Methylation-mediated transcriptional silencing in euchromatin by methyl- CpG binding protein MBD1 isoforms

Naoyuki Fujita, Shin Ichiro Takebayashi, Katsuzumi Okumura, Shinichi Kudo, Tsutomu Chiba, Hideyuki Saya, Mitsuyoshi Nakao

Research output: Contribution to journalArticle

150 Citations (Scopus)

Abstract

DNA methylation of promoter-associated CpG islands is involved in the transcriptional repression of vertebrate genes. To investigate the mechanisms underlying gene inactivation by DNA methylation, we characterized a human MBD1 protein, one of the components of MeCP1, which possesses a methyl-CpG binding domain (MBD) and cysteine-rich (CXXC) domains. Four novel MBD1 isoforms (MBD1v1, MBD1v2, MBD1v3, and MBD1v4) were identified by the reverse transcription-PCR method. We found that these transcripts were alternatively spliced in the region of CXXC domains and the C terminus. Green fluorescent protein-fused MBD1 was localized to multiple foci on the human genome, mostly in the euchromatin regions, and particularly concentrated in the pericentromeric region of chromosome 1. Both the MBD sequence and genome methylation were required for proper localization of the MBD1 protein. We further investigated whether MBD1 isoforms are responsible for transcriptional repression of human genes. A bacterially expressed MBD1 protein bound preferentially to methylated DNA fragments containing CpG islands from the tumor suppressor genes p16, VHL, and E-cadherin and from an imprinted SNRPN gene. All MBD1 isoforms inhibited promoter activities of these genes via methylation. Interestingly, MBD1 isoforms v1 and v2 containing three CXXC domains also suppressed unmethylated promoter activities in mammalian cells. These effects were further manifested in Drosophila melanogaster cells, which lack genome methylation. Sp1-activated transcription of methylated p16 and SNRPN promoters was inhibited by all of the MBD1 isoforms, whereas the isoforms v1 and v2 reduced Sp1-activated transcription from unmethylated promoters as well. These findings suggested that the MBD1 isoforms have different roles in methylation-mediated transcriptional silencing in euchromatin.

Original languageEnglish
Pages (from-to)6415-6426
Number of pages12
JournalMolecular and Cellular Biology
Volume19
Issue number9
Publication statusPublished - 1999 Sep
Externally publishedYes

Fingerprint

Euchromatin
Methylation
Carrier Proteins
Protein Isoforms
snRNP Core Proteins
CpG Islands
DNA Methylation
Genes
Genome
Chromosomes, Human, Pair 1
Gene Silencing
Human Genome
Cadherins
Green Fluorescent Proteins
Tumor Suppressor Genes
Drosophila melanogaster
Reverse Transcription
Cysteine
Vertebrates
Proteins

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Cell Biology

Cite this

Fujita, N., Takebayashi, S. I., Okumura, K., Kudo, S., Chiba, T., Saya, H., & Nakao, M. (1999). Methylation-mediated transcriptional silencing in euchromatin by methyl- CpG binding protein MBD1 isoforms. Molecular and Cellular Biology, 19(9), 6415-6426.

Methylation-mediated transcriptional silencing in euchromatin by methyl- CpG binding protein MBD1 isoforms. / Fujita, Naoyuki; Takebayashi, Shin Ichiro; Okumura, Katsuzumi; Kudo, Shinichi; Chiba, Tsutomu; Saya, Hideyuki; Nakao, Mitsuyoshi.

In: Molecular and Cellular Biology, Vol. 19, No. 9, 09.1999, p. 6415-6426.

Research output: Contribution to journalArticle

Fujita, N, Takebayashi, SI, Okumura, K, Kudo, S, Chiba, T, Saya, H & Nakao, M 1999, 'Methylation-mediated transcriptional silencing in euchromatin by methyl- CpG binding protein MBD1 isoforms', Molecular and Cellular Biology, vol. 19, no. 9, pp. 6415-6426.
Fujita, Naoyuki ; Takebayashi, Shin Ichiro ; Okumura, Katsuzumi ; Kudo, Shinichi ; Chiba, Tsutomu ; Saya, Hideyuki ; Nakao, Mitsuyoshi. / Methylation-mediated transcriptional silencing in euchromatin by methyl- CpG binding protein MBD1 isoforms. In: Molecular and Cellular Biology. 1999 ; Vol. 19, No. 9. pp. 6415-6426.
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