DNA Modifications in Breast Cancer

General Background: Epigenetics
Our genetic material, DNA, encodes all the genes that are required to make and maintain our development from a tiny embryo right through to adulthood. It is stored in a special cellular compartment, the nucleus. Each individual is born with the same unique set of genes in almost every cell in their body. However, at different developmental times certain genes may be only occasionally required to be active (i.e. expressed). This differential gene activity, whether they are on or off, can be regulated by the packaging of DNA. If it is highly packaged (condensed) then genes are usually silent. In an open conformation, the genes can be ‘on’ and direct the synthesis (via mRNA’s) of the different proteins that make up our cells and bodies. The nucleus has evolved a ‘barcode’ system to recognize the on or off state of genes and this barcode pattern will differ between cell types e.g. liver versus breast. This system is called ‘Epigenetics’ and involves putting chemical tags on the genetic material so the right genes are expressed at the right time and place. In chemical terms, the attachment of a methyl or a hyrdroxymethyl group to cytosine (a building unit of DNA) influences the use and storage of DNA. It is very obvious that this process is disrupted in many human diseases, for example the epigenetic ‘barcode’ of cancer cells is altered when compared to their normal counterparts and is associated with changes in ‘Gene expression’. This alteration is thought to contribute to the cancer development and one aspect of our work is to describe how this process may occur in breast cancer.

DNA Modification
As outlined above an intensive area of research is the ‘DNA methylation toolbox’. This consists of the enzymes (DNA methyltransferases and TET1-3 (non-heme iron-dependent dioxygenase’s)) that deposit the chemical tags (methyl and hydroxyl-methyl groups) on DNA. Cytosine is modified to 5-methyl cytosine (5mC); DNA methylation, which can be further modified to 5-hydroxymethylcytosine (hmC) by the TETs). Methylated DNA signals gene silencing and is considered a protective mechanism in normal cells that prevents inappropriate expression of genes and other DNA elements in our genome. For example genes that may be important for liver function would be out of place if they were expressed in the reproductive organs and vice versa.

So we study what, when and where genes are methylated and what happens if this process is perturbed either experimentally or in disease states. We want to know the mechanism by which the process works. We study how DNA methylation contributes to gene silencing, does it do it by-self or in combination with other factors? Is DNA methylation the only ‘barcode’ that contributes to gene silencing or does it work in combination with another ‘barcode’ found on the nuclear DNA storage proteins; the histones? We try to challenge what we think we know about the role of epigenetics in gene regulation to find out if our theories are correct or need to be rethought. We wish to apply this important basic knowledge to the analysis of breast tumours. An important project is the delineation of the abnormal patterns of methylation in different types of breast tumours. Do they match known pathologies and are they important in the direct alteration of gene expression patterns known to occur in cancer or are they a consequence?

Our research was stimulated by the recent discovery of 5-hydroxymethylcytosine in many tissue types including breast. hmC is formed by adding a methyl group to cytosine and subsequently an hydroxy group, in a reaction mediated by the TET family of enzymes. Its importance in epigenetics is that the hydroxymethyl group is suggested to further alter the biological properties of methylated DNA. However, it is clear that hmC is less abundant then 5mC, and the latter is still the most prominent modification in vertebrate DNA in many tissues. Initial analysis suggests that hmC is predominantly associated with the gene bodies of highly expressed genes. We would like to know if this relationship is preserved in cancer. Exciting new published data suggests that inhibition of TET function contributes to the generation of cancer specific methylation patterns. We wish to know the role of hmC in the development of breast cancer by studying where it is and if the patterns are altered in tumours compared to normal cells. If it is, it may provide new ways of classifying tumours and offer new therapeutic pathways.

All our projects rely heavily on collaborations and interactions with Breakthrough colleagues and other members of the Edinburgh scientific and clinical communities.