Arp2/3 inhibition induces amoeboid-like protrusions in MCF10A epithelial cells by reduced cytoskeletal-membrane coupling and focal adhesion assembly.

Abstract

Here we demonstrate that Arp2/3 regulates a transition between mesenchymal and amoeboid protrusions in MCF10A epithelial cells. Using genetic and pharmacological means, we first show Arp2/3 inhibition impairs directed cell migration. Arp2/3 inhibition results in a dramatically impaired cell adhesion, causing deficient cell attachment and spreading to ECM as well as an 8-fold decrease in nascent adhesion assembly at the leading edge. While Arp2/3 does not play a significant role in myosin-dependent adhesion growth, mature focal adhesions undergo large scale movements against the ECM suggesting reduced coupling to the ECM. Cell edge protrusions occur at similar rates when Arp2/3 is inhibited but their morphology is dramatically altered. Persistent lamellipodia are abrogated and we observe a markedly increased incidence of blebbing and unstable pseuodopods. Micropipette-aspiration assays indicate that Arp2/3-inhibited cells have a weak coupling between the cell cortex and the plasma membrane, and suggest a potential mechanism for increased pseudopod and bleb formation. Pseudopods are not sensitive to reduced in formin or myosin II activity. Collectively, these results indicate that Arp2/3 is not necessary for rapid protrusion of the cell edge but plays a crucial role in assembling focal adhesions required for its stabilization.