Award details

The pathways to prion formation in the response to oxidative stress

Reference	BB/S005420/1
Principal Investigator / Supervisor	Professor Christopher Grant
Co-Investigators / Co-Supervisors
Institution	The University of Manchester
Department	School of Biological Sciences
Funding type	Research
Value (£)	414,175
Status	Completed
Type	Research Grant
Start date	03/06/2019
End date	02/03/2023
Duration	45 months

Abstract

We will use the yeast Saccharomyces cerevisiae as a model organism to elucidate the consequences of oxidative stress-induced protein aggregate formation. Our recent studies have shown that oxidative stress promotes the formation of both large amorphous protein aggregates as well as heritable prion aggregates and the aim of this project is to understand the processes which drive the formation of these different types of aggregates. Evolutionarily conserved protein deposit sites have been identified which effectively sequester misfolded and aggregated proteins away from their normal productive pathways, protecting against potential cytotoxic effects. However, it is unclear what role the localization of misfolded proteins to these sites plays in amyloid formation, protein heritability and protein degradation. We will use microscopy and biochemical approaches to define the contribution of intracellular protein deposits (IPOD, JUNQ, stress granules) to protein aggregate formation and fate. This will include using mutants that disrupt protein deposit formation to determine whether protein localization is required to facilitate protein-based inheritance. A key question to be addressed is whether such protein localization acts to facilitate autophagic clearance of aggregates versus heritable protein aggregate (prion) formation. These studies will make use of the well-characterized yeast [PSI+] prion to enable us to study the formation and heritability of protein aggregates. We hypothesize that certain protein aggregates may promote stress tolerance and we will use a biochemical approach to isolate the amyloid-like proteins which are formed in response to oxidative stress conditions. We will determine whether protein aggregates can act as a form of memory providing oxidant tolerance. These studies will have broad implications for understanding the role of oxidative protein damage as a trigger of amyloid formation in the eukaryotic cell.

Summary

Prions are protein-only infectious agents associated with a group of transmissible neurodegenerative diseases typified by human Creutzfeldt Jakob Disease (CJD). Although CJD is a rare disease, it shares many pathological features with other more common, non-infectious diseases of the brain such as Alzheimer's disease. In spite of its infectious nature, the majority of cases of human CJD appear spontaneously, without any evidence of infection by the associated infectious entity, the prion. However, the molecular basis of how prions form spontaneously into infectious structures is poorly understood at present. Prions are remarkable infectious agents because they consist only of a single protein that is a structurally altered form of a protein normally found in the brain. Yet we know very little about how prions form spontaneously to cause sporadic CJD or for that matter what will trigger their formation. To help us address these questions we are proposing to study prions that are found in Baker's yeast (Saccharomyces cerevisiae). Prions were first described in this fungus some 30 years ago and the subsequent studies on yeast prions have revealed many fascinating new aspects of prion biology. In our recent research we have discovered that certain dangerous forms of oxygen known as reactive oxygen species (or ROS) can trigger the spontaneous formation of prions in yeast, because cells lacking the defence system preventing such oxidative damage, form prions spontaneously at a remarkably high frequency. We are now interested in determining how oxidative damage to a normally soluble protein triggers its conversion into the prion form. Importantly, our novel genetic approaches will be relevant to understanding the underlying mechanisms of prion formation in man and animals. In the long term, findings from these studies may therefore reveal candidates for potential therapeutic intervention in the treatment/management of prion diseases.

Impact Summary

Sporadic Creutzfeldt - Jakob disease (CJD) accounts for over 80% of the verified cases of this fatal neurodegenerative disease in the UK, yet we know remarkably little about the underlying mechanism or triggers for disease development. The emergence of the disease correlates with the appearance of PrPSc, a novel conformational form of the cellular PrP protein. This entity, known as a prion, replicates through a cycle of seeded polymerisation and fragmentation, and it is assumed that certain genetic or environmental factors can trigger the conformational change in the absence of any pre-existing PrPSc 'seeds'. We have recently shown that protein oxidation underlies the switch from a soluble to a prion form of a protein, using the yeast prion [PSI+], which is the prion form of the Sup35 protein. We have proposed that the accumulation of oxidative damage in certain proteins such as Sup35 can lead to the formation of a heritable amyloid-like form of the protein in the absence of any underlying genetic change. The same may also apply to the much more prevalent common 'protein misfolding' diseases that lead to dementia and death and which are increasing in proportion to the increased life span of the human, namely Alzheimer's Disease, Huntington's Disease and Parkinson's Disease. This current proposal builds on our initial findings and will provide a detailed understanding of the mechanisms whereby oxidatively damaged proteins spontaneously form both heritable and non-heritable protein aggregates. The major impacts of this research nationally and internationally will therefore be on the bioscience research community with specific interests in the fields of protein quality control and the causes of protein misfolding that lead to disease in humans and/or farmed animals. In the longer term, findings from these studies may reveal candidates for potential therapeutic intervention in the treatment/management of prion diseases. This is a broader issue since a number of non-transmissible neurodegenerative diseases, including Parkinson's disease, Huntington's disease and Amylogenic Lateral Sclerosis, whilst not infectious, are also accompanied by accumulation of insoluble protein aggregates. It is therefore important to understand the protein homeostasis systems which protect against misfolding diseases if we are to develop potential therapeutic targets for the prevention and treatment of such pathologies. Results will be disseminated through research seminars, presentations at conferences and publications in scientific journals. Funding is requested to attend national and international research conferences to allow the researchers to publicize this research. Resources generated from this project are likely to include yeast strains and plasmids and will be made available to the scientific community upon request. Detailed protocols and primary data will be made freely available to academic collaborators. Manchester University has a good track record of encouraging public engagement. This includes regular open days to inform school children and the public about University research and tours of the research facilities at Manchester. This will allow the researchers to share their research findings with the wider public and to raise awareness of the importance of basic research. Manchester University maintains excellent links with the business sector which will allow us to exploit any potential for collaboration with industry.

Committee	Research Committee D (Molecules, cells and industrial biotechnology)
Research Topics	Animal Health, Microbiology
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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