Investigating the role of PIP4K2B, nuclear phosphoinositides and TAF3 in transcription and genome organisation during myogenic differentiation

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Cells are constantly being instructed (programmed) to modulate their behaviour because of changes to environment of the body. In particular, continual repair of muscle tissue is essential to our daily lives but unfortunately is a process which deteriorates as we age. Deterioration in muscle function not only decreases our movement and function but also makes us more susceptible to metabolic type diseases such as diabetes and cardiovascular disease. Satellite cells are specialised cells that are present in small numbers within muscle tissue. When muscles are damaged either by exercise or during disease, the satellite cells become active and produce more muscle cells, a process called differentiation that helps to repair muscle tissue. The activation of satellite cells is complex and depends on signals generated within the environment of the damaged muscle. As organisms age the ability of satellite cells to respond to these signals and generate more muscle cells decreases and in part is responsible for age-induced deterioration in muscle function. Using a model of muscle cell generation we have found that by controlling an enzyme called PIP4K2B we can increase the ability of a cell to differentiate to produce muscle cells. PIP4K2B controls the levels of naturally occurring molecules called phosphoinositides that are present in the cells. It is well established knowledge that phosphoinositides are present in the plasma membrane of cells where they control many different cellular functions. However, over the years there has been growing evidence that phosphoinositides are also present in the nucleus, the cells' control centre, where their levels change in response to different environments. In fact in response to signals generated during muscle cell differentiation the levels of phosphoinositides in the nucleus go up. We now know that nuclear phosphoinositides interact with and change the function of special proteins in the nucleus that are involved in generating instructions that control the behaviour of the cells. One such protein is TAF3, which is part of different protein complexes that control the programming of cells. Interaction of TAF3 with phosphoinositides reprograms the cell to increase muscle cell differentiation. In this proposal we will use a novel state of the art technology called CRISPR CAS to change the DNA of cells so that we can investigate how PIP4K2B and nuclear phosphoinositides control TAF3 and its various complexes to increase muscle differentiation. We believe that specialised TAF3 complexes direct specific instructions to the cell in response to the interaction of TAF3 with phosphoinositides and we can investigate this using a technique called CHiP-Seq. We also think that nuclear phosphoinositides together with TAF3 act as a platform that helps to organise which instructions are given and how these are coordinated to increase muscle differentiation. We will investigate this using a novel technique called promoter capture HiC which will allow us to understand the three dimensional aspect of DNA in the nucleus that is used to generate these instructions. PIP4K2B is a very druggable protein as is the site of interaction between phosphoinositides and TAF3 and we hope that eventually we might be able to use our knowledge to develop drugs to harness this control process within a cell's nucleus and help satellite cells differentiate more effectively, thus aiding the process of muscle tissue repair.