Abstract
Theorientation of actin filaments regulates protrusion and contractile movement of cells because it determines the direction of actin elongation and myosin movement. Little is known about the orientation change of a single actin filament in living cells because of the lack of the method to probe the orientation of actin filaments in living cells. Here we report a method to visualize the orientation of single actin filament. We labeled N and C-termini of tropomyosin with different fluorescent dyes, and introduced it into living cells by electroporation. We performed dual fluorophore-single molecule localization analysis and orientation of actin filaments was determined by the relative position of N and C-termini. We observed the orientation of actin filaments, which flow near focal adhesions(FAs), and found that not only actin filaments rotate rapidly near the FAs, but also they rotate inwardlyin the frontal region of FAs. Actin filaments are also disrupted in the anterior region of FAs. The actin remodeling zone surrounding FAs may be the place for reconstructing the FAs-linked actin network.
