Fungal contamination leading to reduced harvest and post-harvest food spoilage accounts for the loss of 10- 20% of food production worldwide. A major aspect is the loss of food due to contamination with mycotoxins such as aflatoxins, which are potent carcinogens and can cause severe liver damage. The rising threat of fungicide resistance further urges the demand for research into essential cellular pathways to find new targets for drugs against plant and animal pathogenic fungi.
Here, we have identified the multi-subunit endoribonuclease RNase MRP as a promising new target for anti-fungal drug discovery. RNase MRP is conserved in all eukaryotes and often mutated in human disease, such as cartilage-hair hypoplasia (CHH) and other skeletal dysplasias, but its catalytic mechanism is not fully understood. Notably, fungal RNase MRP contains two essential protein components that are not present in multicellular organisms and could therefore be targeted with new anti-fungal drugs, but the roles of these fungal-specific components are not clear. We also do not know if the human complex contains proteins not present in fungi.
While RNase MRP plays a role in cell cycle regulation, it is mainly known as a key player in the production of ribosomes, the large RNA-protein machineries that synthesise all cellular proteins. Ribosome production is absolutely essential for all cells and directly determines their proliferative rate. Defects in ribosome biogenesis are also linked to ~20 inherited human diseases (called ribosomopathies). Key events in ribosomal RNA (rRNA) maturation, a pathway best-characterised in budding yeast (Saccharomyces cerevisiae), are cleavages, for example catalysed by RNase MRP, that release the mature rRNAs from a precursor transcript (pre-rRNA). Orthologues of most yeast ribosome biogenesis factors are present or predicted in all eukaryotes including fungi, but functional studies in models for pathogenic fungi (e.g. Aspergillus nidulans) are in their infancy.
The aim of this DTP2 partnership PhD project with the main base at Newcastle University and an external project at the University of Liverpool will combine multidisciplinary approaches to study RNase MRP and its role in pre-ribosomal RNA cleavage in three biological systems, aiming to identify chemical compounds that specifically inhibit fungal, but not human RNase MRP.
Project outline and training
Understanding the importance of RNase MRP and the role of its fungal- or human-specific components in different organisms, in particular in a model for pathogenic fungi, is key to validate RNase MRP as a potential target for new anti-fungal drugs. Targeted in vivo modification of pre-rRNA cleavage sites to specifically inhibit pre-rRNA processing in budding yeast, Aspergillus nidulans and human cells will therefore be performed to reveal the importance of individual cleavage events in different organisms and help to elucidate the catalytic mechanism of key enzymes such as RNase MRP. Genetic approaches will also be applied to study the role of RNase MRP and its fungal-specific components in yeast and Aspergillus nidulans.
Affinity-purification of RNase MRP complexes from fungi and human cells will further be employed to characterise and compare their 3D-structures, catalytic activities and, importantly, sensitivity to chemical compounds. This will involve cryo-EM studies of purified particles (in collaboration with a team based in Toulouse, France), as well as in vitro RNA cleavage assays to test the catalytic activity of purified enzymes. To identify drugs specifically affecting RNase MRP activity, these assays will be combined with the LifeArc Drug Library available from the High Throughput Screening Facility at Newcastle University, focusing on compounds that are toxic to budding yeast, but not human cells. In parallel, the student will establish in vivo systems (using Saccharomyces cerevisiae as a model system as well as human cell culture) to test selected compounds. The effect of the identified fungal-specific compounds(s) will then be analysed in Aspergillus nidulans.
The successful PhD candidate will be trained in a wide range of genetic, molecular biology and biochemical techniques. They will also benefit from the PhD development programme at the Newcastle University Graduate School, which offers a variety of core and transferable skills courses and a flourishing postgraduate culture. Career progression will be monitored yearly by an academic progression panel and a designated PhD mentor. The student will also be able to present their data at both national and international meetings.
For further information see the website: https://www.ncl.ac.uk/camb
Please complete the online application form and attach a full CV and covering letter. Informal enquiries may be made to [email protected]
Wells, G.R., Weichmann, F., Sloan, K.E., Colvin, D., Watkins, N.J. and Schneider, C. (2017) The ribosome biogenesis factor yUtp23/hUTP23 coordinates key interactions in the yeast and human pre-40S particle and hUTP23 contains an essential PIN domain. Nucleic Acids Res. 45(8):4796-4809. doi: 10.1093/nar/gkw1344.