Research in our lab is aimed at understanding fundamental mechanisms of epigenetic genome regulation. In particular we are interested in in the interplay between chromatin regulation and non-coding RNAs, especially small RNAs associated with the RNA interference pathway.
Our model of choice is fission yeast (Schizosaccharomyces pombe), a single celled eukaryote that is genetically tractable and has both RNAi and chromatin modification pathways similar to those in higher eukaryotes. In fission yeast, the RNAi pathway processes non-coding centromeric transcripts into small RNAs that direct assembly of repressive heterochromatin at corresponding genomic loci. This process is essential for proper centromere function, and has proven a convenient and tractable model for understanding RNA-mediated chromatin regulation more generally. Many factors involved in this process have been identified, and the principles learned have helped shape our understanding of related pathways in higher eukaryotes. However, key questions remain to be addressed, including how this regulatory pathway is itself regulated.
Recent evidence from several systems suggests that both RNAi and chromatin factors are targets for modification by ubiquitin, a small regulatory proteins can be covalently attached to other proteins to alter their activity or interactions. Indeed, one of the key protein complexes involved in RNAi-directed chromatin modification in fission yeast (a complex called CLRC, responsible for histone methylation) is also predicted to mediate ubiquitin attachment, although its protein targets are unknown. Until recently, efforts to study the regulatory roles of ubiquitin have been hampered by a lack of information on modification sites. However, we have recently conducted a large scale analysis that has generated a comprehensive dataset of ubiquitination sites in the fission yeast proteome. Included in this are a number of modifications to factors implicated in RNAi and/or chromatin modification.
The aim of this project is to exploit this new dataset to investigate if and how ubiquitin modification of factors involved in RNAi or chromatin modification contributes to their function in genome regulation. For candidate factors, we will mutate identified ubiquitination sites and assess whether this results in any impact on target chromatin domains. In cases where it does, we will then go on to investigate the molecular basis of the effect, for example testing for changes in protein abundance, subcellular localisation, or protein-protein interactions. We will also explore potential targets of the ubiquitination activity of the histone methyltransferase complex CLRC.
The project will provide training in a wide range of molecular and cell biology techniques including DNA and RNA analysis, chromatin IP, protein co-immunoprecipitation and microscopy. There will also be the opportunity to gain experience in quantitative proteomic techniques. Transferable skills including critical thinking, communication and presentation will be developed through regular participation in lab meetings, journal clubs and seminars.
If you wish to apply for this project, please go this link.