title: Proteinase inhibitor 9, an inhibitor of granzyme B-mediated apoptosis, is a primary estrogen-inducible gene in human liver cells.
abstract: Although liver is an estrogen target tissue, the number of hepatic genes known to be directly induced by estrogen is very small. We identified proteinase inhibitor 9, or PI-9, as being rapidly and strongly induced by estrogen in an estrogen receptor-positive human liver cell line (HepG2-ER7). Since PI-9 mRNA was also induced by estrogen in a human liver biopsy sample, PI-9 is a genuine estrogen-regulated human gene. PI-9 is a potent inhibitor of granzyme B and of granzyme B-mediated apoptosis. Estrogens induced PI-9 mRNA within 2 h, PI-9 mRNA levels reached a plateau of 30-40-fold induction in 4 h, and induction was not blocked by cycloheximide, indicating that induction of PI-9 mRNA is a primary response. The antiestrogen trans-hydroxytamoxifen was a partial agonist for PI-9 mRNA induction, whereas the antiestrogen ICI 182, 780 was a pure antagonist. Western blot analysis showed that estrogen strongly increases PI-9 protein levels. Inhibition of transcription with actinomycin D resulted in identical rates of PI-9 mRNA decay in the presence and absence of estrogen. We isolated genomic clones containing the PI-9 promoter region, identified a putative transcription start site, and carried out transient transfections of PI-9-luciferase reporter gene constructs. The estrogen, moxestrol, elicited a robust induction from the PI-9-luciferase reporter. Mutational inactivation of three potential imperfect estrogen response elements in the PI-9 5'-flanking region had no effect on moxestrol estrogen receptor induction.
authors.affiliation: Department of Biochemistry, University of Illinois, Urbana, Illinois 61801, USA.
authors.forename: H
authors.initials: H
authors.lastname: Kanamori
authors.forename: S
authors.initials: S
authors.lastname: Krieg
authors.forename: C
authors.initials: C
authors.lastname: Mao
authors.forename: V A
authors.initials: VA
authors.lastname: Di Pippo
authors.forename: S
authors.initials: S
authors.lastname: Wang
authors.forename: D A
authors.initials: DA
authors.lastname: Zajchowski
authors.forename: D J
authors.initials: DJ
authors.lastname: Shapiro
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D001483:Base Sequence; D015152:Blotting, Northern; D006528:Carcinoma, Hepatocellular / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004965:Estrogen Antagonists / Q000494:pharmacology; D004967:Estrogens / Q000494:pharmacology*; D004997:Ethinyl Estradiol / Q000031:analogs & derivatives / Q000494:pharmacology; D005786:Gene Expression Regulation; D053804:Granzymes; D006801:Humans; D008099:Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000187:drug effects; D020218:Response Elements; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012697:Serine Endopeptidases / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors / Q000235:genetics / Q000502:physiology; D015843:Serpins / Q000235:genetics / Q000502:physiology*; D013997:Time Factors; D014158:Transcription, Genetic / Q000187:drug effects; D014407:Tumor Cells, Cultured
chemical_list: D004965:Estrogen Antagonists; D004967:Estrogens; D012333:RNA, Messenger; C102316:SERPINB9 protein, human; D015842:Serine Proteinase Inhibitors; D015843:Serpins; D004997:Ethinyl Estradiol; C011310:moxestrol; C509213:GZMB protein, human; D053804:Granzymes; D012697:Serine Endopeptidases