A cytochalasin-sensitive actin filament meshwork is a prerequisite for local wound wall deposition inNitella internodal cells

Abstract

Reorganization of the actin cytoskeleton following cell wall puncturing of characean internodal cells was studied by immunofluorescence and confocal laser scanning microscopy. Injury locally destroyed the parallel subcortical actin filament bundles and cortical actin strands that are characteristic of unwounded regions. At wounds, a delicate three-dimensional interlaced structure of actin strands, with meshes up to 5 μm wide, formed by de novo assembly of isolated filaments and by the elongation of residual subcortical actin bundles and cortical actin strands. The actin meshwork persisted for up to 2 h, corresponding to the duration of intense wound wall secretion. Actin filament bundles continuous with the subcortical bundles outside the wound then regenerated, their parallel alignment probably assisted by endoplasmic flow. Cytochalasin D concentrations that arrested cytoplasmic streaming completely inhibited the formation of the actin meshwork, wound wall deposition and recovery of actin bundles. Concentrations that only reduced streaming velocity delayed meshwork formation and wound walls were thinner than in controls. The actual amount of F-actin within the meshwork, however, was clearly greater in the presence of low cytochalasin concentrations. In late stages of recovery, the actin bundles became very thick and intervening spaces became wider thereby forming a conspicuous, three-dimensional lattice that was continuous with interwebbing subcortical bundles and cortical actin around the periphery of the wound. Our experiments suggest that actin meshwork formation is a prerequisite for plasma membrane-directed transport of vesicles involved in wounding-induced exocytosis in characean internodes. Stabilization of the meshwork by subinhibitory concentrations of cytochalasin D is probably caused by actinbinding properties of the drug that either induce bundling or impede function of associated proteins.