title: CHD7 functions in the nucleolus as a positive regulator of ribosomal RNA biogenesis.
abstract: De novo mutation of the gene encoding chromodomain helicase DNA-binding protein 7 (CHD7) is the primary cause of CHARGE syndrome, a complex developmental disorder characterized by the co-occurrence of a specific set of birth defects. Recent studies indicate that CHD7 functions as a transcriptional regulator in the nucleoplasm. Here, we report based on immunofluorescence and western blotting of subcellular fractions that CHD7 is also constitutively localized to the nucleolus, the site of rRNA transcription. Standard chromatin immunoprecipitation (ChIP) assays indicate that CHD7 physically associates with rDNA, a result that is also observable upon alignment of whole-genome CHD7 ChIP coupled with massively parallel DNA sequencing data to the rDNA reference sequence. ChIP-chop analyses demonstrate that CHD7 specifically associates with hypomethylated, active rDNA, suggesting a role as a positive regulator of rRNA synthesis. Consistent with this hypothesis, siRNA-mediated depletion of CHD7 results in hypermethylation of the rDNA promoter and a concomitant reduction of 45S pre-rRNA levels. Accordingly, cells overexpressing CHD7 show increased levels of 45S pre-rRNA compared with control cells. Depletion of CHD7 also reduced cell proliferation and protein synthesis. Lastly, compared with wild-type ES cells, the levels of 45S pre-rRNA are reduced in both Chd7(+/-) and Chd7(-/-) mouse ES cells, as well as in Chd7(-/-) whole mouse embryos and multiple tissues dissected from Chd7(+/-) embryos. Together with previously published studies, these results indicate that CHD7 dually functions as a regulator of both nucleoplasmic and nucleolar genes and provide a novel avenue for investigation into the pathogenesis of CHARGE syndrome.
authors.affiliation: Department of Genetics, Case Western Reserve University, Cleveland, OH 44106, USA.
authors.forename: Gabriel E
authors.initials: GE
authors.lastname: Zentner
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Hurd
authors.forename: Michael P
authors.initials: MP
authors.lastname: Schnetz
authors.forename: Lusy
authors.initials: L
authors.lastname: Handoko
authors.forename: Chuanping
authors.initials: C
authors.lastname: Wang
authors.forename: Zhenghe
authors.initials: Z
authors.lastname: Wang
authors.forename: Chialin
authors.initials: C
authors.lastname: Wei
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Tesar
authors.forename: Maria
authors.initials: M
authors.lastname: Hatzoglou
authors.forename: Donna M
authors.initials: DM
authors.lastname: Martin
authors.forename: Peter C
authors.initials: PC
authors.lastname: Scacheri
mesh_terms: D000818:Animals; D058747:CHARGE Syndrome / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002466:Cell Nucleolus / Q000235:genetics / Q000378:metabolism*; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D019175:DNA Methylation; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D011485:Protein Binding; D021381:Protein Transport; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012335:RNA, Ribosomal / Q000235:genetics* / Q000378:metabolism
chemical_list: C507367:Chd7 protein, mouse; D004268:DNA-Binding Proteins; D012322:RNA Precursors; D012335:RNA, Ribosomal; D004265:DNA Helicases; C491434:CHD7 protein, human