Distribution of Microtubule Organizing Centers

This study was designed to investigate the relationship between the position of the microtubule organizing center (MTOC) and the direction of migration of a sheet of endothelial cells (EC) . Using immunofluorescence and phase microscopy the MTOC's of migrating EC were visualized as the cells moved into an in vitro experimental wound produced by mechanical denudation of part of a confluent monolayerculture . Although the MTOC's in nonmigrating EC were randomly positioned in relation to the nucleus, in migrating cells the position of the MTOC's changed so that 80% of the cells had the MTOC positioned in front of the nucleus toward the direction of movement of the endothelial sheet . This repositioning of the MTOC occurred within the first 4 h after wounding and was associated with the beginning of migration of EC's into the wounded area as seen by time-lapse cinemicrophotography . These studies focus attention on the MTOC as a cytoskeletal structure that may play a role in determining the direction of cell movement . Although the intracellular mechanisms controlling and regulating cell movement are unknown, numerous studies have suggested that the distribution of the various components of the cytoskeleton, particularly the microfilaments and their associated proteins play an important role in the generation of force and in determining the direction ofmovement during cell migration (1-7) . Several studies have also explored the role of micrombules (MT's) (8, 9) and cilia (6) in cell migration . Badleyiet al . (8), comparing the cytoskeleton in singlemigrating and stationary chick fibroblasts using immunofluorescence microscopy, concluded that the distribution of MT's does not alter significantly during the conversion from the migratory to the stationary state. There are, however, studies that show that coordinated movement in one direction requires the presence of MT's and that movement is either reduced (9), inhibited (10), or can occur only randomly (11-13) when MT's are disrupted as in colchicine-treated cells. Malech et al . (14) showed that colchicine had no effect on random migration of human neutrophils ; however, activated random migration was minimally decreased and directed migration was markedly inhibited . They also showed that the position of the centriole and its associated MT's appear to be important in establishing the direction of migration of neutrophils . The distribution of MT"s in cells is controlled by microtubule organizing centers (MTOC's) which are sites capable of initiating the polymerization of MT's from tubulin both in vivo and in vitro (15-24) . Numerous MT's can be demonstrated to radiate in different directions throughout the cytoplasm from the MTOC's commonly found in the perinuclear region near the centrioles (2527) . THE JOURNAL OF CELL BIOLOGY " VOLUME 91 NOVEMBER 1981 589-594 ©The Rockefeller University Press " 0021-9525/81/11/0589/06$1 .00 To study the role of MTOC's and MT's in cell migration we examined sheets ofendothelial cells (EC's) migrating in a welldefined direction after experimental wounding of a confluent monolayer (28) . Immunofluorescence microscopy with antibodies to tubulin and phase contrast microscopy of living cells were used to visualize the changes in the position ofthe MTOC in relation to the nucleus and the direction of movement . In nonmigrating cells the MTOC's were randomly positioned in relation to the nucleus. In migrating sheets of EC's however, the position of the MTOC's changed in the great majority of cells along the wound edge, so that they were found in front of the nucleus facing the direction of movement . These studies support the hypothesis that the MTOC's may be an important cytoskeletal structure which, together with intact MT's, play a role in determining the direction of movement . MATERIALS AND METHODS