The post describes the work of Lu Yang, who just completed her PhD, and is now doing a post-doc in Germany
Lu’s recent study relates to one particular epigenetic mark, the addition of a methyl groups (CH3) to Cytosine (the C residues in the sequences of DNA). Epigenetic marks like this are known to be associated with turning genes on and off but how? We asked whether the methyl group could block the binding of a protein that was required to regulate genes.
To make it simple, picture the gene regulatory protein we work on (it’s called GATA1 since it binds the bases G-A-T-A in DNA), as a pigeon. Imagine the human genes as all being sections of a clothesline – the chromosome. Basically, we were asking the question – if you put on a methyl group, a clothes peg, at the beginning of one section of the clothesline to make meC-G-A-T-A, will that prevent the pigeon from landing?
The answer is – yes, it will. That’s it. Methylation can protect the gene from being turned on or off. If you remove the methylation, then the gene can be regulated.
Lu did her experiments in the context of blood stem cells differentiating into red blood cells. The methylation of DNA changes as the stem cells gradually mature into red blood cells and different genes become available for regulation by GATA1. Understanding this helps explain how biology works, but also how we might make artificial genetic switches.
What’s most surprising is that no one had discovered the sensitivity of GATA1 to methylation before. I remember that when I was an under-graduate I was taught about DNA-binding proteins that were unable to bind DNA when it was methylated. For example, restriction enzymes that recognize and then cut DNA can be blocked by methyl groups. Oddly, very few human DNA-binding proteins have been discovered to be sensitive to methylation. There are very few, if any other examples, where a single methylation site blocks the binding of a key gene regulatory protein.
The reason for the lack of work in this area is partly that until the development of CRISPR, it was impossible to test even the simple hypothesis that one methylation site influenced gene expression. CRISPR really has changed everything.
Lu’s work involved traditional experiments in vitro and then the analysis of genetically modified mice that were made here at UNSW by Lars Itner’s group, using CRISPR gene editing. Lu first showed that in a test tube the methylation of the C in a C-G-A-T-A sequence in DNA blocks the binding of GATA1. She then showed that in red blood cell lines grown in petri dishes that GATA1 could bind and regulate certain genes from unmethylated C-G-A-T-A elements but not from elements modified by the addition of a methyl group on the C. Then she looked at the modified mice that had a single mutation in one key C-G-A-T-A element in an important red blood cell gene. The blood cell profile in the mice was abnormal. This implied that in a living mouse a single methylated C residue was absolutely required for normal gene regulation across blood cell maturation.
So, she effectively showed that a clothes peg on a certain gene can prevent at least one type of bird, a pigeon, or the regulatory protein GATA1, from landing and regulating one gene. The work is a nice example of why methylation affects which genes are turned on or off, as it shows that methylation can prevent gene regulatory proteins from accessing their target genes. This is a good result and could be a nice textbook example.
A lot has been written about epigenetics. The field has captured the public imagination because it smacks of Lamarckian inheritance, the idea that environmental effects (including the addition of methyl groups to DNA in response to diet, stress, aging or just normal development) can influence our genes and possibly the genes we inherited or those we pass on to our offspring. Having a more precise understanding of how DNA methylation at specific genes affects gene regulation and cellular differentiation will ultimately enable us to understand the contribution of epigenetics to human development and evolution and will help in efforts to artificially regulate genes for therapeutic or agricultural purposes.
Our lab is particularly interested in boosting the output of the foetal globin gene in order to alleviate Sickle Cell Disease. It is known that methylation reduces the foetal globin output and we are now exploring whether our results help explain that phenomenon.
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