The role of Nuclear phosphoinositides in epigenetic signalling

If genetics is represented by a book containing the sequence of DNA encoding humans then epigenetics can be considered as the notes written in the margins by a previous reader. They do not alter the code itself but remind and instruct the next reader how they might interpret the written code. Epigenetics provides the means for environmental signals, such as how much food has been eaten or how warm it is, to be written on top of the genetic code to help instruct how cells and organisms might respond. Like the DNA code the notes can also be inherited. In essence epigenetics provides a rational model for the nature versus nurture hypothesis.

DNA is complexed with proteins to form a structure called chromatin that enables the large amount of DNA to be packed into the very small volume of the cell. Problematically this packing prevents the DNA from being used easily. The epigenetic notes which are deposited on to the components of chromatin help to pack and un-pack the chromatin. The notes are deposited, removed and interpreted by a series of enzyme complexes. Exactly how these complexes are regulated in response to environmental signals is not very well understood.

We have found that one of these enzyme complexes is regulated by an enzyme called PIP5K1A. In response to environmental signals PIP5K1A produces a small molecule called PIP2. Our studies suggest that PIP2 sticks to the enzyme complex and controls how well it works. This leads to changes in the epigenetic notes that are deposited and eventually to how the cell behaves. In this way we think that PIP2 acts a messenger informing and controlling the enzyme complex in response to changes in the environment.

In this proposal we intend to understand how PIP2 sticks to and changes how well the enzyme complex works. The enzyme complex is made up of many different protein components and we have found that one of these proteins can stick to PIP2. By using different molecular technologies we want to find out exactly how PIP2 sticks to this protein and if this alters how well the enzyme complex deposits the epigenetic notes. Understanding the molecular details of this process is important as it increases our basic understanding of how cells respond to their environment. Furthermore the enzyme complexes that deposit epigenetic notes often do not work properly which can change how organisms age. It can also lead to the development of different human diseases such as cancer and diabetes. Understanding how the enzyme complexes are controlled will help us to design medicines that alter how well these complexes work which could be useful in modulating how well we age and how we combat diseases.

DNA and histones are packaged into chromatin. Epigenetic signaling enzymes that modify chromatin alter the packaging and consequently transcriptional output. For example histoneH3 lysine 4 (H3K4) tri-methylation is associated with enhanced transcription. H3K4 methylation is controlled by six different methylation complexes (MLL-SET) and impacts on differentiation, development, aging and transgenerational epigenetic memory. While much is known about H3K4 signalling and its relationship to transcription much less is known about how MLL-SET complexes are regulated.

Nuclear Phosphoinositides (PI), such as Phosphatidylinositol(4,5)bisphosphate (PIP2), are lipid messengers that change during cell cycle progression, differentiation and adaptive stress responses. Nuclear PI operate by interacting with and regulating proteins often involved in chromatin regulation.

We have identified a direct functional interaction between PIP5K1A, which synthesises PIP2, and an essential core component of all MLL-SET complexes. We suggest that nuclear PIP2 regulates H3K4 methylation by interacting with this core component. We will use an array of molecular and structural technologies to identify: A. components that constitute the nuclear PIP2/H3K4 methylation pathway; B. how PIP5K1A and PIP2 interact with the MLL-SET core component and C. how these interactions might regulate MLL-SET activity in vitro and in cells. Next generation sequencing will investigate how nuclear PIP2 influences global transcriptional output and H3K4 methylation. Finally, we will develop novel technologies to investigate how different sub-nuclear pools of PIP2 impact on H3K4 methylation and transcription.

Epigenetics will provide novel targets to impact on healthy ageing and diseases. Defining how small molecule such as PIP2 might allosterically regulate MLL-SET activity will likely provide highly selective, less toxic approaches to intervene in H3K4 methylation signalling.

People. This project has the potential to impact directly on the general public through impacting on health. PI and epigenetic signalling are implicated in cancer, muscular degeneration and in mental health and inflammatory diseases. Our studies bring these two pathways together and in doing so reveal novel target sites for the potential development of therapies that could be used to combat these diseases. Furthermore recent studies in model organisms show that PI and epigenetic signalling impact on the ageing process and on health span, potentially placing our studies of value in this area. The western world has an increasing aged population above 65 and both medicinal and non-medicinal interventions will be essential to maximise the health span of the population. Therefore the outputs from this research and their further commercial developments have potentially enormous impact on public health, although the realistic timescales to such therapeutic use are in the order of tens of years.
The increased knowledge gained from this proposal and its impact on human health and ageing are topics frequently highlighted in the news and are of direct interest to the public. Engaging the public in discussions of the wider issues encompassing our research increases knowledge and fosters an information based society which indirectly impacts on the ability of research councils and universities to lobby government on research funding policies.

Industrial beneficiaries. The research outputs outlined in this proposal could impact on industrial beneficiaries such as small and larger scale pharmaceutical companies. Development of therapies aimed at components of PI and epigenetic signalling pathways are already being developed illustrating their known potential in human health. Our studies will define exactly how a small regulatory molecule, PIP2, interacts with and controls an epigenetic signalling complex providing novel target sites for commercially exploitation. Industrial partnerships could be instigated within two years. These partnerships will likely be aimed at developing proof of principle cellular and animal models. Within the time frame of the grant we would hope to strengthen and generate productive industrial partnership interactions.

Economy. If target molecules can be realised then the potential to the overall economy is large. Revenue from the technology and its exploitation together with increased societal health will impact on the economy of the nation. The second potential for impact into the economy is through training and teaching. The research participants within this project will be trained in sophisticated biological and non-biological areas thereby enabling their contribution to both the academic and professional science base as well as non-academic environments. This research will also provide research placements and lecture material for undergraduate students, thereby contributing to the development of an educated workforce.