Focal Adhesions Formation and Cell Migration: Differential Roles of Orai1, STIM1 and PLA2G6

Abstract

Store-operated Ca2+ entry (SOCE), and its major components (Orai1, STIM1 and PLA2G6) are known to be important for cell migration, but mechanism of their involvement in this complex process is unknown. In this study we used state-of-the-art imaging techniques and advanced molecular approaches to characterize the role of Orai1, STIM1 and PLA2G6 in lamellipodia and focal adhesion (FA) formation in HEK293 cells. We found that while transfection of the cells with siRNA against Orai1, STIM1 or PLA2G6 resulted in a similar reduction in the velocity and distance of cell migration, their effects on lamellipodia, FAs and the actin cytoskeleton were not the same. Downregulation of Orai1 or PLA2G6, but not STIM1 caused significant reduction in lamellipodia formation and affected actin cytoskeleton. Analysis of FAs showed that molecular downregulation of PLA2G6 changed the appearance and overall distribution of FAs. Compared to FAs in control (scrRNA treated) cells, mean area and aspect ratio of FAs in PLA2G6-deficient cells were significantly reduced. Moreover, FAs accumulated closer to the leading edge of polarized cells: about 80% of FAs were found within a short (up to 4 μm) distance from the leading edge, compared to less than 50% of FAs in control cells, and FAs beyond 10 μm distance virtually disappeared. In contrast, downregulation of STIM1 did not affect either mean area of FAs, or their aspect ratio. FAs in STIM1 siRNA treated cells showed much wider distribution throughout the lamellipodium, and could easily be found far beyond the leading edge. These data suggest that Orai1, PLA2G6 and STIM1 may be involved in different steps of FA formation, maturation and cytoskeleton integration that are all essential for accurate cell migration.