ACTIN IN LIVING AND FIXED CHARACEAN INTERNODAL CELLS - IDENTIFICATIONOF A CORTICAL ARRAY OF FINE ACTIN STRANDS AND CHLOROPLAST ACTIN RINGS

We report on the novel features of the actin cytoskeleton and its deve lopment in characean internodal cells. Images obtained by confocal las er scanning microscopy after microinjection of living cells with fluor escent derivatives of F-actin-specific phallotoxins, and by modified i mmunofluorescence methods using fixed cells, were mutually confirmator y at all stages of internodal cell growth. The microinjection method a llowed capture of 3-dimensional images of high quality even though pho tobleaching and apparent loss of the probes through degradation and up take into the vacuole made it difficult to record phallotoxin-labelled actin over long periods of time. When injected at appropriate concent rations, phallotoxins affected neither the rate of cytoplasmic streami ng nor the long-term viability of cells. Recently formed internodal ce lls have relatively disorganized actin bundles that become oriented in the subcortical cytoplasm approximately parallel to the newly establi shed long axis and traverse the cell through transvacuolar strands. In older cells with central vacuoles not traversed by cytoplasmic strand s, subcortical bundles are organized in parallel groups that associate closely with stationary chloroplasts, now in files. The parallel arra ngement and continuity of actin bundles is maintained where they pass round nodal regions of the cell, even in the absence of chloroplast fi les. This study reports on two novel structural features of the charac ean internodal actin cytoskeleton: a distinct array of actin strands n ear the plasma membrane that is oriented transversely during cell grow th and rings of actin around the chloroplasts bordering the neutral li ne, the zone that separates opposing flows of endoplasm.