Loss of Myoferlin Redirects Breast Cancer Cell Motility towards Collective Migration

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.

Myoferlin depletion elevates focal adhesion kinase and paxillin phosphorylation and enhances cell-matrix adhesion in breast cancer cells

“…Previous studies by our group and others have indicated that MYOF is involved in many aspects of cancer cell function (16,32). We have shown that RNAi silencing of MYOF in MDA-MB-231 cells resulted in a mesenchymal-epithelial transition, in molecular, morphological, and biomechanical terms (17,32).…”

“…We have shown that RNAi silencing of MYOF in MDA-MB-231 cells resulted in a mesenchymal-epithelial transition, in molecular, morphological, and biomechanical terms (17,32). Additionally, we have reported that the degree of MYOF expression corresponds with invasive potential in breast epithelial (very low in MCF-10A) and cancer (MCF-7, T47D, MDA-MB-231, and BT549) cell lines (17).…”

“…Specifically, we demonstrated that shRNA-mediated ablation of MYOF reverted cells from the parental mesenchymal phenotype to a polarized morphology and epithelial gene profile (17). Furthermore, we reported that loss of MYOF diminished the invasive capacity and caused highly directed, collective migration, in contrast to the random motility behavior in wild-type MDA-MB-231 cells or their control counterparts (17,32). In the midst of our earlier investigations, we also discovered that MYOF depletion enhanced, by nearly twofold, breast cancer cell adhesion to substrates (32).…”

Cancer cells become less deformable and more invasive with activation of β-adrenergic signaling

“…Myosin II binds to actin and regulates F-actin assembly and disassembly dynamics (Murrell et al, 2015), and also regulates intracellular tension (or pre-stress) and cell contractility, which can contribute to cell stiffness Murrell et al, 2015). Indeed, cancer cells that exhibit greater traction stresses show increased invasive behavior in vitro (Kraning-Rush et al, 2012;Volakis et al, 2014) and in vivo (Paszek et al, 2005;Volakis et al, 2014). Our results suggest that βAR signaling could enhance the contractility of cancer cells.…”

Myoferlin plays a key role in VEGFA secretion and impacts tumor‐associated angiogenesis in human pancreas cancer

“…Recently, myoferlin was found to be important for breast cancer invasion. [13][14][15] Our laboratory showed that myoferlin, is a key regulator of EGFR activity in human breast cancer cells. 16 Interestingly, it was reported, in a Kaplan-Meier survival curve, that patients with myoferlin-positive PDAC had a significant lower overall survival than those with a myoferlinnegative tumor.…”