular signaling [9]. This interaction between the tumor and stroma cells with the ECM represents a primary factor in epithelial to mesenchymal transition (EMT) [14]

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Conclusion