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 microM extravesicular Ca(2+), B[a]P-7,8-dione exhibited threshold and EC(50) values of 0.4 and 2 microM, respectively, and a maximal release rate of 2 micromol 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 (< or = 20 microM) 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.
Pessah
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IN
Department of Molecular Biosciences, School of Veterinary Medicine, University of California, Davis, California, USA. inpessah@ucdavis.edu
Beltzner
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C
Burchiel
S W
SW
Sridhar
G
G
Penning
T
T
Feng
W
W
D000818:Animals; D001564:Benzo(a)pyrene / Q000494:pharmacology*; D001580:Benzopyrenes / Q000494:pharmacology*; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D066298:In Vitro Techniques; D008861:Microsomes / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D010084:Oxidation-Reduction; D011809:Quinones / Q000378:metabolism; D011817:Rabbits; D019837:Ryanodine Receptor Calcium Release Channel / Q000502:physiology*; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases
D001580:Benzopyrenes; D011809:Quinones; D019837:Ryanodine Receptor Calcium Release Channel; D001564:Benzo(a)pyrene; C076588:benzo(a)pyrene-7,8-dione; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; D002118:Calcium