Sequential order of target-recognizing domains in multispecific DNA-methyltransferases.

Wilke, Kathleen; Rauhut, E; Noyer-Weidner, Mario; Lauster, Roland; Pawlek, B.; Behrens, B.; Trautner, Thomas A.

doi:10.1002/j.1460-2075.1988.tb03110.x

Cited by 115 publications

(85 citation statements)

References 49 publications

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“…These mutated expression vectors can be tested in the same experimental system for de novo methyltransferase activity in vivo. It is known that a cysteine-to-serine alteration in the P-C motif of methyltransferases destroys the catalytic activity without compromising other functions of these proteins (18,30,31). A single replacement of cysteine by serine at position 706 of Dnmt3a protein and position 657 of Dnmt3b1 protein was made to alter this known catalytic site of cytosine methylases.…”

Section: Resultsmentioning

confidence: 99%

In Vivo Activity of Murine De Novo Methyltransferases, Dnmt3a and Dnmt3b

Hsieh

1999

Molecular and Cellular Biology

244

164

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The putative de novo methyltransferases, Dnmt3a and Dnmt3b, were reported to have weak methyltransferase activity in methylating the 3 long terminal repeat of Moloney murine leukemia virus in vitro. The activity of these enzymes was evaluated in vivo, using a stable episomal system that employs plasmids as targets for DNA methylation in human cells. De novo methylation of a subset of the CpG sites on the stable episomes is detected in human cells overexpressing the murine Dnmt3a or Dnmt3b1 protein. This de novo methylation activity is abolished when the cysteine in the P-C motif, which is the catalytic site of cytosine methyltransferases, is replaced by a serine. The pattern of methylation on the episome is nonrandom, and different regions of the episome are methylated to different extents. Furthermore, Dnmt3a also methylates the sequence methylated by Dnmt3a on the stable episome in the corresponding chromosomal target. Overexpression of human DNMT1 or murine Dnmt3b does not lead to the same pattern or degree of de novo methylation on the episome as overexpression of murine Dnmt3a. This finding suggests that these three enzymes may have different targets or requirements, despite the fact that weak de novo methyltransferase activity has been demonstrated in vitro for all three enzymes. It is also noteworthy that both Dnmt3a and Dnmt3b proteins coat the metaphase chromosomes while displaying a more uniform pattern in the nucleus. This is the first evidence that Dnmt3a and Dnmt3b have de novo methyltransferase function in vivo and the first indication that the Dnmt3a and Dnmt3b proteins may have preferred target sites.CpG methylation has been associated with reduced transcription (1, 13, 23), decreased DNase I sensitivity (14), and decreased site-specific recombination (8). Some of the effects of CpG methylation on chromatin structure are mediated by binding of a methylation binding protein (MeCP2) to methylated CpGs and recruitment of histone deacetylase (12,19). CpG methylation is tightly regulated during replication and differentiation of somatic cells. A maintenance methyltransferase (designated DNA methyltransferase 1 [DNMT1] in humans and Dnmt1 in mice) functions during DNA replication to preserve the methylated pattern of preexisting methylated region (16). In general, genes lose their CpG methylation within the promoter when they become activated, whereas genes acquire CpG methylation after they are no longer transcribed (for reviews, see references 2 and 22). It has been demonstrated recently that protein binding can lead to demethylation through a replication-dependent process within the specific binding sites (11,17). However, it remains unclear how regions of DNA acquire methylation in somatic cells (de novo methylation) or over what time intervals these changes occur, although many sequences have been reported to acquire methylation during development or the neoplastic process (25,26).De novo methylation is thought to establish the genomic CpG methylation pattern shortly after implantation in mammals ...

