Maize Chromomethylase Zea methyltransferase2 Is Required for CpNpG Methylation

Abstract

A cytosine DNA methyltransferase containing a chromodomain, Zea methyltransferase2 (Zmet2), was cloned from maize. The sequence of ZMET2 is similar to that of the Arabidopsis chromomethylases CMT1 and CMT3 , with C-terminal motifs characteristic of eukaryotic and prokaryotic DNA methyltransferases. We used a reverse genetics approach to determine the function of the Zmet2 gene. Plants homozygous for a Mutator transposable element insertion into motif IX had a 13% reduction in methylated cytosines. DNA gel blot analysis of these plants with methylation-sensitive restriction enzymes and bisulfite sequencing of a 180-bp knob sequence showed reduced methylation only at CpNpG sites. No reductions in methylation were observed at CpG or asymmetric sites in heterozygous or homozygous mutant plants. Our research shows that chromomethylase Zmet2 is required for in vivo methylation of CpNpG sequences.

Hypomethylation of CNG targets induced with dihydroxypropyladenine is rapidly reversed in the course of mitotic cell division in tobacco

“…Almost 100% of CCG and 30-50% of CWG motifs could be demethylated in 5S rDNA repeats (Fulnecek et al 1998). The pattern of DNA hypomethylation induced with DHPA was very similar to that observed in chromomethylase mutants in Arabidopsis (Lindroth et al 2001) and maize (Papa et al 2001) in which the CNG sites underwent hypomethylation while methylation at the CG sites remained largely unaffected. Furthermore, outcrossing of a mutated Zmet2 locus resulted in partial restoration of DNA methylation patterns in maize (Papa et al 2001).…”

“…The pattern of DNA hypomethylation induced with DHPA was very similar to that observed in chromomethylase mutants in Arabidopsis (Lindroth et al 2001) and maize (Papa et al 2001) in which the CNG sites underwent hypomethylation while methylation at the CG sites remained largely unaffected. Furthermore, outcrossing of a mutated Zmet2 locus resulted in partial restoration of DNA methylation patterns in maize (Papa et al 2001). However, since the outcrossing procedure requires at least one meiotic cycle it was not possible to determine at which stage of plant development remethylation occurred.…”

“…3) Finally, it has to be considered that the de novo methylating capacities of CG and CNG methylation enzymes may differ. It has recently been established that CNG methylation in Arabidopsis (Lindroth et al 2001) and maize (Papa et al 2001) is accomplished by a group of specific enzymes called chromomethylases. Though a tobacco analog of these enzymes has not yet been isolated, it is likely that a similar gene is functional in tobacco.…”

Beyond Watson and Crick: DNA Methylation and Molecular Enzymology of DNA Methyltransferases

“…Chromomethylases have been shown to be responsible for methylation at CNG sites, which only occurs in plants and contributes to epigenetic gene silencing. [135,253,254] The abundance of DNA MTases in plants, as well as the presence of at least two independent methylation codes (CG and CNG), might be related to the fact that plants as sessile organisms cannot escape from unfavorable environmental conditions. Therefore, they require a fine adaptation of their genetic program to the microenvironment at each habitat.…”

Characterization of two rice DNA methyltransferase genes and RNAi-mediated reactivation of a silenced transgene in rice callus

“…Dnmt2 MTases contain only a methyltransferase domain and lack significant activity both in vivo and in vitro (Okano et al 1998). Whereas the Dnmt3 class contains putative de novo DNA MTases that are found in animals and plants (Okano et al 1999;Cao et al 2000;Robertson et al 2000;Cao and Jacobsen 2002), chromomethylases (CMTs) have thus far only been found in plants and are implicated in CpNpG methylation (Henikoff and Comai 1998;Bartee et al 2001;Papa et al 2001) and, possibly, in heterochromatin methylation.…”