Calyculin A-induced neurite retraction is critically dependent on actomyosin activation but not on polymerization state of microtubules.
Calyculin A (CL-A), a toxin isolated from the marine sponge Discodermia calyx, is a strong inhibitor of protein phosphatase 1 (PP1) and 2A (PP2A). Although CL-A is known to induce rapid neurite retraction in developing neurons, the cytoskeletal dynamics of this retraction have remained unclear. Here, we investigated the cytoskeletal dynamics during CL-A-induced neurite retraction in cultured rat hippocampal neurons, using fluorescence microscopy as well as polarized light microscopy, which can visualize the polymerization state of the cytoskeleton in living cells. We observed that MTs were bent while maintaining their polymerization state during the neurite retraction. In addition, we also found that CL-A still induced neurite retraction when MTs were depolymerized by nocodazole or stabilized by paclitaxel. These results imply a mechanism other than depolymerization of MTs for CL-A-induced neurite retraction. Our pharmacological studies showed that blebbistatin and cytochalasin D, an inhibitor of myosin II and a depolymerizer of actin, strongly inhibited CL-A-induced neurite retraction. Based on all these findings, we propose that CL-A generates strong contractile forces by actomyosin to induce rapid neurite retraction independently from MT depolymerization.
Department of Biophysics, Graduate School of Science, Kyoto University, Oiwake, Kitashirakawa, Sakyo-ku, Kyoto 606-8502, Japan.
Ayumu
A
Inutsuka
Makoto
M
Goda
Yoshinori
Y
Fujiyoshi
D000205:Actomyosin / Q000096:biosynthesis*; D000818:Animals; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006624:Hippocampus / Q000166:cytology; D008387:Marine Toxins; D008870:Microtubules / Q000378:metabolism; D016501:Neurites / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D010080:Oxazoles / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D051381:Rats
D004791:Enzyme Inhibitors; D008387:Marine Toxins; D010080:Oxazoles; C059041:calyculin A; D000205:Actomyosin; D010749:Phosphoprotein Phosphatases