Methytransferases and RNAi Pathways
DNA methyltransferase
- MET1 DNA methyltransferase
- DRM1 and DRM2 DNA methyltransferase
- DRM3 DNA methytransferase
MET1 is responsible for maintaining CG methylation
DRM2 is the primary enzyme implicated in the RNAi mediated methylation pathway. Other The enzyme constantly targets CHH sites (Widman et. al, 2009). DRM2 is expressed at higher levels than DRM1 and is the main enzyme involved in de novo methylation (Meyer, 2005).
Henderson et. al (2006) discovered that DRM3 is required for non-CG DNA methylation. They tested two T-DNA insertion strains (drm3-1 and drm3-2), finding a reduction of methylation of MEA-ISR. After performing a southern blot on the Msp1 (methyl-sensitive restriction enzyme) digested MEA-ISR repeat and sequencing samples of this DNA using the bisulfite method, the researchers concluded that DRM3 is necessary for proper DRM2 functioning.
(A) Southern blots hybridized for the MEA-ISR repeat using DNA digested with the methyl-sensitive restriction enzyme MspI. The 4.3 kb band and 2 kb bands represent digested and undigested DNA respectively. (B) Graphical representation of bisulfite sequencing at MEA-ISR with % cytosine methylation shown for each genotype. CG sequence contexts are shaded black, CHG are shaded grey and white represents CHH. The 95% confidence limits shown are given by the Wilson score interval. (C) Graphical representation of bisulfite sequencing at the methylated promoter repeats of the FWA endogene with % cytosine methylation shown for each genotype and shaded according to sequence context as in (B). (D) Quantitative PCR measurement of AtSN1 genomic DNA following restriction endonuclease digestion with DNA methylation sensitive enzyme HaeIII. The value on the y-axis represents the ratio of DNA measured by quantitative PCR with and without HaeIII digestion. The data represents three biological replicates and the error bars represent standard error.
