Actin-organelle interaction: Association with chloroplast in Arabidopsis leaf mesophyll cells

The role of the cytoskeleton in the regulation of chloroplast motility and positioning has been investigated by studying: (1) structural relationship of at tin microfilaments, microtubules, and chloroplasts in cryofixed and F reeze-substituted leaf cells of Arabidopsis; and (2) the effects of anti-ac tin (Latrunculin B; LAT-B) and anti-microtubule (Oryzalin) drugs on intrace llular distribution of chloroplasts. Immunolabeling of leaf cells with two plant-actin specific antibodies, which react equivalently with all the expr essed Arabidopsis actins, revealed two arrangements of actin microfilaments : longitudinal arrays of thick actin bundles and randomly oriented thin act in filaments that extended From the bundles. Chloroplasts were either align ed along the actin bundles or closely associated with the fine filaments. B askets of actin microfilaments were also observed around the chloroplasts. The leaf cells labeled with an anti-tubulin antibody showed dense transvers e arrays of cortical microtubules that exhibited no apparent association wi th chloroplasts. The application of LAT-B severely disrupted actin filament s and their association with chloroplasts. In addition, LAT-B induced aberr ant aggregation of chloroplasts in the mesophyll and bundle sheath cells. D ouble labeling of LAT-B heated cells with anti-actin and anti-tubulin antib odies revealed that the microtubules in these cells were unaffected. Moreov er, depolymerization of microtubules with Oryzalin did not affect the distr ibution of chloroplasts. These results provide evidence for the involvement of actin, but not tubulin, in the movement and positioning and chloroplast s in leaf cells. We propose that using motor molecules, some chloroplasts m igrate: along the actin cables directly, while others are pulled along the cables by the fine actin filaments. The baskets of microfilaments may ancho r the chloroplasts during streaming and allow control over proper three-dim ensional orientation to light. (C) 1999 Wiley-Liss Inc.