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Characterization at the Molecular Level using Robust Biochemical Approaches of a New Kinase Protein
11:23

Characterization at the Molecular Level using Robust Biochemical Approaches of a New Kinase Protein

Published on: June 30, 2019

Molecular mechanism of cofilin dephosphorylation by ouabain

Jaehoon Jung1, Moonhee Kim, Suenghee Choi

  • 1College of Pharmacy, Center for Cell Signaling Research and Division of Molecular Life Sciences, Ewha Womans University, Seoul 120-750, Republic of Korea.

Cellular Signalling
|May 23, 2006
PubMed

Insights

Ouabain, a Na,K-ATPase inhibitor, dephosphorylates cofilin via the Src/EGFR/Ras/Raf/MEK pathway. This process impacts cytoskeletal reorganization and cell volume regulation, revealing a new molecular mechanism.

Area of Science:

  • Cellular Biology
  • Molecular Signaling
  • Biochemistry

Background:

  • Previously, a link was established between phosphorylated cofilin-triosephosphate isomerase (TPI) complex, Na,K-ATPase activity, and Rho-mediated signaling.
  • Cofilin phosphorylation by the Na,K-ATPase pump was shown to enhance pump activity.

Purpose of the Study:

  • To investigate if Na,K-ATPase inhibition by ouabain induces cofilin dephosphorylation.
  • To elucidate the molecular pathway involved in ouabain-induced cofilin dephosphorylation.

Main Methods:

  • Utilized various human cell lines treated with ouabain.
  • Employed inhibitors for Src, EGFR, Raf-1 kinase, and MEK.
  • Transfected cells with a Ras dominant-negative mutant.
  • Performed immunofluorescence and flow cytometry.

Main Results:

  • Ouabain decreased cofilin phosphorylation in a time- and dose-dependent manner.
  • Cofilin dephosphorylation was independent of Src, EGFR, Raf, MEK, and Ras signaling.
  • Ouabain activated the Ras/Raf/MEK pathway while down-regulating the ROCK/LIMK/cofilin pathway.
  • Ouabain-induced active cofilin correlated with cytoskeletal reorganization and cell volume regulation.

Conclusions:

  • Ouabain inhibits Na,K-ATPase, leading to cofilin dephosphorylation via the Src/EGFR/Ras/Raf/MEK pathway.
  • A cross-talk exists between the Ras/Raf/MEK and ROCK/LIMK/cofilin pathways modulated by ouabain.
  • This study reveals a novel mechanism for cofilin dephosphorylation linked to Na,K-ATPase inhibition.

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