title: Transsulfuration Is a Significant Source of Sulfur for Glutathione Production in Human Mammary Epithelial Cells.
abstract: The transsulfuration pathway, through which homocysteine from the methionine cycle provides sulfur for cystathionine formation, which may subsequently be used for glutathione synthesis, has not heretofore been identified as active in mammary cells. Primary human mammary epithelial cells (HMEC's) were labeled with35S-methionine for 24 hours following pretreatment with a vehicle control, the cysteine biosynthesis inhibitor propargylglycine or the gamma-glutamylcysteine synthesis inhibitor buthionine sulfoximine. Cell lysates were prepared and reacted with glutathione-S-transferase and the fluorescent labeling compound monochlorobimane to form a fluorescent glutathione-bimane conjugate. Comparison of fluorographic and autoradiographic images indicated that glutathione had incorporated35S-methionine demonstrating that functional transsulfuration occurs in mammary cells. Pathway inhibitors reduced incorporation by roughly 80%. Measurement of glutathione production in HMEC's treated with and without hydrogen peroxide and/or pathway inhibitors indicates that the transsulfuration pathway plays a significant role in providing cysteine for glutathione production both normally and under conditions of oxidant stress.
authors.affiliation: Department of Chemistry, Marshall University, One John Marshall Drive, Huntington, WV 25755-0003, USA.
authors.forename: Andrea D
authors.initials: AD
authors.lastname: Belalcázar
authors.affiliation: Department of Pharmacology, Physiology & Toxicology, Marshall University Joan C. Edwards School of Medicine, One John Marshall Drive, Huntington, WV 25755-0003, USA.
authors.forename: John G
authors.initials: JG
authors.lastname: Ball
authors.affiliation: Department of Chemistry, Marshall University, One John Marshall Drive, Huntington, WV 25755-0003, USA.
authors.forename: Leslie M
authors.initials: LM
authors.lastname: Frost
authors.affiliation: Department of Pharmacology, Physiology & Toxicology, Marshall University Joan C. Edwards School of Medicine, One John Marshall Drive, Huntington, WV 25755-0003, USA.
authors.forename: Monica A
authors.initials: MA
authors.lastname: Valentovic
authors.affiliation: Department of Anatomy and Pathology, Marshall University Joan C. Edwards School of Medicine, One John Marshall Drive, Huntington, WV 25755-0003, USA.
authors.forename: John
authors.initials: J
authors.lastname: Wilkinson