Dnmt2-dependent methylomes lack defined DNA methylation patterns

Abstract

Several organisms have retained methyltransferase 2 (Dnmt2) as their only candidate DNA methyltransferase gene. However, information about Dnmt2-dependent methylation patterns has been limited to a few isolated loci and the results have been discussed controversially. In addition, recent studies have shown that Dnmt2 functions as a tRNA methyltransferase, which raised the possibility that Dnmt2-only genomes might be unmethylated. We have now used whole-genome bisulfite sequencing to analyze the methylomes of Dnmt2-only organisms at single-base resolution. Our results show that the genomes of Schistosoma mansoni and Drosophila melanogaster lack detectable DNA methylation patterns. Residual unconverted cytosine residues shared many attributes with bisulfite deamination artifacts and were observed at comparable levels in Dnmt2-deficient flies. Furthermore, genetically modified Dnmt2-only mouse embryonic stem cells lost the DNA methylation patterns found in wild-type cells. Our results thus uncover fundamental differences among animal methylomes and suggest that DNA methylation is dispensable for a considerable number of eukaryotic organisms.

Genome methylation in D. melanogaster is found at specific short motifs and is independent of DNMT2 activity

“…The relative rarity of methylation in Drosophila, even at stage 5 where it has been shown to be most abundant (Lyko et al 2000a), has made it a subject of controversy. Some have recently suggested that it may be absent, or its presence is spurious and a reflection of nonspecific activity by MT2 (Marhold et al 2004;Schaefer and Lyko 2010a;Raddatz et al 2013); the near-normal phenotype of flies lacking functional MT2 has supported this point of view (Kunert et al 2003;Phalke et al 2009;Schaefer and Lyko 2010b). Although we find that methylation can be present on a substantial proportion of alleles within methylated regions, methylation is virtually never present on more than half of the alleles.…”

“…Inactivation of Mt2 does not visibly alter embryonic development (Kunert et al 2003), although it can suppress position-effect variegation (Phalke et al 2009). No clear pattern of genomic cytosine methylation has been established by any study, and genomewide bisulfite sequencing studies of wild type Drosophila embryos at an average depth of 5.6-fold (Zemach et al 2010) and 32-fold (Raddatz et al 2013) did not report methylated cytosines. The finding that MT2 can methylate tRNAs has led to speculation that its primary function in Drosophila is RNA methylation (Goll et al 2006;Schaefer et al 2010;Durdevic et al 2013), and that genomic cytosine methylation is random and spurious (Schaefer and Lyko 2010b).…”

“…The second study (Raddatz et al 2013) has sufficient coverage for most of the resequenced regions (Supplemental Table S3; Supplemental Fig. S7); only three of the regions with sufficient coverage show no evidence for methylation in that data set.…”

“…This indicates that such regions could be detected by shotgun bisulfite sequencing if sufficient sequencing depth were achieved. Two previously published studies used whole genome bisulfite sequencing to conclude that methylation is absent from the Drosophila genome (Zemach et al 2010;Raddatz et al 2013).…”

“…Despite this, the pattern and function of cytosine methylation in Drosophila has never been clear, and recently considerable doubt has been expressed as to the presence of any significant quantity of genomic methylation (Goll et al 2006;Schaefer and Lyko 2010a;Zemach et al 2010;Raddatz et al 2013). Inactivation of Mt2 does not visibly alter embryonic development (Kunert et al 2003), although it can suppress position-effect variegation (Phalke et al 2009).…”