About the Project
Peroxisomal Biogenesis Disorders (PBDs) such as Zellweger syndrome (1) refers to a group of diseases caused by genetic mutations in one of the membrane proteins (PEXs) responsible for biogenesis of the peroxisomes. Peroxisomes are organelles that generate reactive oxygen species (ROS) that trigger an essential process called pexophagy where damaged peroxisomes are cleared by autophagy (i.e., a cellular ‘self-eating’ process via lysosomal degradation). Emerging evidence has linked upregulated pexophagy to the pathogensis of PBDs (2,3). However, how pexophagy is regulated remains largely unknown. It is known that pexophagy in mammals is driven by hypoxia-inducible factor (HIF2α) (4) that undergoes a protein post-translational modification called SUMOylation which attaches the Small Ubiquitin MOdifier protein called SUMO (5). It is also known that increased ROS increases the stability of an enzyme called SENP3 (6) that removes SUMO from SUMOylated proteins in a process called deSUMOylation. Intriguingly our preliminary results suggest that SENP3 contributes to ROS-induced pexophagy. These findings have led us to hypothesize that SENP3-mediated deSUMOylation of HIF2α is required for pexophagy induction. This exciting project will explore the potential role for HIF2α deSUMOylation by SENP3 in pexophagy regulation related to the pathogenesis of PBDs, and the findings from this project would better our understanding of molecular mechanisms underlying PBDs and identify new target(s) for future therapeutic intervention for this type of genetic diseases.
The proposed work will involve a combination of techniques, including molecular biology (e.g., cloning, tagging, mutagenesis and CRISPR-Cas9 technology), protein chemistry (e.g., glutathione S-transferase/Histidine pulldowns, western blotting for LC3 conversion and p62 degradation, co-immunoprecipitations, protein purification and assays for SUMOylation/deSUMOyation), cell biology (e.g., cultures of clonal cell lines, neuronal cell lines and primary rat cortical neurons; DNA & sh/siRNA transfections, pexophagy induction by cell stressors such as hypoxia, hydrogen peroxide and clofibrate), imaging detection of pexophagy (e.g., immunocytochemistry, fluorescence, confocal and electron microscopic analysis of morphological changes in peroxisomes as well as “pexophagic flux” in cells expressing a novel biosensor pexo-pHfluorin), and peroxisome function analysis (e.g., ROS production and cell viability examinations upon and following stressed conditions).
Science Graduate School
As a PhD student in one of the science departments at the University of Sheffield, you’ll be part of the Science Graduate School. You’ll get access to training opportunities designed to support your career development by helping you gain professional skills that are essential in all areas of science. You’ll be able to learn how to recognise good research and research behaviour, improve your communication abilities and experience the breadth of technologies that are used in academia, industry and many related careers. Visit www.sheffield.ac.uk/sgs to learn more.
First class or upper second 2(i) in a relevant subject. To formally apply for a PhD, you must complete the University's application form using the following link: http://www.sheffield.ac.uk/postgraduate/research/apply/applying
All applicants should ensure that both references are uploaded onto their application as a decision will be unable to be made without this information.
1. O. Y. Al-Dirbashi et al., Zellweger syndrome caused by PEX13 deficiency: report of two novel mutations. American journal of medical genetics. Part A 149a, 1219-1223 (2009).
2. C. M. Cipolla, I. J. Lodhi, Peroxisomal Dysfunction in Age-Related Diseases. Trends in endocrinology and metabolism: TEM 28, 297-308 (2017).
3. T. Y. Nazarko, Pexophagy is responsible for 65% of cases of peroxisome biogenesis disorders. Autophagy 13, 991-994 (2017).
4. K. M. Walter et al., Hif-2alpha promotes degradation of mammalian peroxisomes by selective autophagy. Cell metabolism 20, 882-897 (2014).
5. M. van Hagen, R. M. Overmeer, S. S. Abolvardi, A. C. Vertegaal, RNF4 and VHL regulate the proteasomal degradation of SUMO-conjugated Hypoxia-Inducible Factor-2alpha. Nucleic acids research 38, 1922-1931 (2010).
6. C. Huang et al., SENP3 is responsible for HIF-1 transactivation under mild oxidative stress via p300 de-SUMOylation. The EMBO journal 28, 2748-2762 (2009).