title: Chromatin immunoprecipitation scanning identifies glucocorticoid receptor binding regions in the proximal promoter of a ubiquitously expressed glucocorticoid target gene in brain.
abstract: While the actions of glucocorticoids on brain functions have been comprehensively studied, the underlying genomic mechanisms are poorly understood. In this study, we show that glucocorticoid-induced leucine zipper (GILZ) mRNA is strongly and ubiquitously induced in rat brain. To decipher the molecular mechanisms underlying these genomic effects, it is of interest to identify the regulatory sites in the promoter region. Alignment of the rat GILZ promoter with the well-characterized human promoter resulted in poor sequence homology. Consequently, we analyzed the rat 5' flanking sequence by Matrix REDUCE and identified two high-affinity glucocorticoid response elements (GRE) located 2 kb upstream of the transcription start site. These findings were corroborated using the glucocorticoid receptor (GR) expressing Ns-1 PC12 rat cell-line. In these cells, dexamethasone treatment leads to a progressive increase of GILZ mRNA expression levels via a GR-dependent mechanism. Subsequently, using chromatin immunoprecipitation assays we show that the two high-affinity GREs are located within the GR-binding regions. Lastly, we demonstrate using multiple tissue in situ hybridization a marked increase in mRNA expression levels in spleen, thymus, heart, lung, liver, muscle, testis, kidney, colon, ileum, as well as in brain and conclude that the GILZ gene can be used to study glucocorticoid effects in many additional rodent tissues.
authors.affiliation: Division of Medical Pharmacology, Leiden/Amsterdam Centre for Drug Research (LACDR), Leiden, The Netherlands. siem.laan@gmail.com
authors.forename: Siem
authors.initials: S
authors.lastname: van der Laan
authors.forename: R Angela
authors.initials: RA
authors.lastname: Sarabdjitsingh
authors.forename: Marcel F
authors.initials: MF
authors.lastname: Van Batenburg
authors.forename: Servane B
authors.initials: SB
authors.lastname: Lachize
authors.forename: Hualing
authors.initials: H
authors.lastname: Li
authors.forename: Thomas F
authors.initials: TF
authors.lastname: Dijkmans
authors.forename: Erno
authors.initials: E
authors.lastname: Vreugdenhil
authors.forename: E Ron
authors.initials: ER
authors.lastname: de Kloet
authors.forename: Onno C
authors.initials: OC
authors.lastname: Meijer
mesh_terms: D024506:5' Flanking Region / Q000235:genetics; D000818:Animals; D001665:Binding Sites / Q000187:drug effects / Q000235:genetics; D001921:Brain / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation / Q000379:methods; D003907:Dexamethasone / Q000378:metabolism / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics*; D005938:Glucocorticoids / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008967:Molecular Biology / Q000379:methods; D016716:PC12 Cells; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000235:genetics*; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011965:Receptors, Glucocorticoid / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014781:Viscera / Q000378:metabolism
chemical_list: D005938:Glucocorticoids; D012333:RNA, Messenger; D011965:Receptors, Glucocorticoid; D014157:Transcription Factors; C498609:Tsc22d3 protein, rat; D003907:Dexamethasone