Tài liệu Báo cáo Y học: Regulation of transcription of the Dnmt1 gene by Sp1 and Sp3 zinc finger

Regulation of transcription of the Dnmt1 gene by Sp1 and Sp3 zinc

finger proteins

Shotaro Kishikawa1,2, Takehide Murata1

, Hiromichi Kimura2

, Kunio Shiota2 and Kazunari K. Yokoyama1

1

Gene Engineering Division, Department of Biological Systems, BioResource Center, RIKEN (The Institute of Physical & Chemical

Research), Japan; 2

Department of Animal Resource Science/Veterinary Medical Sciences, The University of Tokyo, Japan

The Sp family is a family of transcription factors that

bind to cis-elements in the promoter regions of various

genes. Regulation of transcription by Sp proteins is based

on interactions between a GC-rich binding site

(GGGCGG)in DNA and C-terminal zinc finger motifs

in the proteins. In this study, we characterized the

GC-rich promoter of the gene for the DNA methyl￾transferase (Dnmt1)that is responsible for methylation of

cytosine residues in mammals and plays a role in gene

silencing. We found that a cis-element (nucleotides )161

to )147)was essential for the expression of the mouse

gene for Dnmt1. DNA-binding assays indicated that

transcription factors Sp1 and Sp3 bound to the same cis￾element in this region in a dose-dependent manner. In

Drosophila SL2 cells, which lack the Sp family of tran￾scription factors, forced expression of Sp1 or Sp3

enhanced transcription from the Dnmt1 promoter. Sti￾mulation by Sp1 and Sp3 were independent phenomena.

Furthermore, cotransfection reporter assays with a p300-

expression plasmid revealed the activation of the

promoter of the Dnmt1 gene in the presence of Sp3. The

transcriptional coactivator p300 interacted with Sp3 in

vivo and in vitro. Our results indicate that expression of

the Dnmt1 gene is controled by Sp1 and Sp3 and that

p300 is involved in the activation by Sp3.

Keywords:Dnmt1 gene; activation of transcription; Sp1; Sp3;

p300.

Transcription is regulated by the combinational actions of

proteins that bind to distinct promoter and enhancer

elements. In general, a limited number of cis-acting

DNA elements is recognized, not that by a single transcrip￾tion factor exclusively but, rather, by a set of different

proteins that are structurally related [1]. The promoter

regions of many eukaryotic genes contain GC-rich sequences

[2] and some of the most widely distributed promoter

elements are GC boxes and related motifs [2].

The Sp family of transcription factors includes the

proteins Sp1, Sp3 and Sp4, which recognize and bind to

GC boxes as well as to GT/A-rich motifs with similar

affinity, and Sp2, which binds preferentially to GT/A-rich

sequences [2,3]. Sp1 and Sp3 are expressed in a wide variety

of mammalian cells whereas Sp4 has been detected

predominantly in neuronal tissues. The regulation of gene

expression by Sp transcription factors is complex. Although

certain promoters can be activated by either Sp1 or Sp3 in

assays in vivo and are occasionally activated by both Sp1

and Sp3 that act in a synergistic manner [4–6], there are

other promoters that show a definite preference for Sp1 or

Sp3 [7]. Furthermore, Sp3 can function as an activator or a

repressor of transcription, depending on the gene in

question [8,9].

The genes for several mammalian activators and

repressors of transcription have been cloned. The gene

for p300 was first cloned as the gene for an E1A￾associated protein with properties of a transcriptional

adapter [10]. The protein was found later to possess

intrinsic histone acetyltransferase (HAT)activity and

to function as a coactivator in MyoD-, p53-, and SRC￾1-mediated transcription [11,12]. Furthermore, p300

appears to play a critical role in progression of the cell

cycle and the differentiation of cells [11,12].

The methylation of DNA plays a role in the regulation

of gene expression [13,14], genomic imprinting [15] and

inactivation of the X chromosome [16] and it has been

shown to be essential for mammalian development [17,18].

Altered patterns of DNA methylation has been implicated

in tumorigenesis [19]. However, the mechanisms by which

DNA methylation is regulated during development and

tumorigenesis remain largely unknown. Five distinct

families of gene for DNA methyltransferases, designated

Dnmt1, Dnmt2, Dnmt3a, Dnmt3b and Dnmt3L, have been

identified in mammalian cells [20]. Dnmt1 is expressed

constitutively in proliferating cells, it is associated with foci

of DNA replication [21] and methylates CpG dinucleo￾tides [22]. These findings are consistent with the hypothesis

that Dnmt1 is a maintenance methyltransferase that

restores appropriate patterns of DNA methylation to the

genome shortly after DNA replication [23]. Representative

sites for initiation of transcription have been found in the

promoter of the Dnmt1 gene, namely, an oocyte-specific

site, a somatic cell-specific site and a spermatocyte-specific

site. In adult somatic cells, most of the available data

indicate that the identification by Bigey et al. of many

Correspondence to K. K. Yokoyama, Gene Engineering Division,

Department of Biological Systems, BioResource Center, Tsukuba

Institute, RIKEN (The Institute of Physical & Chemical Research),

3-1-1 Koyadai, Tsukuba, Ibaraki 305-0074, Japan.

Fax: + 81 298 36 9120, Tel.: + 81 298 36 3612,

E-mail: [email protected]

Abbreviations: ChIP, chromatin immunoprecipitate; ODN,

oligodeoxynucleotides; HAT, histone acetyltransferase.

(Received 28 January 2002, revised 25 April 2002,

accepted 30 April 2002)

Eur. J. Biochem. 269, 2961–2970 (2002)
FEBS 2002 doi:10.1046/j.1432-1033.2002.02972.x