CONTROL OF OLIGODENDROCYTE DEVELOPMENT BY OLIG2 AND CHROMATIN REMODELLING COMPLEXES

Amazingly, the thousands of different cell types that make up our body - blood cells, muscle cells, nerve cells, for example - all develop from the same single cell, the fertilized egg. Hence, all cell types contain the same DNA, yet each contains a set of specialized proteins that are unique to that particular cell type, on top of a core set of "housekeeping" proteins found in all cells. The "cell-type-specific" proteins are what give a cell its particular identity - what defines it as a blood cell or nerve cell, for example. Examples are haemoglobin (present or "expressed" only in red blood cells), keratin (only in skin cells), insulin (only in pancreatic cells) and so on. How are these characteristic proteins expressed in only one or a few cell types despite the fact that all cells contain the same DNA - the same collection of genes? If we could understand the mechanisms that keep some genes shut down and others highly expressed, we might learn how to convert one cell type into another - for example, to cure diseases in which a particular type of cell is damaged or destroyed. Examples of such diseases are type-one diabetes, in which the pancreatic cells that make insulin are destroyed by the immune system, or motor neuron disease, in which spinal neurons that control muscle movement die for unknown reasons. If we could manufacture replacement cells from healthy cells in the body, or in a dish, this could be extremely helpful.

There are many proteins in cells whose function is purely to activate or repress other protein-coding genes, by binding specific DNA sequences next to those genes. Such DNA-binding proteins are called "transcription factors" because they control whether a given gene is "transcribed" into the instructions for assembling the corresponding protein. For example, the transcription factor OLIG2 is present uniquely in cells in the central nervous system called "oligodendrocytes". These cells make "myelin", spiral wraps of insulating membrane around "axons", the long thin extensions of nerve cells that carry electrical impulses from one part of the brain to another. This greatly increases the speed at which information travels around the brain. Without myelin, we literally would not be able to think quickly, or at all! Moreover, when myelin is damaged, as it is during the demyelinating disease multiple sclerosis, nervous function can be seriously compromised. We therefore want to understand how OLIG2 can activate myelin-forming genes uniquely in oligodendrocytes.

DNA in chromosomes ("chromatin") is normally tightly wound into a form that makes its encoded information inaccessible. Transcription factors like OLIG2 cannot by themselves unravel the DNA - they need to interact with many other proteins to form large "chromatin remodelling complexes" that can together release a gene from its tightly folded, "closed" state. These complexes are of several types (e.g. INO80 and ISWI complexes) whose individual functions are poorly understood. We recently found that OLIG2 is associated tightly with both of these complexes and others, raising the question of why different complexes are needed, and what do they individually do?

Our hypothesis is that the different chromatin remodelling complexes come into play at different stages of oligodendrocyte development to activate different sets of genes that allow progression along the path from early embryonic stem cell to fully mature, myelin-forming oligodendrocyte. Our project will test this idea by identifying the genes associated with the INO80 and ISWI complexes, determining whether and how OLIG2 directs the INO80 and ISWI complexes to those genes and whether different sets of genes are engaged as oligodendrocytes develop. Our experiments will help to illuminate general mechanisms of transcriptional regulation, applicable to all cell types, as well as helping us understand the detailed workings of the mammalian brain.

Oligodendrocytes (OLs) make myelin in the central nervous system (CNS). Myelin envelops axons, speeding conduction of action potentials as well as transferring energy substrates (e.g. lactate) into axons. OLs continue to be made during adult life in mice and are important for learning and memory. Therefore, OLs and myelin are essential for normal brain function and their loss during demyelinating diseases like multiple sclerosis causes severe neuropathology. Understanding OL development might suggest ways of enhancing cognitive performance and repairing demyelination.

The transcription factor OLIG2 is expressed at all stages of OL development and performs specific roles at each stage - possibly through a changing repertoire of partner proteins and gene targets. We identified OLIG2 binding proteins by mass spectrometry; many are components of known ATP-dependent chromatin remodelling complexes, including the INO80 and ISWI complexes. We aim to uncover the roles of INO80 and ISWI complexes in OLs, and the nature of their association with OLIG2.

Pilot data from purified mouse OL lineage cells show that INO80 and SMARCA5 (the central ATPases of the INO80 and ISWI complexes) are differentially regulated as OLs differentiate in culture, and that INO80 is required for OL precursors to proliferate. We will next examine OL lineage-specific knockouts of INO80 and SMARCA5 to ask whether, and at what stage, OL development arrests in vivo. We will identify genomic targets of INO80 and SMARCA5 by ChIPseq, determine whether OLIG2 is required for targeting, whether INO80/ ISWI complexes activate or repress specific genes and whether they act by targeting the histone variant H2A.Z in nucleosomes.

These experiments will start to reveal general principles of transcription regulation during development, as well as providing specific information about the OL lineage that could suggest targets for stimulating OL production in vivo.

Myelin is the multi-layered glial-sheath surrounding axons in the vertebrate nervous system. Myelin enables extremely rapid propagation of action potentials, allowing high-speed computation and a powerful brain. The cells that supply myelin in the central nervous system are oligodendrocytes (OLs). Intensive studies have been conducted to reveal how OL development is controlled, yet the interplay between transcription factors and epigenetic regulators during OL differentiation and myelination are not well understood. Our research will have immediate impact in the academic circle and may have social and economic impact in the long term.
Impact on academic community - This project will advance our understanding of OL differentiation and myelination. We will produce evidence on how the OL master transcription factor, OLIG2, in conjunction with the chromatin remodellers INO80 and SMARCA5, can regulate OL development in different ways and at different stages of OL lineage development. This will inspire other researchers to design experiments to reveal the interplay between transcription factors and epigenetic factors during development of other CNS (including OL) and non-CNS cell lineages. In addition, the postdoctoral research associate supported by this grant will benefit from excellent training in molecular biology and neurobiology. We anticipate that, if this grant is awarded, we will be successful in attracting a funded PhD student who will also benefit from tuition and training in the field of molecular genetics.
Impact on business/industry. The death and dysfunction of OLs can have devastating neurological consequences, e.g. leading to human diseases such as multiple sclerosis in adults and cerebral palsy in young children. Inducing endogenous OL precursors to differentiate and form new OLs to replace those that are dead or damaged is one potential treatment strategy for such diseases. Therefore, this study has the potential to uncover new drug targets and attract the interest of the pharmaceutical industry. In addition, OL development has been shown to continue throughout young adulthood in mice and we recently showed that newly-formed OLs are required for mice to learn new motor and cognitive skills. Therefore there is potential in the longer term to develop drugs to control and improve human learning and/or memory performance.
Impact on the general public: Understanding how our brain works and what happens in our brain if something goes wrong is inherently interesting to the general public, especially in an era of increasing incidence of and/or awareness of mental illness, and in an ageing society. We hope that our work can inspire more young people to take up neuroscience-related subjects at University and ultimately to move into neuroscience research in an academic or applied context.