Role of Ca2+/calmodulin-dependent phosphatase 2B in thrombin-induced endothelial cell contractile responses

Alexander D. Verin, Clare Cooke, Maria Herenyiova, Carolyn E. Patterson, Joe G.N. Garcia

Research output: Contribution to journalArticlepeer-review

50 Scopus citations

Abstract

Thrombin-induced Ca2+ mobilization, activation of Ca2+/calmodulin- dependent myosin light chain (MLC) kinase (MLCK), and increased phosphorylation of MLCs precede and are critical to endothelial cell (EC) barrier dysfunction. Net MLC dephosphorylation after thrombin is nearly complete by 60 min and involves type 1 phosphatase (PPase 1) activity. We now report that thrombin does not alter total PPase 1 activity in EC homogenates but rather decreases myosin-associated PPase 1 activity. The PPase 1 inhibitor calyculin fails to prevent thrombin-induced MLC dephosphorylation. However, thrombin significantly increased the activity of Ca2+-dependent PPase 2B in EC homogenates (~1.5- to 2-fold), with PPase 2B activation correlating with phosphorylation of the PPase 2B catalytic subunit. Western immunoblotting revealed PPase 2B to be present in cytoskeletal EC fractions, with specific PPase 2B inhibitors such as cyclosporin (200 nM) and deltamethrin (100 nM to 1 μM) attenuating thrombin-induced cytoskeletal protein dephosphorylation, including EC MLC dephosphorylation. These results suggest a model whereby thrombin-inducible contraction is determined by the phosphorylation status of EC MLC regulated by the balance between EC MLCK, PPase 1 (constitutive), and PPase 2B (inducible) activities.

Original languageEnglish (US)
Pages (from-to)L788-L799
JournalAmerican Journal of Physiology - Lung Cellular and Molecular Physiology
Volume275
Issue number4 19-4
DOIs
StatePublished - 1998
Externally publishedYes

Keywords

  • Bovine pulmonary artery endothelium
  • Selective phosphatase 2B inhibitors
  • Thrombin-stimulated endothelial cell permeability
  • Thrombin-stimulated myosin light chain phosphorylation

ASJC Scopus subject areas

  • Physiology
  • Pulmonary and Respiratory Medicine
  • Physiology (medical)
  • Cell Biology

Fingerprint Dive into the research topics of 'Role of Ca<sup>2+</sup>/calmodulin-dependent phosphatase 2B in thrombin-induced endothelial cell contractile responses'. Together they form a unique fingerprint.

Cite this