SLCO/OATP-like transport of glutathione in FasL-induced apoptosis: glutathione efflux is coupled to an organic anion exchange and is necessary for the progression of the execution phase of apoptosis.
Apoptosis is characterized by the activation of specific biochemical pathways that lead to the organized demise of cells. Intracellular GSH depletion has been observed during apoptosis; however, neither the mechanisms involved in the reduction of the intracellular GSH concentration, [GSH](i), nor its link to the progression of apoptosis have been elucidated. We have studied this issue using Fas ligand (FasL)-induced apoptosis in Jurkat cells where changes in [GSH](i) can be analyzed biochemically and at the single cell level by flow cytometry. A reduction in the total [GSH](i) in response to FasL occurs in two distinct stages prior to the loss of membrane integrity. Jurkat cells express several members of the multidrug resistance protein (ABCC/MRP), and the organic anion-transporting polypeptide protein (SLCO/OATP) families of GSH efflux pumps at the mRNA level. Glutathione loss and its accumulation in the extracellular medium, induced by FasL, was trans-stimulated by the organic substrates MK571, probenecid, taurocholic acid, estrone sulfate, and bromosulfophthalein and inhibited by high concentrations of extracellular GSH. Single cell analysis demonstrated that intracellular GSH loss was paralleled by the activation of an organic anion uptake process, supporting the role of an anion exchange mechanism (SLCO/OATP-like transport) in GSH efflux induced by FasL. Additionally, high extracellular GSH inhibited the activation of the execution caspases, the cleavage of their substrates poly(ADP-ribose) polymerase (PARP) and alpha-fodrin, and DNA degradation. In contrast, the trans-stimulation of GSH efflux by MK571 increased the cleavage of the execution caspases and their substrates. Together these results suggest that GSH efflux during FasL-induced apoptosis is mediated by a SLCO/OATP-like transport mechanism that modulates the progression of the execution phase of apoptosis.
Franco
Rodrigo
R
Laboratory of Signal Transduction, NIEHS, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
Cidlowski
John A
JA
D017209:Apoptosis / Q000502:physiology*; D001692:Biological Transport; D053222:Fas Ligand Protein / Q000737:chemistry / Q000502:physiology*; D005978:Glutathione / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D007424:Intracellular Fluid / Q000737:chemistry / Q000378:metabolism; D019169:Jurkat Cells; D008024:Ligands; D027361:Organic Anion Transporters / Q000502:physiology*; D010084:Oxidation-Reduction; D019014:fas Receptor / Q000737:chemistry / Q000378:metabolism*
D053222:Fas Ligand Protein; D008024:Ligands; D027361:Organic Anion Transporters; D019014:fas Receptor; D005978:Glutathione