Different effects of Hg 2+ and Cu 2+ on mussel ( Mytilus galloprovincialis) plasma membrane Ca 2+-ATPase: Hg 2+ induction of protein expression

Abstract

Deregulation of Ca 2+ homeostasis can produce serious effects on cell functioning due to an alteration of Ca 2+ signaling. The aim of this study was to evaluate variations in plasma membrane Ca 2+-ATPase (PMCA) induced in mussels by in vivo exposure to Cu 2+ or Hg 2+. PMCA activity was assayed using a cytochemical method allowing localization and in situ quantification of Ca 2+-ATPase on cryostat tissue sections. The effects of fixed concentrations of Cu 2+ (0.6 μM) or Hg 2+ (1.3 μM) were evaluated after different times of exposure (1, 4, 6 days), while those of increasing amounts of Cu 2+ (0.3, 0.6, 1.3 μM) or of Hg 2+ (0.6, 1.3, 2.4 μM) were evaluated after 4 days. Cu 2+ produces dose-dependent inhibition of PMCA in the digestive gland, with a minimum at the fourth day of treatment and a recovery at the sixth day. Conversely, Hg 2+ induces a significant rise of PMCA activity, with a maximum at the fourth day. Similar results have been found after biochemical assay of PMCA, using plasma membranes obtained from density-gradient separation of gill homogenates. PMCA expression has been assessed by immunoprecipitation and Western immunoblotting on digestive gland homogenates, showing an induction after exposure to Hg 2+ but not to Cu 2+. In conclusion, Cu 2+ does not vary PMCA expression but reduces PMCA activity, indicating PMCA inhibition; conversely, Hg 2+ increases PMCA expression more than PMCA activity, suggesting that it also produces PMCA inhibition, but the induction of PMCA expression leads to a net increase in enzyme activity.