RhoA and Rho kinase regulate the epithelial Na+/H+ exchanger NHE3. Role of myosin light chain phosphorylation

Katalin Szászi, Kazuyoshi Kurashima, András Kapus, Anders Paulsen, Kozo Kaibuchi, Sergio Grinstein, John Orlowski

Research output: Contribution to journalArticlepeer-review

66 Citations (Scopus)

Abstract

The activity of the Na+/H+ exchanger NHE3 isoform, which is found primarily in epithelial cells, is sensitive to the state of actin polymerization. Actin assembly, in turn, is controlled by members of the small GTPase Rho family, namely Rac1, Cdc42, and RhoA. We therefore investigated the possible role of these GTPases in modulating NHE3 activity. Cells stably expressing NHE3 were transiently transfected with inhibitory forms of Rac1, Cdc42, or RhoA and transport activity was assessed using microfluorimetry. NHE3 activity was not adversely affected by either dominant-negative Rac1 or Cdc42. By contrast, the inhibitory form of RhoA greatly depressed NHE3 activity, without noticeably altering its subcellular distribution. NHE3 activity was equally reduced by inhibiting p160 Rho-associated kinase I (ROK), a downstream effector of RhoA, with the selective antagonist Y-27632 and a dominant-negative form of ROK. Furthermore, inhibition of ROK reduced the phosphorylation of myosin light chain. A comparable net dephosphorylation was achieved by the myosin light chain kinase inhibitor ML9, which similarly inhibited NHE3. These data suggest that optimal NHE3 activity requires a functional RhoA-ROK signaling pathway which acts, at least partly, by controlling the phosphorylation of myosin light chain and, ultimately, the organization of the actin cytoskeleton.

Original languageEnglish
Pages (from-to)28599-28606
Number of pages8
JournalJournal of Biological Chemistry
Volume275
Issue number37
DOIs
Publication statusPublished - 15-09-2000
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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