Background: R-loops (DNA/RNA hybrids) are conserved across all domains of life. They serve important functions for regulating gene expression and may cover ~5% of the human genome. Importantly, their aberrant accumulation triggers genome instability causing age associated disease such as dementia, thus tight control of R-loop homeostasis is essential. Indeed, genes involved in R-loop processing are linked neurodegenerative diseases.
R-loops often form during transcription when the nascent RNA molecule hybridises back to the transcribed DNA strand, thereby displacing the complementary DNA strand. Defects in DNA repair, for instance the failure to remove stalled Topoisomerase-DNA complexes, can increase R-loop formation. In order to understand the link between R-loops and neurodegeneration better, and as a starting point to screen for potential therapeutic approaches we are currently creating zebrafish mutant models with defects in R-loop homeostasis.
The prospective PhD student will analyse previously created rnaseh2a mutants, which are predicted to be defective in R-loop processing and, in addition, another newly identified DNA repair factor that is also predicted to increase levels of R-loops and affect cerebellar functions. Both mutations lead to neurodegeneration in embryos fully deficient for these proteins. The project will use genetics approaches to ascertain if the neurodegenerative phenotype is due to their role in R-loop processing. This will be achieved by rescue experiments using specific mutants generated in the lab, first by RNA injection, and with selected constructs also by transgenesis, or CRISPR/Cas9 editing approaches. Where required, further models can now efficiently be generated using CRISPR/CRISPANT approaches.
The generated disease models will be used to determine the precise in vivo effect on neuronal survival, R-loop homeostasis, DNA damage and inflammatory markers. Importantly, these models will serve as platforms to explore potential “treatments” to suppress the observed phenotypes, which has a much broader impact in neuroscience, given the role of R-loops in multiple neurodegenerative disorders. Therefore, the student will also do pilot chemical screens to search for chemical suppressors of the neurodegenerative phenotypes. A candidate modulator that we have identified (HIF) will be included in this.
The student will employ a wide variety of genetic techniques in zebrafish, CRISPR/Cas9, immunohistochemistry, in situ hybridisation, microinjection, use of fluorescent proteins, confocal microscopy, qRT-PCR, in addition NGS approaches may be employed to dissect mutant phenotypes.
Applications are open to students from both the UK and overseas, though we note that due to funding constraints the availability of positions for students with overseas fee status will be more limited. We anticipate competition for these studentships to be very intense. We would expect applicants to have an excellent undergraduate degree in a relevant discipline. We would also expect applicants to have completed or be undertaking a relevant master’s degree to a similar very high standard (or have equivalent research experience).
--
Please complete a University Postgraduate Research Application form available here: https://www.sheffield.ac.uk/postgradapplication/
Please clearly state the prospective main supervisor in the respective box and select ‘Neuroscience’ as the department.
After the application closing date, we will shortlist applicants for an online interview. We expect to carry out interviews (each lasting approximately 30 minutes) on Tuesday 27th April (am, GMT) and Tuesday 4th May (pm, GMT). If you are shortlisted for interview, we will aim to inform you of this no later than the end of Friday 23rd April. If you are unable to attend at the specified times, please let us know if we confirm that we would like to interview you.
Continue with Facebook
