Noel Buckley

Our interests are focused upon the transcriptional programs that operate to regulate neuronal gene expression. During embryonic life most of our 100 billion nerve cells are born. They are derived from neural stem cells, multipotent cells that have the potential to turn into any type of nerve cell. There are hundreds if not thousands of types of nerve cell - each type differing in its shape, connectivity and chemistry. This diversity of form, or phenotype, is essential to correct formation and functioning of the nervous system. Establishment and maintenance of phenotype requires that particular sets of genes are activated whilst others are shut down. Gene activity is regulated by transcription factors - proteins that bind to specific sets of genes and act as on/off or dimmer switches and microRNAs - short RNA molecules that switch off gene expression by destabilizing mRNA or blocking its translation. To date, our knowledge of how these 'transcriptional programmes' are established in nerve cells is almost non-existent. In no case do we know the complete set of targets of any singular transcription factor. One 'switch' we use for our studies is REST, a multifunction transcription factor that represses or silences many genes in both neural, and non-neural cells and is required for normal embryogenesis and development of the heart, as well as for neural stem cell differentiation. Furthermore, the regulation of REST and its target genes play important roles in several neuropathological conditions, including the response to ischaemic or epileptic insults, Down's syndrome, Huntington's disease and in some medulloblastomas. We have used bioinformatic approaches to identify all potential REST binding sites and target genes across multiple vertebrate genomes. Using chromatin immunoprecipiatation (ChIP) combined with DNA microarrays we can identify which target genes are operated on in which cell type and furthermore, we can map the epigenetic signature around theses sites. Combined with manipulating and measuring gene expression we are building up a profile of the cofactor platforms and chromatin modifications associated with each site. The application of this combined biochemical and bioinformatics approach allows several fundamental questions to be addressed: How many REST binding sites are there across the entire genome? Which genes are operated on in which cell types? Does this change with developmental stage? How does co-factor recruitment vary across different loci? Is this the cause or effect of distinct 'epigenetic signatures'? Currently, we are applying these approaches to neural stem cells to see how the epigenetic signature changes during neuronal and glial differentiation. Is the multipotentiality of a neural stem cells reflected in its chromatin structure? How does this change as commitment and differentiation proceed? Another application of this approach is the study of transcriptional dysfunction during neurological illness. For these studies we focus on the interaction between REST and huntingtin, an interaction that is disrupted in the presence of the mutant huntingtin allele leading to decreased expression of specific genes such as BDNF, a vital survival factor for striatal neurons. This gives a genome-wide perspective on transcriptional dysfunction in Huntington's disease and can potentially identify biomarkers or therapeutic targets.

Transcriptional and epigenetic analysis of neural stem cells.