Abstract
Cells migrating over 2D substrates are required to polymerise actin at the leading edge to form lamellipodia protrusions and nascent adhesions to anchor the protrusion to the substrate. The major actin nucleator in lamellipodia formation is the Arp2/3 complex, which is activated by the WAVE regulatory complex (WRC). Using inducible Nckap1 floxed mouse embryonic fibroblasts (MEFs), we confirm that the WRC is required for lamellipodia formation, and importantly, for generating the retrograde flow of actin from the leading cell edge. The loss of NCKAP1 also affects cell spreading and focal adhesion dynamics. In the absence of lamellipodium, cells can become elongated and move with a single thin pseudopod, which appears devoid of N-WASP. This phenotype was more prevalent on collagen than fibronectin, where we observed an increase in migratory speed. Thus, 2D cell migration on collagen is less dependent on branched actin.
Highlights
Cell motility is a highly regulated but dynamic process that is fundamental throughout biology.Many cells, including immune cells, pathogens, and cancerous cells, migrate and traverse biological barriers, often in response to a stimulus
We found that Nckap1 KO mouse embryonic fibroblasts (MEFs) have fewer adhesions (Figure 3A,C) similar to observations of WAVE knockdown cells [12]
The focal adhesion analysis server (FAAS), to quantify adhesion dynamics, we found that overall focal adhesion turnover is slower in Nckap1 KO MEFs compared to control cells (Figure 3F–H)
Summary
Cell motility is a highly regulated but dynamic process that is fundamental throughout biology.Many cells, including immune cells, pathogens, and cancerous cells, migrate and traverse biological barriers, often in response to a stimulus. Classical 2D mesenchymal migration is defined by the formation of a protrusive front and retracting rear. This is predominantly driven by the Arp2/3 complex that induces the nucleation of branched actin filaments at the membrane to form a broad, flat protrusion at the leading edge, termed a lamellipodium [2]. These newly forming lamellipodia are constantly interacting with the surrounding matrix and relaying signals through integrin-based adhesions, a process commonly known as mechanotransduction [3]
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