Rho-kinase phosphorylates COOH-terminal threonines of ezrin/radixin/moesin (ERM) proteins and regulates their head-to-tail association

Takeshi Matsui, Masato Maeda, Yoshinori Doi, Shigenobu Yonemura, Mutsuki Amano, Kozo Kaibuchi, Sachiko Tsukita, Shoichiro Tsukita

Research output: Contribution to journalArticle

690 Citations (Scopus)

Abstract

The ezrin/radixin/moesin (ERM) proteins are involved in actin filament/plasma membrane interaction that is regulated by Rho. We examined whether ERM proteins are directly phosphorylated by Rho-associated kinase (Rho-kinase), a direct target of Rho. Recombinant full-length and COOH- terminal half radixin were incubated with constitutively active catalytic domain of Rho-kinase, and ~30 and ~100% of these molecules, respectively, were phosphorylated mainly at the COOH-terminal threonine (T564). Next, to detect Rho-kinase-dependent phosphorylation of ERM proteins in vivo, we raised a mAb that recognized the T564-phosphorylated radixin as well as ezrin and moesin phosphorylated at the corresponding threonine residue (T567 and T558, respectively). Immunoblotting of serum-starved Swiss 3T3 cells with this mAb revealed that after LPA stimulation ERM proteins were rapidly phosphorylated at T567 (ezrin), T564 (radixin), and T558 (moesin) in a Rho- dependent manner and then dephosphorylated within 2 min. Furthermore, the T564 phosphorylation of recombinant COOH-terminal half radixin did not affect its ability to bind to actin filaments in vitro but significantly suppressed its direct interaction with the NH2-terminal half of radixin. These observations indicate that the Rho-kinase-dependent phosphorylation interferes with the intramolecular and/or intermolecular head-to-tail association of ERM proteins, which is an important mechanism of regulation of their activity as actin filament/plasma membrane crosslinkers.

Original languageEnglish
Pages (from-to)647-657
Number of pages11
JournalJournal of Cell Biology
Volume140
Issue number3
DOIs
Publication statusPublished - 09-02-1998

All Science Journal Classification (ASJC) codes

  • Cell Biology

Fingerprint Dive into the research topics of 'Rho-kinase phosphorylates COOH-terminal threonines of ezrin/radixin/moesin (ERM) proteins and regulates their head-to-tail association'. Together they form a unique fingerprint.

  • Cite this