DEPHOSPHORYLATION AND DEACTIVATION OF CA2+ CALMODULIN-DEPENDENT PROTEIN-KINASE-II IN BETA-TC3-CELLS IS MEDIATED BY MG2+-SENSITIVE AND OKADAIC-ACID-SENSITIVE PROTEIN PHOSPHATASES/

The alpha-toxin-permeabilized beta TC3 cell has been utilized as an ex perimental model for the identification of protein phosphatases respon sible for the dephosphorylation and deactivation of Ca2+/calmodulin-de pendent protein kinase II (CaM kinase II) in situ. In this model, the elevation of Ca2+ from 0.05 to 10 mu M induced the near-total conversi on of CaM kinase II into a Ca2+/calmodulin-independent (autonomous) fo rm characteristic of autophosphorylated, activated enzyme. On the remo val 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-l (PP-1) and PP-2A, okadaic aci d and calyculin A, and by the selective chelation of Mg2+ by the addit ion 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 t o calyculin A relative to okadaic acid, characteristic of the involvem ent of PP-1. CaM kinase II deactivation was not affected by FK-506, el iminating the involvement of PP-2B in this process. These data suggest that CaM kinase II dephosphorylation and deactivation in the pancreat ic beta-cell is mediated by the combined action of an okadaic-acid-sen sitive phosphatase and a Mg2+-dependent phosphatase, such as PP-2C.