Abstract
The focal adhesion (FA) is an integrin-based structure built in/on the plasma membrane (PM), linking the extracellular matrix to the actin stress-fibers, working as cell migration scaffolds. Previously, we proposed the archipelago architecture of the FA, in which FA largely consists of fluid membrane, dotted with small islands accumulating FA proteins: membrane molecules enter the inter-island channels in the FA zone rather freely, and the integrins in the FA-protein islands rapidly exchanges with those in the bulk membrane. Here, we examined how Rac1, a small G-protein regulating FA formation, and its activators αPIX and βPIX, are recruited to the FA zones. PIX molecules are recruited from the cytoplasm to the FA zones directly. In contrast, majorities of Rac1 molecules first arrive from the cytoplasm on the general inner PM surface, and then enter the FA zones via lateral diffusion on the PM, which is possible due to rapid Rac1 diffusion even within the FA zones, slowed only by a factor of two to four compared with that outside. The constitutively-active Rac1 mutant exhibited temporary and all-time immobilizations in the FA zone, suggesting that upon PIX-induced Rac1 activation at the FA-protein islands, Rac1 tends to be immobilized at the FA-protein islands. © 2013 Wiley Periodicals, Inc
Highlights
The focal adhesion (FA) is an integrin-based structure built in and on the plasma membrane (PM), mechanically linking the extracellular matrix with the termini of actin stress-fibers, providing key scaffolds for the cells to migrate in tissues (Fig. 1A) [Galbraith et al, 2007; Petrie et al, 2012]
Our results indicate that whereas aPIX and bPIX are recruited from the cytoplasm to the FA zone directly, majorities of Rac1 molecules, including the wild-type (WT), constitutively-active (CA) G12V, and dominantnegative (DN) T17N Rac1 molecules, are recruited to the FA zone by way of the general PM area via lateral diffusion on the PM
By using the micro-printed fibronectin, FA zones tended to exhibit similar shapes and sizes, the analysis of the experimental results was facilitated
Summary
The focal adhesion (FA) is an integrin-based structure built in and on the plasma membrane (PM), mechanically linking the extracellular matrix with the termini of actin stress-fibers, providing key scaffolds for the cells to migrate in tissues (Fig. 1A) [Galbraith et al, 2007; Petrie et al, 2012]. Integrin b3 readily enters the FA zone, and repeatedly undergoes the cycles of temporary immobilization and diffusion in the FA zone, whereas $1/3 of integrin b3 becomes immobilized there [Shibata et al, 2012; see a follow-up study by Rossier et al, 2012]. Based on these observations, we proposed the archipelago architecture of the FA (‘‘Archipelago Model’’ in Fig. 1B), or rather the FA zone, in which many small islands of integrins and n 162 Shibata et al
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