One of the things I wanted to do on this blog was to highlight recent plant science journal articles, and when I found back-to-back papers on Arabidopsis research in Science I thought they would be a good place to start.
But when I started to read, I realised the obvious – my cell wall biochemistry background will be no help at all when trying to understand other areas of plant science research. But I still want to highlight high-impact articles like this on the blog, so I decided to have a Summary for non-specialists series. Please feel free to comment if I’ve got something horribly wrong, and of course if anyone would like to provide a Summary by a Specialist that would be great!
Qian et al., June 2012. A Histone Acetyltransferase Regulates Active DNA Demethylation in Arabidopsis. Science 336: 1445-1447
Prior to this research, little was known about the regulation of DNA methylation, or how DNA and histone modifications were related. Here, Qian et al. define a process in which histone modification is an essential part of DNA methylation. This research opens the door to deeper understanding of the regulation and mechanism of DNA modification, and possible manipulation of epigenetic mechanisms.
(Relevant) basic introduction to epigenetics
Information gleaned from this review: Law and Jacobsen (2010) Nature Reviews Genetics.
Broadly speaking, DNA is arranged around histones and compacted into chromatin. Gene expression is affected by the physical properties of the DNA, histones and chromatin within this general superstructure.
In eukaryotes there are 5 histone types. Histone H3 is the most heavily modified. Positions on histones are defined and named, for example H3K18 (Histone H3, position K18).
Both DNA and histones can be methylated or unmethylated. If DNA is methylated, it is ‘switched off’. If a histone is methylated, the DNA on it is ‘switched on.’
DNA methyltransferase enzymes methylate DNA, and demethylation of DNA is carried out by DNA glycolases. Both processes are RNA-directed. Many of the enzymes and pathways involved are characterized, but this is a fairly new field so there is still more to uncover.
The mechanism of DNA de/methylation regulation, and how or if it is related to histone modification or chromatin structure, is still largely unknown.
The researchers screened Arabidopsis T-DNA insertion mutants for hypermethylation on a specific gene (DT-77, AT1G26400) on the genome. They identified two lines in which this gene was highly methylated. Both lines had the T-DNA insertion in the same gene, which they named INCREASED DNA METHYLATION 1 (IDM1).
Alongside their analysis of the newly found IDM1, the team analysed DNA glycolase ROS1, an enzyme known to be part of the DNA demethylation pathway.
Both the idm1 and ros1 mutants had hypermethylated DNA at more than a thousand loci, while only around a hundred loci showed less methylation compared to the wildtype. However the idm1-ros1 double mutant did not show additive effects. ROS1 expression was not reduced in idm1 and the proteins do not co-localize. Transcripts from both genes were reduced in Arabidopsis mutants with highly unmethylated DNA. At this stage, the researchers could hypothesise that ROS1 and IDM1 were part of the same demethylation pathway.
The next phase of research was characterising the structure of IDM1. There are three major predicted domains:
- The MBD domain binds to Histone H3. Binding occurs at areas of low histone methylation because of …
- … the PHD finger, which specifically binds the N-terminal of Histone H3 at an unmethylated H3K4 domain. Binding is inhibited by H3K4 or H3R2 methylation. This binding activity was confirmed by in vitro affinity pull-down assays.
- The N-acetyltransferase domain, with histone acetyl transferase (HAT) activity. It acetylates positions H3K18 and H3K23.
All three domains were confirmed to be necessary for IDM1 activity. The team attempted to rescue the idm1 mutant line with fragments of IDM1 MBD, PHD or HAT domains, but none of the fragments could reduce levels of hypermethylation idm1.
IDM1 is a regulator of demethylation by the ROS1 subfamily of DNA glycosylases. It recognises histone H3 by its PHD finger and binds to it by the MBD. The N-acetyl transferase region acetylates H3K18 and H3K23. The acetylated histone regions are tags marking the DNA on the histone for demethylation by ROS and its cohort.