Title: Recruitment of CREB1 and histone deacetylase 2 (HDAC2) to the mouse Ltbp-1 promoter regulates its constitutive expression in a dioxin receptor-dependent manner.
Authors: Gomez-Duran, Aurea; Ballestar, Esteban; Carvajal-Gonzalez, Jose M; Marlowe, Jennifer L; Puga, Alvaro; Esteller, Manel; Fernandez-Salguero, Pedro M
Published In J Mol Biol, (2008 Jun 27)
Abstract: Latent TGFbeta-binding protein 1 (LTBP-1) is a key regulator of TGFbeta targeting and activation in the extracellular matrix. LTBP-1 is recognized as a major docking molecule to localize, and possibly to activate, TGFbeta in the extracellular matrix. Despite this relevant function, the molecular mechanisms regulating Ltbp-1 transcription remain largely unknown. Previous results from our laboratory revealed that mouse embryonic fibroblasts (MEF) lacking dioxin receptor (AhR) had increased Ltbp-1 mRNA expression and elevated TGFbeta activity, suggesting that AhR repressed Ltbp-1 transcription. Here, we have cloned the mouse Ltbp-1 gene promoter and analysed its mechanism of transcriptional repression by AhR. Reporter gene assays, AhR over-expression and site-directed mutagenesis showed that basal Ltbp-1 transcription is AhR-dependent. Chromatin immunoprecipitation (ChIP) and RNA interference (RNAi) revealed that AhR regulates Ltbp-1 transcription by a mechanism involving recruitment of co-activators such as CREB1 and co-repressors such as HDAC2 to the Ltbp-1 promoter. In AhR-expressing (AhR+/+) MEF cells, the recruitment of HDAC1, 2 and 4 correlated with decreased K8H4 acetylation and impaired binding of pCREB(Ser133) to the Ltbp-1 promoter, likely maintaining a constitutive repressed state. AhR-/- MEF cells had the opposite pattern of HDACs and pCREB1(Ser133) binding to Ltbp-1 promoter, and therefore, over-expressed Ltbp-1 mRNA. In agreement, siRNA for HDAC2 increased Ltbp-1 expression and K8H4 acetylation in AhR+/+ but not in AhR-/- MEF cells. We suggest that HDAC2 binding keeps Ltbp-1 promoter repressed in AhR+/+ MEF cells, whereas in AhR-null MEF cells the absence of HDAC2 and the binding of pCREB(Ser133) allow Ltbp-1 transcription. Thus, epigenetics can contribute to constitutive Ltbp-1 repression by a mechanism requiring AhR activity.
PubMed ID: 18508077
MeSH Terms: Acetylation; Animals; Base Sequence; Cloning, Molecular; Cyclic AMP Response Element-Binding Protein/metabolism*; DNA Methylation; Gene Expression Regulation*; Genotype; Histone Deacetylase 2; Histone Deacetylases/genetics; Histone Deacetylases/metabolism*; Histones/metabolism; Latent TGF-beta Binding Proteins/genetics*; Latent TGF-beta Binding Proteins/metabolism; Mice; Molecular Sequence Data; Mutagenesis, Site-Directed; Promoter Regions, Genetic/genetics*; Protein Binding; RNA Interference; Receptors, Aryl Hydrocarbon/metabolism*; Repressor Proteins/genetics; Repressor Proteins/metabolism*; Response Elements/genetics