title: Targeting human glutathione transferase A3-3 attenuates progesterone production in human steroidogenic cells.
abstract: hGSTA3-3 (human Alpha-class glutathione transferase 3-3) efficiently catalyses steroid Delta(5)-Delta(4) double-bond isomerization in vitro, using glutathione as a cofactor. This chemical transformation is an obligatory reaction in the biosynthesis of steroid hormones and follows the oxidation of 3beta-hydroxysteroids catalysed by 3beta-HSD (3beta-hydroxysteroid dehydrogenase). The isomerization has commonly been ascribed to a supplementary function of 3beta-HSD. The present study is the first to provide evidence that hGSTA3-3 contributes to this step in steroid hormone biosynthesis in complex cellular systems. First, we find glutathione-dependent Delta(5)-Delta(4) isomerase activity in whole-cell extracts prepared from human steroidogenic cells. Secondly, effective inhibitors of hGSTA3-3 dramatically decrease the conversion of Delta(5)-androstene-3,17-dione into Delta(4)-androstene-3,17-dione in cell lysates. Thirdly, we show that RNAi (RNA interference) targeting hGSTA3-3 expression decreases by 30% the forskolin-stimulated production of the steroid hormone progesterone in a human placental cell line. This effect is achieved at low concentrations of two small interfering RNAs directed against distinct regions of hGSTA3-3 mRNA, and is weaker in unstimulated cells, in which hGSTA3-3 expression is low. The results concordantly show that hGSTA3-3 makes a significant contribution to the double-bond isomerization necessary for steroid hormone biosynthesis and thereby complements the indispensable 3beta-hydroxysteroid oxidoreductase activity of 3beta-HSD. The results indicate that the lower isomerase activity of 3beta-HSD is insufficient for maximal rate of cellular sex hormone production and identify hGSTA3-3 as a possible target for pharmaceutical intervention in steroid hormone-dependent diseases.
authors.affiliation: Department of Biochemistry and Organic Chemistry, Uppsala University, Biomedical Center, Husargatan 3, Box 576, SE-75123 Uppsala, Sweden. Francoise.Mathieu@biorg.uu.se
authors.forename: Françoise
authors.initials: F
authors.lastname: Raffalli-Mathieu
authors.forename: Carolina
authors.initials: C
authors.lastname: Orre
authors.forename: Mats
authors.initials: M
authors.lastname: Stridsberg
authors.forename: Maryam
authors.initials: M
authors.lastname: Hansson Edalat
authors.forename: Bengt
authors.initials: B
authors.lastname: Mannervik
mesh_terms: D002460:Cell Line; D004791:Enzyme Inhibitors / Q000737:chemistry; D005260:Female; D018390:Gene Targeting / Q000379:methods; D005982:Glutathione Transferase / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000502:physiology; D012739:Gonadal Steroid Hormones / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology; D006801:Humans; D010920:Placenta / Q000166:cytology* / Q000201:enzymology* / Q000378:metabolism; D011374:Progesterone / Q000096:biosynthesis* / Q000235:genetics*; D034622:RNA Interference; D013237:Stereoisomerism
chemical_list: D004791:Enzyme Inhibitors; D012739:Gonadal Steroid Hormones; D011374:Progesterone; D005982:Glutathione Transferase; C495975:glutathione S-transferase A3-3, human