Dephosphorylation and deactivation of Ca2+/calmodulin-dependent protein kinase II in betaTC3-cells is mediated by Mg2+- and okadaic-acid-sensitive protein phosphatases

Biochem J. 1998 Jan 15;329 ( Pt 2)(Pt 2):283-8. doi: 10.1042/bj3290283.

Abstract

The alpha-toxin-permeabilized betaTC3 cell has been utilized as an experimental model for the identification of protein phosphatases responsible for the dephosphorylation and deactivation of Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) in situ. In this model, the elevation of Ca2+ from 0.05 to 10 microM induced the near-total conversion of CaM kinase II into a Ca2+/calmodulin-independent (autonomous) form characteristic of autophosphorylated, activated enzyme. On the removal of Ca2+, the activation state of CaM Kinase II rapidly returned to prestimulated levels. This reversal was slowed, but not prevented, by the inhibitors of protein phosphatase-1 (PP-1) and PP-2A, okadaic acid and calyculin A, and by the selective chelation of Mg2+ by the addition of EDTA. Near-complete prevention of enzyme deactivation, however, was observed in the combined presence of both okadaic acid and EDTA. Under these conditions, CaM kinase II phosphatase was more sensitive to calyculin A relative to okadaic acid, characteristic of the involvement of PP-1. CaM kinase II deactivation was not affected by FK-506, eliminating the involvement of PP-2B in this process. These data suggest that CaM kinase II dephosphorylation and deactivation in the pancreatic beta-cell is mediated by the combined action of an okadaic-acid-sensitive phosphatase and a Mg2+-dependent phosphatase, such as PP-2C.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2
  • Calcium-Calmodulin-Dependent Protein Kinases / antagonists & inhibitors
  • Calcium-Calmodulin-Dependent Protein Kinases / metabolism*
  • Cell Line
  • Enzyme Inhibitors / pharmacology*
  • Magnesium / pharmacology*
  • Mice
  • Mice, Transgenic
  • Okadaic Acid / pharmacology*
  • Phosphoprotein Phosphatases / antagonists & inhibitors
  • Phosphoprotein Phosphatases / metabolism*
  • Phosphorylation
  • Protein Phosphatase 1

Substances

  • Enzyme Inhibitors
  • Okadaic Acid
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2
  • Calcium-Calmodulin-Dependent Protein Kinases
  • Phosphoprotein Phosphatases
  • Protein Phosphatase 1
  • Magnesium