Molecular mechanism of cofilin dephosphorylation by ouabain.
We previously reported that phosphorylated cofilin-triosephosphate isomerase (TPI) complex interacts with Na,K-ATPase and enhances the pump activity through the phosphorylation of cofilin via Rho-mediated signaling pathway. In this study, we tested the hypothesis that the dephosphorylation of cofilin may be induced through Na,K-ATPase inhibition by ouabain. The phosphorylation level of cofilin by ouabain which decreases in a time- and dose-dependent manner in various human cell lines, remains unchanged by pretreatment with Src inhibitor, PP2; epidermal growth factor receptor (EGFR) inhibitor, AG1478; Raf-1 kinase (Raf) inhibitor, GW5074; and ERK kinase (MEK) inhibitor, PD98059, and by transfection of Ras dominant negative mutant (RasN17). This suggests that ouabain dephosphorylates cofilin through the Src/EGFR/Ras/Raf/MEK pathway. Ouabain activates Ras/Raf/MEK pathway, but down-regulates Rho kinase (ROCK)/LIM kinase (LIMK)/cofilin pathway, implying that there may be a cross-talk by ouabain between the Ras/Raf/MEK and the ROCK/LIMK/cofilin pathways. Immunofluorescence and flow cytometry suggest that ouabain-induced active form of cofilin may be involved in cytoskeletal reorganization and cell volume regulation. Thus, these findings demonstrate a new molecular mechanism for the dephosphorylation of cofilin through the inhibition of Na,K-ATPase by ouabain.
Jung
Jaehoon
J
College of Pharmacy, Center for Cell Signaling Research and Division of Molecular Life Sciences, Ewha Womans University, Seoul 120-750, Republic of Korea.
Kim
Moonhee
M
Choi
Suenghee
S
Kim
Min-Jeong
MJ
Suh
Jae-kyung
JK
Choi
Eung Chil
EC
Lee
Kyunglim
K
D051339:Actin Depolymerizing Factors / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D020935:MAP Kinase Signaling System; D051379:Mice; D008954:Models, Biological; D041681:NIH 3T3 Cells; D010042:Ouabain / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D013997:Time Factors
D051339:Actin Depolymerizing Factors; D004791:Enzyme Inhibitors; D010042:Ouabain