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Section: Resultsmentioning

confidence: 99%

In Vivo Activity of Murine De Novo Methyltransferases, Dnmt3a and Dnmt3b

Hsieh

1999

Molecular and Cellular Biology

244

164

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“…Interestingly, these phages carry genes encoding MTases that can recognize more than one target sequence due to the presence of multiple TRDs (53). Such multispecific MTases have been identified in the temperate Bacillus subtilis phages ø3T, p11B, and SP␤ (54), which each recognize two target sequences, while phage SPR carries a gene that encodes a type II MTase specific for three different recognition sites (55).…”

Section: Bacteriophage Mtases Possible Function and Prevalencementioning

confidence: 99%

Bacteriophage Orphan DNA Methyltransferases: Insights from Their Bacterial Origin, Function, and Occurrence

Murphy

Mahony

Ainsworth

et al. 2013

Appl Environ Microbiol

205

137

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“…The cysteine residue is conserved in all DNA methyltransferases, and its mutation to serine abolishes catalytic activity but does not affect DNA binding (32,41,61,62). We introduced the Cys12293Ser (C1229S) mutation into a functional minigene of mouse Dnmt1 composed of ϳ5 kb of cDNA and ϳ10 kb of genomic sequence that contains the regulatory sequence and the first three exons (56).…”

Section: Conservative Point Mutation In Dnmt1mentioning

confidence: 99%

Biological Functions of DNA Methyltransferase 1 Require Its Methyltransferase Activity

Damelin

Bestor

2007

Molecular and Cellular Biology

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DNA methyltransferase 1 (DNMT1) has been reported to interact with a wide variety of factors and to contain intrinsic transcriptional repressor activity. When a conservative point mutation was introduced at the key catalytic residue, mutant DNMT1 failed to rescue any of the phenotypes of Dnmt1-null embryonic stem (ES) cells, which indicated that the biological functions of DNMT1 are exerted through the methylation of DNA. ES cells that expressed the mutant protein did not survive differentiation. Intracisternal A-particle family retrotransposons were no longer methylated and were transcribed at high levels. The proper localization of DNMT1 depended on normal genomic methylation, and we discuss the implications of this finding for epigenetic dysregulation in cancer.Patterns of methylated and unmethylated cytosines at CpG dinucleotides are propagated with a fidelity of Ͼ99%, and their stable inheritance for Ͼ80 cell generations has been documented (48). DNA methylation is required for the transcriptional silencing of transposons, imprinted genes, and genes on the inactive X chromosome. The methylation landscape of the human genome reflects this function: methylation is concentrated in transposons and repeat sequences and is rare in promoter regions, except for certain imprinted genes, certain cancer-testis antigen genes, and genes on the inactive X chromosome in females (49). DNA methylation is also required for normal chromosome structure and stability. Demethylation and chromosome rearrangements at juxtacentromeric heterochromatin are observed in patients with immunodeficiency, centromere instability, and facial anomaly syndrome (OMIM 242860), which is caused by mutations in the DNA methyltransferase 3B (DNMT3B) gene (63).Cancer cells commonly exhibit aberrant genomic methylation patterns (3,20). The epigenetic dysregulation is complex, and the causes are not known. Hypomethylation of repeat sequences often coexists with hypermethylation of a subset of promoters. Hypomethylation and hypermethylation can independently predict tumor malignancy (21). A loss of imprinting increased the incidence of tumorigenesis (31, 51), and aberrant methylation had different effects on the progressions of different cancers (64).The mechanism of transcriptional repression of methylated DNA is not known. The chromatin-associated factors MeCP2, methyl-binding domain 1 (MBD1), MBD2, and Kaiso have been reported to repress methylated promoters, but methylated promoters are not reactivated in the absence of these factors, and the phenotypes of mice that lack these factors do not resemble those of mice that lack DNA methyltransferases (29,45,57,66).DNA methylation in mammals is catalyzed by three methyltransferases, DNMT1, DNMT3A, and DNMT3B (27). Biochemical and genetic data support a model in which all three enzymes have de novo and maintenance activities and cooperate to establish and maintain genomic methylation patterns. Of the methyltransferases, DNMT1 has the highest expression level in vivo and the highest specific activity in vi...

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Sequential order of target-recognizing domains in multispecific DNA-methyltransferases.

Cited by 115 publications

References 49 publications

In Vivo Activity of Murine De Novo Methyltransferases, Dnmt3a and Dnmt3b

In Vivo Activity of Murine De Novo Methyltransferases, Dnmt3a and Dnmt3b

Bacteriophage Orphan DNA Methyltransferases: Insights from Their Bacterial Origin, Function, and Occurrence

Biological Functions of DNA Methyltransferase 1 Require Its Methyltransferase Activity

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