DNA methylation in wheat

Abstract

The origin and function of the large amount of 5-methylcytosine in plant DNA is not well understood. As a tool for in vitro studies of methylcytosine formation in plants we have isolated and characterized the DNA methyltransferase present in germinating wheat embryo. An enzyme fraction enriched 300-fold over the tissue homogenate was obtained by salt extraction of nuclei, chromatography on DEAE-cellulose, Sephadex (3-75, blue Sepharose and on DNA immobilized on cellulose. It catalyzes the methylation of cytosine residues in doublestranded DNAs isolated from wheat, maize, calf thymus or bacteria using S-adenosylmethionine as methyl donor. The efficient methylation of both an unmethylated plasmid DNA and its hemimethylated derivative indicate that the wheat DNA methylase can function de now and in maintenance methylation. A relative molecular mass of 50000 -55 000 was estimated by gel permeation chromatography and sucrose density gradient centrifugation. Polyacrylamide gel electrophoresis showed the presence of a protein of M , = 50000 and one other componentThe preference for endogenous, double-stranded DNA as substrate and the lower molecular mass distinguish wheat DNA methyltransferase from the DNA methylases obtained from mammalian sources. The properties of the wheat enzyme resemble, however, those of the DNA methylase isolated from the alga Chlamydomonas reinhardii, suggesting that plant cells possess their own type of DNA methyltransferase for the biosynthesis of their high methylcytosine content in DNA. ( M , = 35000).Plant DNA is distinguished from the DNA of all other organisms by its substantial content of the modified base, 5-methylcytosine, which can constitute up to 8 mo1/100 mol of the total base composition, or one-third of all cytosine residues [l, 21. The origin and functions of this high level of DNA methylation in plants are unknown. Vertebrates have much lower proportions of their cytosine residues in DNA methylated (approximately 0.9 mo1/100 mol). Here, changes in methylation are involved in the control of gene expression during development and cell differentiation, with a general correlation between undermethylation in active gene sequences and full methylation in non-transcribed regions [3, 41. Whether a similar signal function occurs in plant tissues is not well established; the above correlation seems to hold for zein genes, but not for the alcohol dehydrogenase gene of maize [5, 61. In any event it is difficult to conceive that the entire amount of msCyt in plants takes part in control mechanisms.In spite of the abundance of 5-methylcytosine in plants, its biosynthesis has not received much attention. A DNA methyltransferase was characterized in the green algae, Chlumydomonus reinhardii [7, 81 but because the unicellular Correspondence to H . Follmann,

Locus-specific control of asymmetric and CpNpG methylation by the DRM and CMT3 methyltransferase genes

“…These results are consistent with those of other studies of asymmetric methylation in plants, where CpA and CpT have been found in more abundance than CpC methylation (6,17,48). These results are also consistent with nearest neighbor analysis of plant DNA (5), and with studies of the in vitro substrate preference of purified plant methyltransferases (49,50). Interestingly, mouse embryonic stem cells also contain a significant amount CpA and CpT methylation and lower amounts of CpC methylation, and indirect evidence suggests an important role for Dnmt3 genes (the closest mammalian DRM homologs) in maintaining this methylation (2).…”

Light-Regulated Transcription of Genes Encoding Peridinin Chlorophyll a Proteins and the Major Intrinsic Light-Harvesting Complex Proteins in the DinoflagellateAmphidinium carterae Hulburt (Dinophycae)1

“…Our studies establish the presence of significant and variable levels of CpG and CpNpG methylation in dinoflagellates, suggesting the presence of a higher-plant-like methyltransferase(s) (Theiss et al, 1987). Our data are consistent with the hypothesis that methylation systems may have arisen as a means of noise reduction in organisms with large genomes (Bestor, 1990;Bird, 1995;Jablonka and Regev, 1995).…”

Purification and characterization of rice DNA methyltransferase

“…Up to now, only a few DNA MTase proteins have been purified from algae and plants [32,38,43]. Rice DNA MTase with a molecular weight of 54,000 had been previously purified from cultured rice cells with 380 purification folds [11] and showed strong preference on hemi-methylated DNA substrate.…”