ACTIN-BASED VESICLE DYNAMICS AND EXOCYTOSIS DURING WOUND WALL FORMATION IN CHARACEAN INTERNODAL CELLS

Characean internodal cells readily form wound walls upon local membran e damage. In the present study we documented the dynamics of vesicles involved in wound wall secretion and compared them with actin organiza tion in equivalent cells using immunofluorescence. Single exocytotic e vents (spreading of vesicle contents) could be visualized using image enhancement by video microscopy. In control unwounded cells vesicles m oved unidirectionally along parallel actin bundles and rarely contacte d the plasma membrane. The wound response started with (1) local inhib ition of active cytoplasmic streaming (unidirectional movements) due t o inactivation, depolymerization, or mechanical displacement of the su bcortical actin bundles. Accordingly, vesicles performed only oscillat ing motions and moved slowly with the same velocity and direction as p assive endoplasmic flow. (2) Several minutes after wounding, vesicles started to perform random saltatory movements with frequently changing velocities, punctuated by oscillating motion and periods of immobilit y (docking) at the plasma membrane. Vesicle trajectories correlated wi th a fine-meshed actin network at the wound site. (3) Several hours af ter wounding, vesicles moved again unidirectionally along regenerated subcortical actin bundles. Spreading of vesicles (vesicle contents) wa s observed during wound wall formation, i.e., during the period of sal tatory movements when vesicles had access to the plasma membrane. Depe ndent on the type of wound wall being secreted, three variants could b e distinguished: (1) slow and continuous spreading over a time period of several seconds up to 30 min near the plasma membrane, (2) fast spr eading within 80 ms inside an already formed wound wall, and/or (3) fa st spreading at the plasma membrane. We conclude from our study that w ounding-induced changes in vesicle dynamics are due to transient reorg anization of the actin cytoskeleton from parallel bundles to a fine-me shed network. Furthermore, our results indicate that spreading of vesi cle contents varies considerably with time and may be delayed by vesic le docking and/or discharge. (C) 1996 Wiley-Liss. Inc.