Biochemistry and biology of mammalian DNA methyltransferases

Hermann, Andrea; Gowher, Humaira; Jeltsch, Albert

doi:10.1007/s00018-004-4201-1

Cited by 479 publications

(371 citation statements)

References 182 publications

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“…Among Sp family proteins, Sp1 and Sp3 were shown to be ubiquitously expressed in human tissues, and transcriptional activity enhanced by Sp1 was shown to be suppressed by Sp3 (32). Therefore, in such genes, CpG motifs may play an essential role in gene expression, especially in the expression of the DR3 gene (33,34). This idea needs further investigation.…”

Section: Discussionmentioning

confidence: 99%

Hypermethylated promoter region of DR3, the death receptor 3 gene, in rheumatoid arthritis synovial cells

Takami

Osawa

Miura

et al. 2006

Arthritis & Rheumatism

138

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Objective. To examine the promoter activity and protein expression of the death receptor 3 gene DR3, a member of the apoptosis-inducing Fas gene family, with particular reference to the methylation status of its promoter region in rheumatoid arthritis (RA).Methods. Genomic DNA was prepared from peripheral blood mononuclear cells obtained from healthy individuals and from patients with RA and synovial cells obtained from patients with RA and osteoarthritis. The methylation status of the DR3 promoter was analyzed by bisulfite genomic sequencing and methylation-specific polymerase chain reaction techniques. Gene promoter activity and protein expression were examined using the luciferase reporter and Western blotting techniques.Results. The promoter region of the DR3 gene contained many CpG motifs, including one CpG island that was specifically hypermethylated in synovial cells from patients with RA. Promoter assays showed that the promoter CpG island was essential for the transactivation of the DR3 gene and that forced hypermethylation of the CpG island with the bacterial methylase Sss I in vitro resulted in inhibition of the DR3 gene expression.

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Section: Discussionmentioning

confidence: 99%

Hypermethylated promoter region of DR3, the death receptor 3 gene, in rheumatoid arthritis synovial cells

Takami

Osawa

Miura

et al. 2006

Arthritis & Rheumatism

138

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“…Another process by which epigenetic modifications directly effect gene expression and chromatin organization is methylation of DNA at cytosine residues by DNA methyltransferases (DNMTs) (for reviews, see Refs. [5,10]). Several reports have detailed how deregulation of methylation can be the direct cause or result of cellular dysfunction.…”

Section: Hhmi Author Manuscriptmentioning

confidence: 99%

“…Small organic molecules and peptides and most types of cellular macromolecules, including DNA, RNA, protein, and lipids, can serve as substrates for methylation (for detailed reviews, see Refs. [2][3][4][5][6][7]). …”

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confidence: 99%

A universal competitive fluorescence polarization activity assay for S-adenosylmethionine utilizing methyltransferases

