title: Regulation of murine hepatic hydroxysteroid sulfotransferase expression in hyposulfatemic mice and in a cell model of 3'-phosphoadenosine-5'-phosphosulfate deficiency.
abstract: The cytosolic sulfotransferases (SULTs) catalyze the sulfate conjugation of nucleophilic substrates, and the cofactor for sulfonation, 3'-phosphoadenosine-5'-phosphosulfate (PAPS), is biosynthesized from sulfate and ATP. The phenotype of male knockout mice for the NaS1 sodium sulfate cotransporter includes hyposulfatemia and increased hepatic expression of mouse cytoplasmic sulfotransferase Sult2a and Sult3a1. Here we report that in 8-week-old female NaS1-null mice, hepatic Sult2a1 mRNA levels were ∼51-fold higher than they were in a wild-type liver but expression of no other Sult was affected. To address whether hyposulfatemia-inducible Sult2a1 expression might be due to reduced PAPS levels, we stably knocked down PAPS synthases 1 and 2 in HepG2 cells (shPAPSS1/2 cells). When a reporter plasmid containing at least 233 nucleotides (nt) of Sult2a1 5'-flanking sequence was transfected into shPAPSS1/2 cells, reporter activity was significantly increased relative to the activity that was seen for reporters containing 179 or fewer nucleotides. Mutation of an IR0 (inverted repeat of AGGTCA, with 0 intervening bases) nuclear receptor motif at nt -191 to 180 significantly attenuated the PAPSS1/2 knockdown-mediated increase. PAPSS1/2 knockdown significantly activated farnesoid X receptor (FXR), retinoid-related orphan receptor, and pregnane X receptor responsive reporters, and treatment with the FXR agonist GW4064 [3-(2,6-dichlorophenyl)-4-(3'-carboxy-2-chlorostilben-4-yl)oxymethyl-5-isopropylisoxazole] increased Sult2a1 promoter activity when the IR0 was intact. Transfection of shPAPSS1/2 cells with FXR small interfering RNA (siRNA) significantly reduced the Sult2a1 promoter activity. The impact of PAPSS1/2 knockdown on Sult2a1 promoter activity was recapitulated by knocking down endogenous SULT2A1 expression in HepG2 cells. We propose that hyposulfatemia leads to hepatic PAPS depletion, which causes loss of SULT2A1 activity and results in accumulation of nonsulfated bile acids and FXR activation.
authors.affiliation: Institute of Environmental Health Sciences, 259 Mack Avenue, Room 4118, Wayne State University, Detroit, MI 48201, USA.
authors.forename: Kathleen G
authors.initials: KG
authors.lastname: Barrett
authors.forename: Hailin
authors.initials: H
authors.lastname: Fang
authors.forename: Mary D
authors.initials: MD
authors.lastname: Gargano
authors.forename: Daniel
authors.initials: D
authors.lastname: Markovich
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Kocarek
authors.forename: Melissa
authors.initials: M
authors.lastname: Runge-Morris
mesh_terms: D000818:Animals; D027682:Cation Transport Proteins / Q000502:physiology; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D056945:Hep G2 Cells; D006801:Humans; D008099:Liver / Q000201:enzymology*; D051379:Mice; D009097:Multienzyme Complexes / Q000502:physiology; D010724:Phosphoadenosine Phosphosulfate / Q000172:deficiency*; D011401:Promoter Regions, Genetic; D000077003:Sodium Sulfate Cotransporter; D013430:Sulfate Adenylyltransferase / Q000502:physiology; D013431:Sulfates / Q000097:blood; D015238:Sulfotransferases / Q000235:genetics*; D027981:Symporters / Q000502:physiology
chemical_list: D027682:Cation Transport Proteins; D009097:Multienzyme Complexes; D000077003:Sodium Sulfate Cotransporter; D013431:Sulfates; D027981:Symporters; D010724:Phosphoadenosine Phosphosulfate; C022322:PAPS synthetase; D013430:Sulfate Adenylyltransferase; D015238:Sulfotransferases; C028152:alcohol sulfotransferase