The Arp2/3 complex mediates multigeneration dendritic protrusions for efficient 3‐dimensional cancer cell migration

Abstract

Arp2/3 is a protein complex that nucleates actin filament assembly in the lamellipodium in adherent cells crawling on planar 2-dimensional (2D) substrates. However, in physiopathological situations, cell migration typically occurs within a 3-dimensional (3D) environment, and little is known about the role of Arp2/3 and associated proteins in 3D cell migration. Using time resolved live-cell imaging and HT1080, a fibrosarcoma cell line commonly used to study cell migration, we find that the Arp2/3 complex and associated proteins N-WASP, WAVE1, cortactin, and Cdc42 regulate 3D cell migration. We report that this regulation is caused by formation of multigeneration dendritic protrusions, which mediate traction forces on the surrounding matrix and effective cell migration. The primary protrusions emanating directly from the cell body and prolonging the nucleus forms independent of Arp2/3 and dependent on focal adhesion proteins FAK, talin, and p130Cas. The Arp2/3 complex, N-WASP, WAVE1, cortactin, and Cdc42 regulate the secondary protrusions branching off from the primary protrusions. In 3D matrices, fibrosarcoma cells as well as migrating breast, pancreatic, and prostate cancer cells do not display lamellipodial structures. This study characterizes the unique topology of protrusions made by cells in a 3D matrix and show that these dendritic protrusions play a critical role in 3D cell motility and matrix deformation. The relative contribution of these proteins to 3D migration is significantly different from their role in 2D migration.

EB1 and cytoplasmic dynein mediate protrusion dynamics for efficient 3‐dimensional cell migration

“…2 A ), cells embedded in 3D matrix displayed highly branched dendritic protrusions (Fig. 2 B , J ) [see also Giri et al (5)]. These branched protrusions contained both microtubules and F-actin (Fig.…”

“…Recent studies have shown a strong correlation between 3D cell migration speed and protrusion activity, defined here as the number of protrusions generated per unit time, not the length or lifetime of protrusions (5, 11, 12). Therefore, we hypothesized and verified that pharmacological manipulation of microtubules using nocodazole and taxol would reduce protrusion activity because these treatments reduce cell speed in 3D matrices (Fig.…”

Physical view on migration modes

“…During the last two decades, lamellipodial-driven migration has been predominately investigated on flat two-dimensional (2-D) substrates. However, recent studies have pointed out that several aspects of cell motility significantly differ in a three dimensional microenvironment (Wu et al, 2014;Mierke et al, 2010;Giri et al, 2013). In summary, the basic mechanical concept of the lamellipodia-dependent migration is that the actin polymerization induces the protrusion formation of the membrane in the direction of the migration.…”

Three-dimensional cell migration does not follow a random walk

“…However, cell migration in vivo often forces cells to remodel, exert pulling forces on, and move through a 3D collagen I-rich matrix. Recent work has demonstrated that mechanisms of 3D migration are often different from their 2D counterparts (9)(10)(11)(12)(13)(14)(15). Migration on 2D dishes, which induces a basal-apical polarization of the cell, is driven by actomyosin contractility of stress fibers between large focal adhesions and the formation of a wide lamellipodium terminated by thin filopodial protrusions at the leading cellular edge (4,16).…”

“…The same cells in collagen-rich 3D matrix do not display a lamellipodium or filopodia. Instead, they display highly dendritic pseudopodial protrusions controlled by distinct proteins that rely both on acto-myosin contractility and microtubule assembly/disassembly dynamics (11,17). 3D cell migration depends on the expression of metalloproteinases (MMPs), which are dispensable in 2D migration, and physical properties of the 3D matrix (18), such as pore size (6).…”