(137a) Endothelial Sarcomere Fluctuations Arise From Actin Polymerization at Focal Adhesions

Generation and maintenance of intracellular tension is necessary for endothelial cells to adhere to the basement membrane and resist external mechanical stresses. Tension in endothelial cells is generated in stress fibers which are observed not only in vitro, but also in vivo. Tensile force is generated in endothelial stress fibers through tens of repeating contractile units called sarcomeres. In a recent paper, we showed that there exists an exponential distribution of contraction lengths in sarcomeres of severed stress fibers. By hypothesizing that sarcomeres in a stress fiber may continually fluctuate with time, but with an average velocity that is negative (i.e. causes contraction), we could explain the observed exponential distribution of contraction distances. To test this hypothesis, we performed imaging of sarcomeres in living endothelial cells. Sarcomeres were observed to continually fluctuate in length and in some cases lengthen and break. Nascent sarcomeres were observed to continually flow into pre-existing fibers at focal adhesions. Adjacent sarcomeres were frequently found to fuse end-to-end. Dynamic fluctuations in sarcomere length occurred in cells of constrained shape, were attenuated with serum starvation, and could be triggered in serum-starved cells by the addition of LPA which triggers actin polymerization. Together, these results suggest that sarcomeres are not static structures but continually fluctuate in living stress fibers, and that these fluctuations require a continual flow of nascent sarcomeres from focal adhesions into intact stress fibers.