PKC phosphorylates MARCKS Ser159 not only directly but also through RhoA/ROCK.
It is well recognized that phorbol 12,13-dibutyrate (PDBu)-activated PKC directly phosphorylates myristoylated alanine-rich C kinase substrate (MARCKS), whose phosphorylation is used as a marker of PKC activation. However, in SH-SY5Y neuroblastoma cells, Western blotting analyses revealed that Rho-associated coiled-coil kinase (ROCK)-specific inhibitor H-1152 inhibited PDBu-induced phosphorylation, and that a small G-protein inhibitor, toxin B, also inhibited MARCKS phosphorylation. Furthermore, in GST pull-down assays, PDBu induced RhoA activation in SH-SY5Y cells, and this activation was inhibited by PKC inhibitor Ro-31-8220. Finally, we showed that the transfection of a dominant negative form of RhoA inhibited PDBu-induced MARCKS phosphorylation in immunocytochemistries. These findings suggest that some PDBu-induced MARCKS phosphorylation includes the RhoA/ROCK pathway in SH-SY5Y cells.
Tanabe
Atsuhiro
A
Laboratory of Pharmacology, Pharmaceutical Sciences, Kitasato University, Minato-ku, Tokyo 108-8641, Japan. tanabea@pharm.kitasato-u.ac.jp
Kamisuki
Yukinori
Y
Hidaka
Hiroyoshi
H
Suzuki
Masaaki
M
Negishi
Manabu
M
Takuwa
Yoh
Y