A bioactive metabolite of benzo[a]pyrene, benzo[a]pyrene-7,8-dione, selectively alters microsomal Ca2+ transport and ryanodine receptor function.

Polycyclic aromatic hydrocarbons are environmental pollutants known to be carcinogenic and immunotoxic. In intact cell assays, benzo[ a ]pyrene (B[ a ]P) disrupts Ca 2+ homeostasis in both immune and nonimmune cells, but the molecular mechanism is undefined. In this study, B[ a ]P and five metabolites are examined for their ability to alter Ca 2+ transport across microsomal membranes. Using a well-defined model system, junctional SR vesicles from skeletal muscle, we show that a single o -quinone metabolite of B[ a ]P, B[ a ]P-7,8-dione, can account for altered Ca 2+ transport across microsomal membranes. B[ a ]P-7,8-dione induces net Ca 2+ release from actively loaded vesicles in a dose-, time-, and Ca 2+ -dependent manner. In the presence of 5 μM extravesicular Ca 2+ , B[ a ]P-7,8-dione exhibited threshold and EC 50 values of 0.4 and 2 μM, respectively, and a maximal release rate of 2 μmol of Ca 2+ min −1 mg −1 . The mechanism by which B[ a ]P-7,8-dione enhanced Ca 2+ efflux was further investigated by measuring macroscopic fluxes and single RyR1 channels reconstituted in bilayer lipid membranes and direct measurements of SERCA catalytic activity. B[ a ]P-7,8-dione (≤ 20 μM) had no measurable effect on initial rates of Ca 2+ accumulation in the presence of ruthenium red to block ryanodine receptor (RyR1), nor did it alter Ca 2+ -dependent (thapsigargin-sensitive) ATPase activity. B[ a ]P-7,8-dione selectively altered the function of RyR1 in a time-dependent diphasic manner, first activating then inhibiting channel activity. Considering that RyR1 and its two alternate isoforms are broadly expressed in mammalian cells and their important role in Ca 2+ -signaling, the present results reveal a mechanism by which metabolic bioactivation of B[ a ]P may mediate RyR dysfunction of pathophysiological significance.