Graves

Zhang

Scott

2008

Analytical Biochemistry

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A high-throughput, competitive fluorescence polarization immunoassay has been developed for the detection of methyltransferase activity. The assay was designed to detect Sadenosylhomocysteine (AdoHcy), a product of all S-adenosylmethionine (AdoMet)-utilizing methyltransferase reactions. We employed commercially available anti-AdoHcy antibody and fluorescein-AdoHcy conjugate tracer to measure AdoHcy generated as a result of methyltransferase activity. AdoHcy competes with tracer in the antibody/tracer complex. The release of tracer results in a decrease in fluorescence polarization. Under optimized conditions, AdoHcy and AdoMet titrations demonstrated that the antibody had more than a 150-fold preference for binding AdoHcy relative to AdoMet. Mock methyltransferase reactions using both AdoHcy and AdoMet indicated that the assay tolerated 1 to 3 μM AdoMet. The limit of detection was approximately 5 nM (0.15 pmol) AdoHcy in the presence of 3 μM AdoMet. To validate the assay's ability to quantitate methyltransferase activity, the methyltransferase catechol-Omethyltransferase (COMT) and a known selective inhibitor of COMT activity were used in proofof-principle experiments. A time-and enzyme concentration-dependent decrease in fluorescence polarization was observed in the COMT assay that was developed. The IC 50 value obtained using a selective COMT inhibitor was consistent with previously published data. Thus, this sensitive and homogeneous assay is amenable for screening compounds for inhibitors of methyltransferase activity. KeywordsFluorescence polarization; Methyltransferase; S-Adenosylhomocysteine; S-Adenosylmethionine; Catechol-O-methyltransferase Methyltransferases are a diverse class of enzymes that catalyze the transfer of a single methyl group from a methyl donor molecule to a methyl acceptor. The vast majority use Sadenosylmethionine (AdoMet) 1 as the methyl donor [1,2]. AdoMet is the second most HHMI Author Manuscript HHMI Author Manuscript HHMI Author Manuscriptwidely used substrate molecule in the cell after ATP. Methylation is an essential and highly choreographed regulatory metabolic process. Small organic molecules and peptides and most types of cellular macromolecules, including DNA, RNA, protein, and lipids, can serve as substrates for methylation (for detailed reviews, see Refs. [2][3][4][5][6][7]).A rapidly growing area of methyltransferase research focuses on methylation of histone tails by histone methyltransferases (HMTs) (for reviews, see Refs. [8,9]). Histone methylation is an epigenetic modification that has a role in formation of heterochromatin, X chromosome inactivation, and modification of gene expression patterns. Histone methylation occurs on arginine and lysine residues and is catalyzed by methyltransferases belonging to three distinct families of proteins: the protein arginine N-methyltransferase 1 (PRMT1) family, the SET-domain-containing protein family, and the non-SET-domain proteins DOT1/DOT1L. This histone methylation alters chromatin structure and, thus, regulates...

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“…DNMT1 functions primarily in maintenance methylation during DNA replication using hemimethylated DNA as substrates (Lyko et al, 1999;Robertson et al, 1999;Rountee et al, 2000;Robert et al, 2003;Suzuki et al, 2004). The reactions of de novo methylation using unmethylated DNA are thought to be primarily catalysed by DNMT3A and DNMT3B in mammals during early development (Okano et al, 1999;Bachman et al, 2001;Brueckner and Lyko, 2004;Hermann et al, 2004;Dodge et al, 2005).…”

Section: Introductionmentioning

confidence: 99%

Cigarette smoke induces demethylation of prometastatic oncogene synuclein-γ in lung cancer cells by downregulation of DNMT3B

et al. 2007

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The prometastatic oncogene synuclein-c (SNCG) is not expressed in normal lung tissues, but it is highly expressed in lung tumors. Here, we show that cigarette smoke extract (CSE) has strong inducing effects on SNCG gene expression in A549 lung cancer cells through demethylation of SNCG CpG island. CSE treatment also augments the invasive capacity of A549 cells in an SNCG-dependent manner. To elucidate the mechanisms underlying the demethylating effects of CSE, we examined expression levels of DNA methyltransferases (DNMTs), 1, 3A and 3B in CSE-treated cells. We show that the mRNA expression of DNMT3B is specifically downregulated by CSE with a kinetics concurrent to SNCG reexpression. Utilizing siRNA to knockdown DNMT3B expression, we show that inhibition of DNMT3B directly increases SNCG mRNA expression. We further show that exogenous overexpression of DNMT3B in an SNCG-positive lung cancer cell line H292 suppresses SNCG mRNA and protein expression and induces de novo methylation of SNCG CpG island, whereas overexpression of DNMT1 or DNMT3A has no effects. Taken together, these new findings demonstrate that tobacco exposure induces the abnormal expression of SNCG in lung cancer cells through downregulation of DNMT3B. This work sheds light on the molecular understanding of demethylation of this oncogene during cancer progression.

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Biochemistry and biology of mammalian DNA methyltransferases

Cited by 479 publications

References 182 publications

Hypermethylated promoter region of DR3, the death receptor 3 gene, in rheumatoid arthritis synovial cells

Hypermethylated promoter region of DR3, the death receptor 3 gene, in rheumatoid arthritis synovial cells

A universal competitive fluorescence polarization activity assay for S-adenosylmethionine utilizing methyltransferases

Cigarette smoke induces demethylation of prometastatic oncogene synuclein-γ in lung cancer cells by downregulation of DNMT3B

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