title: MeCP2 knockdown reveals DNA methylation-independent gene repression of target genes in living cells and a bias in the cellular location of target gene products.
abstract: MeCP2, a methyl-CpG binding domain (MBD) protein, is known to bind to methylated CpG sites via a conserved MBD, leading to transcriptional repression. However, studies in cell-free system for gene repression and MeCP2 binding have suggested that DNA methylation-independent repression also occurs in living cells. It has been difficult to characterize the target genes of MeCP2 because a limited number have been identified to date. In this context, we screened for MeCP2 target genes using knockdown (KD) experiments combined with microarray gene expression analyses. Of the 49 genes that showed a more than three-fold increase in expression in two independent KD experiments conducted with different siRNA sets, unexpectedly, half (24 genes) did not contain promoter CpG islands (CGIs). Of seven selected genes that did contain CGIs, only two were methylated at the CGI, bound MeCP2 before KD, and reduced MeCP2 after KD. For three, MeCP2 was observed to bind to the unmethylated CGI before KD, and for one MeCP2 was reduced after KD. Another two genes neither had DNA methylation nor bound MeCP2 before KD. Gene ontology analysis suggested that MeCP2 represses a certain group of genes. These results suggest that in addition to the canonical gene repression function, MeCP2 can repress gene expression by binding to unmethylated DNA in particular genes in living cells.
authors.affiliation: Department of Biomolecular Sciences, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Shinya
authors.initials: S
authors.lastname: Yakabe
authors.forename: Hidenobu
authors.initials: H
authors.lastname: Soejima
authors.forename: Hitomi
authors.initials: H
authors.lastname: Yatsuki
authors.forename: Hirotaka
authors.initials: H
authors.lastname: Tominaga
authors.forename: Wei
authors.initials: W
authors.lastname: Zhao
authors.forename: Ken
authors.initials: K
authors.lastname: Higashimoto
authors.forename: Keiichiro
authors.initials: K
authors.lastname: Joh
authors.forename: Shinichi
authors.initials: S
authors.lastname: Kudo
authors.forename: Kohji
authors.initials: K
authors.lastname: Miyazaki
authors.forename: Tsunehiro
authors.initials: T
authors.lastname: Mukai
mesh_terms: D002460:Cell Line; D018899:CpG Islands; D019175:DNA Methylation; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D006801:Humans; D051783:Methyl-CpG-Binding Protein 2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D012097:Repressor Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D018875:Vocabulary, Controlled
chemical_list: D051783:Methyl-CpG-Binding Protein 2; D012097:Repressor Proteins