Mammalian (cytosine-5) methyltransferases cause genomic DNA methylation and lethality in Drosophila

Abstract

CpG methylation is essential for mouse development as well as gene regulation and genome stability. Many features of mammalian DNA methylation are consistent with the action of a de novo methyltransferase that establishes methylation patterns during early development and the post-replicative maintenance of these patterns by a maintenance methyltransferase. The mouse methyltransferase Dnmt1 (encoded by Dnmt) shows a preference for hemimethylated substrates in vitro, making the enzyme a candidate for a maintenance methyltransferase. Dnmt1 also has de novo methylation activity in vitro, but the significance of this finding is unclear, because mouse embryonic stem (ES) cells contain a de novo methylating activity unrelated to Dnmt1 (ref. 10). Recently, the Dnmt3 family of methyltransferases has been identified and shown in vitro to catalyse de novo methylation. To analyse the function of these enzymes, we expressed Dnmt and Dnmt3a in transgenic Drosophila melanogaster. The absence of endogenous methylation in Drosophila facilitates detection of experimentally induced methylation changes. In this system, Dnmt3a functioned as a de novo methyltransferase, whereas Dnmt1 had no detectable de novo methylation activity. When co-expressed, Dnmt1 and Dnmt3a cooperated to establish and maintain methylation patterns. Genomic DNA methylation impaired the viability of transgenic flies, suggesting that cytosine methylation has functional consequences for Drosophila development.

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

“…However, as DNMT1 expression is induced by CSE in A549 cells and DNMT1 accounts for >90% of the maintenance activity of DNA methylation, it is unlikely that DNMT1 is involved in the maintenance of SNCG methylation. DNMT3B possesses de novo DNA methyltransferase activity along with DNMT3A and unmethylated DNA is the preferred substrate for these two enzymes (Lyko et al, 1999). To examine the de novo methyltransferase activity of DNMT3B in controlling SNCG methylation status, H292 lung cancer cells in which the CpG island of SNCG is completely unmethylated were transfected with a DNMT3B expression vector.…”

“…DNMT1 is the most abundant form of DNMTs in mammalian cells. 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).…”

Dynamic and reversibility of heterochromatic gene silencing in human disease

“…DNMT3A and DNMT3B are highly expressed in ES cells, early embryos, and developing germ cells where de novo methylation takes place, but expressed at lower rate in somatic tissues of postnatal animals [22]. These enzymes have no preference for hemimethylated DNA [22,23] and can methylate de novo unmethylated CpGs when ectopically expressed in mammalian cells or transgenic flies [24,25]. DNMT3A has also been reported to methylate CpA sites [3].…”

Expression of the drosophila gene disconnected using the UAS/GAL4 system

“…The effects of Disco expression on the developing eye were examined by activating UAS-disco expression using GMR-GAL4 (Freeman, 1996) and eyeless-GAL4 (Lyko et al, 1999). Ectopic expression of disco posterior to the morphogenetic furrow using the GMR-GAL4 driver resulted in small, rough eyes lacking most pigmentation (Fig.…”