title: Epigenetic characterization of the growth hormone gene identifies SmcHD1 as a regulator of autosomal gene clusters.
abstract: Regulatory elements for the mouse growth hormone (GH) gene are located distally in a putative locus control region (LCR) in addition to key elements in the promoter proximal region. The role of promoter DNA methylation for GH gene regulation is not well understood. Pit-1 is a POU transcription factor required for normal pituitary development and obligatory for GH gene expression. In mammals, Pit-1 mutations eliminate GH production resulting in a dwarf phenotype. In this study, dwarf mice illustrated that Pit-1 function was obligatory for GH promoter hypomethylation. By monitoring promoter methylation levels during developmental GH expression we found that the GH promoter became hypomethylated coincident with gene expression. We identified a promoter differentially methylated region (DMR) that was used to characterize a methylation-dependent DNA binding activity. Upon DNA affinity purification using the DMR and nuclear extracts, we identified structural maintenance of chromosomes hinge domain containing -1 (SmcHD1). To better understand the role of SmcHD1 in genome-wide gene expression, we performed microarray analysis and compared changes in gene expression upon reduced levels of SmcHD1 in human cells. Knock-down of SmcHD1 in human embryonic kidney (HEK293) cells revealed a disproportionate number of up-regulated genes were located on the X-chromosome, but also suggested regulation of genes on non-sex chromosomes. Among those, we identified several genes located in the protocadherin β cluster. In addition, we found that imprinted genes in the H19/Igf2 cluster associated with Beckwith-Wiedemann and Silver-Russell syndromes (BWS & SRS) were dysregulated. For the first time using human cells, we showed that SmcHD1 is an important regulator of imprinted and clustered genes.
authors.affiliation: Faculty of Health Sciences, Simon Fraser University, Burnaby, BC, Canada.
authors.forename: Shabnam
authors.initials: S
authors.lastname: Massah
authors.affiliation: Faculty of Health Sciences, Simon Fraser University, Burnaby, BC, Canada.
authors.forename: Robert
authors.initials: R
authors.lastname: Hollebakken
authors.affiliation: Faculty of Health Sciences, Simon Fraser University, Burnaby, BC, Canada.
authors.forename: Mark P
authors.initials: MP
authors.lastname: Labrecque
authors.affiliation: Faculty of Biology, Ludwig Maximilians University Munich, Martinsried, Germany.
authors.forename: Addie M
authors.initials: AM
authors.lastname: Kolybaba
authors.affiliation: Faculty of Health Sciences, Simon Fraser University, Burnaby, BC, Canada.
authors.forename: Timothy V
authors.initials: TV
authors.lastname: Beischlag
authors.affiliation: Faculty of Health Sciences, Simon Fraser University, Burnaby, BC, Canada.
authors.forename: Gratien G
authors.initials: GG
authors.lastname: Prefontaine
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001374:Azacitidine / Q000494:pharmacology; D001483:Base Sequence; D001506:Beckwith-Wiedemann Syndrome / Q000235:genetics; D015820:Cadherins / Q000235:genetics; D002460:Cell Line; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D033481:Chromosomes, Mammalian / Q000235:genetics*; D019175:DNA Methylation / Q000187:drug effects; D044127:Epigenesis, Genetic / Q000187:drug effects; D013006:Growth Hormone / Q000235:genetics*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D056730:Silver-Russell Syndrome / Q000235:genetics; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D015820:Cadherins; D002868:Chromosomal Proteins, Non-Histone; C578241:SMCHD1 protein, human; C529024:SmcHD1 protein, mouse; D013006:Growth Hormone; D001374:Azacitidine