Analysis of mesenchymal stromal/stem cell sub-populations
Human mesenchymal stromal/stem cells (MSCs) are found in adult tissues such as bone marrow and are able to differentiate into osteogenic, chondrogenic and adipogenic tissues. There is intense interest in determining how MSCs may be used in future cell-based therapies, including gene therapy, immunotherapy and tissue engineering, and as in vitro models for fundamental research and drug discovery. However, little is known about MSC identity and research is often performed on heterogeneous mixtures of different MSC sub-populations. Using a process of telomerase-based immortalisation and cell cloning, we have generated several different MSC lines that represent different “types” of MSCs. For example, some MSC lines demonstrate tri-lineage, whereas other are bi-potent, uni-potent or nulli-potent. This project will examine the differentiation characteristics of these MSC lines to determine how they reflect MSC sub-populations in vivo.
Role of redox-signalling and oxidative stress in the regulation of neuronal transcription factors (2016-17)
Synaptic activity-induced changes in neuronal gene expression programs direct neuronal differentiation during development and alter connectivity of adult neurons in response to emotional and sensory stimuli. The Class IIa HDACs, HDAC4 and HDAC5 are transcriptional co-repressors that regulate the activity of the myocyte enhancer factor-2 (MEF2) family of transcription factors. Under basal conditions Class IIa HDACs reside in the nucleus and repress MEF2 transcriptional activity. Derepression of MEF2 factors involves synaptic activity-induced nuclear export of Class IIa HDACs through phosphorylation of two conserved serine residues in their N-termini. Recently, reactive oxygen species (ROS) have been implicated in mediating nuclear export of Class IIa HDACs in muscle through the oxidation of conserved cysteines in HDAC4 and HDAC5. Moreover, HDAC5 displays phosphorylation-independent cyclical changes in subcellular localization (Fogg et al, 2014). This PhD project will examine whether ROS signalling influences HDAC4/5 subcellular localization in primary hippocampal neurons in response to synaptic activity and in response to oxidative stress. It will investigate whether there is an interaction between HDAC4/5 cysteine oxidation and phosphorylation. The project will employ a range of cellular, molecular and biochemical techniques to assess HDAC4/5 localization, phosphorylation and cysteine oxidation in neurons.
FoggPCM, O'NeillJS, DobrzyckiT, CalvertS, LordE, LordRL, ElliottCJH, SweeneyST, HastingsMHand Chawla S (2014). Class IIa histone deacetylases are conserved regulators of circadian function. Journal of Biological Chemistry 289, 34341-34348.
Identifying enhancers and suppressors of Frontotemporal Dementia in a Drosophila model (2015-16)
Frontotemporal Dementia (FTD) is a major cause of early onset dementia. Characterised by atrophy of the frontal and temporal lobes with loss in language and social function, FTD incidence has a strong dominant genetic component. A number of loci have been mapped. Among these is CHMP2B encoding a subunit of ESCRTIII, a complex required for endosomal function. We have used mutant CHMP2B to perform genetic screens in Drosophila for enhancers and suppressors of an FTD related phenotype in the fly eye. We have preliminarily mapped 29 loci, three of which we have identified and characterised in detail: a regulator of the Toll innate immune pathway (Ahmad et al., 2009, PNAS), a component of phagosome maturation (Lu et al., 2013, Mol Cell) and a regulator of the endosomal recycling pathway (paper in preparation). This project will focus on fine mapping and identifying other loci from the screen. Once identified, fly genetics, cell biology and physiology will be used to define the cellular function of such proteins critical to FTD onset and progression. This project is a collaboration with Professor Fen-Biao Gao of the Department of Neurology, University of Massachusetts, Worcester. Exchanges between labs may play an important part of the project.
Novel Regulators of Seedling Growth (2015-16)
RNA polymerase (pol) III synthesizes noncoding RNAs that are essential for growth, e.g. tRNA and 5S rRNA. Elevated tRNA expression has been shown to stimulate cell proliferation and organismal growth in fruitflies. In yeast and animal cells, transcription by pol III is tightly linked to growth conditions, sometimes through epigenetic changes: it is suppressed in quiescent/dormant cells by transcriptional inhibitors; growth stimuli inactivate these inhibitors and induce direct activators of pol III transcription. In Arabidopsis, transcription, methylation and compaction of 5S rRNA genes is regulated during seed development and germination. It is not yet known if tRNA expression is controlled in a similar way to 5S rRNA in plants, although precedents from other organisms predict that this is the case. The project will have four main objectives: 1. Determine how tRNA expression is regulated when Arabidopsis seeds exit dormancy; 2. Test the hypothesis that loss of dormancy triggers epigenetic changes in tRNA genes; 3. Test if tRNA genes in Arabidopsis are regulated by homologues of transcription factors that regulate pol III transcription in animal cells (e.g. MYC, MAF1, RB); 4. Test if manipulating tRNA expression can influence seedling growth.
Co directors: Professor Bob White and Professor Ian Graham