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Induction of actin cytoskeleton rearrangement by methyl okadaate – comparison with okadaic acid
Author(s) -
Vilariño Natalia,
Ares Isabel R.,
Cagide Eva,
Louzao M. Carmen,
Vieytes Mercedes R.,
Yasumoto Takeshi,
Botana Luis M.
Publication year - 2008
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/j.1742-4658.2008.06256.x
Subject(s) - okadaic acid , cytoskeleton , microbiology and biotechnology , actin , actin cytoskeleton , biology , phalloidin , neurite , phosphatase , phosphorylation , biochemistry , chemistry , cell , in vitro
Methyl okadaate is a derivative of the lipophilic polyether okadaic acid (OA), a well‐known inducer of apoptosis. OA inhibits Ser/Thr protein phosphatases (PPs), among them types 1 and 2A (PP1 and PP2A), whereas methyl okadaate lacks PP1/PP2A inhibitory activity in vitro . As progressive loss of neuronal cytoarchitecture is a major event that precedes neuronal death, in this work we studied comparatively the effects of both toxins on actin cytoskeleton organization in human neuroblastoma cells by filamentous actin (F‐actin) labeling with the specific dye Oregon Green 514 Phalloidin. Neither methyl okadaate nor OA modified the amount of F‐actin per cell. However, confocal microscopy imaging showed that methyl okadaate induced reorganization of actin cytoskeleton, loss of the typical flattened morphology and adoption of a round shape, and a reduction in the number of neurites, with a consequent loss of cell attachment. These effects were identical to those induced by OA, although methyl okadaate potency was approximately 10‐fold lower. In order to investigate the role of membrane potential and cytosolic Ca 2+ concentration in morphological changes induced by these toxins, the cells were stained with bis‐(1,3‐dibutylbarbituric acid)‐trimethine oxonol and fura‐2. No toxin effect was detected on membrane potential or calcium influx, indicating that these two signals are not responsible for cytoskeletal/morphological change induction. Methyl okadaate induced an increase of Ser/Thr phosphorylation of cellular proteins detected by western blot, showing similar phosphorylation profiles to OA. Our data suggest that methyl okadaate is an active compound that shares a pharmacological target with OA that may be a Ser/Thr phosphatase, probably different from PP1 and PP2A.