title: The role of DNA methylation in regulation of the murine Lhx3 gene.
abstract: LHX3 is a LIM-homeodomain transcription factor with critical roles in pituitary and nervous system development. Mutations in the LHX3 gene are associated with pediatric diseases featuring severe hormone deficiencies, hearing loss, developmental delay, and other symptoms. The mechanisms that govern LHX3/Lhx3 transcription are poorly understood. In this study, we examined the role of DNA methylation in the expression status of the mouse Lhx3 gene. Pituitary cells that do not normally express Lhx3 (Pit-1/0 cells) were treated with 5-aza-2'-deoxycytidine, a demethylating reagent. This treatment leads to activation of Lhx3 gene expression suggesting that methylation contributes to Lhx3 regulation. Treatment of Pit-1/0 pituitary cells with a combination of a demethylating reagent and a histone deacetylase inhibitor led to rapid activation of Lhx3 expression, suggesting possible crosstalk between DNA methylation and histone modification processes. To assess DNA methylation levels, treated and untreated Pit-1/0 genomic DNAs were subjected to bisulfite conversion and sequencing. Treated Pit-1/0 cells had decreased methylation at specific sites in the Lhx3 locus compared to untreated cells. Chromatin immunoprecipitation assays demonstrated interactions between the MeCp2 methyl binding protein and Lhx3 promoter regions in the Pit-1/0 cell line. Overall, this study demonstrates that DNA methylation patterns of the Lhx3 gene are associated with its expression status.
authors.affiliation: Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, IN, USA.
authors.forename: Raleigh E
authors.initials: RE
authors.lastname: Malik
authors.affiliation: Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, IN, USA; Department of Biology, Indiana University-Purdue University Indianapolis, IN, USA; Department of Cellular and Integrative Physiology, Indiana University School of Medicine, Indianapolis, IN, USA. Electronic address: srhodes@iupui.edu.
authors.forename: Simon J
authors.initials: SJ
authors.lastname: Rhodes
mesh_terms: D000818:Animals; D002460:Cell Line; D019175:DNA Methylation; D015870:Gene Expression / Q000235:genetics; D006657:Histones / Q000235:genetics / Q000378:metabolism; D060850:LIM-Homeodomain Proteins / Q000235:genetics*; D051783:Methyl-CpG-Binding Protein 2 / Q000235:genetics / Q000378:metabolism; D051379:Mice; D010902:Pituitary Gland / Q000378:metabolism / Q000502:physiology; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics*
chemical_list: D006657:Histones; D060850:LIM-Homeodomain Proteins; C094898:Lhx3 protein; C497393:Mecp2 protein, mouse; D051783:Methyl-CpG-Binding Protein 2; D012333:RNA, Messenger; D014157:Transcription Factors
keywords: 5-aza-dc; 5′-aza-2′-deoxycytidine; CPHD; Chromatin; CpG; DNA methyltransferase; DNMT; FSHβ; GnRH-R; HDAC; LHX3; LIM-homeodomain protein 3; MBP; NF1; PRL; Promoter; SP1; TSA; TSS; Transcription; alpha glycoprotein subunit; combined pituitary hormone deficiency; cytosine-phosphate bond-guanine; follicle-stimulating hormone; gonadotropin-releasing hormone receptor; histone deacetylase; methyl-CpG binding protein; nuclear factor 1; prolactin; specificity protein 1; transcription start site; trichostatin A; αGSU