Cellular Functions of PrP

Lead Research Organisation: MRC Prion Unit

Abstract

Prion diseases are fatal progressive neurodegenerative disorders that, like Alzheimer’s and Huntington’s disease, involve accumulation of a misfolded protein. This protein, the prion protein, is normally expressed within most cells of the body but the highest expression is within the brain and in the immune system. It is predominantly found on the surface of cells attached to the plasma membrane via a carbohydrate containing fatty acid linker.
The prion protein has been proposed to have many cellular functions but is has been found that mice that are null for this protein are viable, do not have any overt symptoms but are completely resistant to prion disease. Even though the cellular prion protein is a prerequisite for prion disease, it is not known which parts of this protein are required for prion propagation.

We aim to investigate which regions of the prion protein are required for propagating prions. We also aim to develop cells which can be used to propagate human prions since there are currently no cell lines which can propagate human prions. We also aim set up a primary cell assay to demonstrate that prions can kill neurones and use it to isolate and characterise the toxic species in prion disease.

Technical Summary

According to the widely accepted protein-only hypothesis, prions comprise aggregated forms of misfolded host-encoded cellular prion protein (PrPC) generally referred to as PrPSc. PrPC is a glycosyl phosphatidylinositol (GPI)-anchored cell-surface glycoprotein, expressed in most cell types and tissues, but at the highest levels in the CNS and immune system. PrPC is highly conserved in mammals but a precise cellular function remains unclear although multiple functions have been proposed. However mice, in which the PrP gene has been knocked out, have no overt phenotype and a normal lifespan, but do not propagate prions and are completely resistant to prion disease, indicating that PrPC expression is absolutely required for prion propagation and prion-induced neurotoxicity.

Like most GPI-anchored proteins, PrPC is predominantly found at the cell surface in lipid rafts which has led to the suggestion that it may be involved in signal transduction by acting as a cell-surface receptor. Since PrPC lacks a transmembrane domain for transmitting signals from the cell surface to the inside of the cell, much work has been done to identify binding partners that could transmit the signal from the cell surface to the cell interior. A number of putative binding partners have been identified but the biological significance of many of them is unclear since some are only present within the cytoplasm, whereas others are nuclear. Recent evidence has suggested that PrPC acts as a cell surface receptor for A?-oligomers, the molecular species that mediate toxicity in Alzheimer’s disease. PrPC is also widely expressed in the immune system where it has been suggested to play a role in homeostasis as well as in the immune synapse and signalling.

We have undertaken a reverse genetic approach, utilising alanine mutagenesis, to investigate the regions of PrPC that are required for efficient prion propagation in PK1 cells, a derivative of N2a neuroblastoma cells. This has identified three regions within the unstructured N-terminus as critical for prion propagation: residues 23-25 (KKR, Charge Cluster 1), glutamine 41 and residues 90-111 (Charge Cluster 2). Our aim is now to identify the proteins that interact with these regions.

We also aim to develop cells that can efficiently propagate human sCJD and vCJD prions to set up an automated bioassay for human prions that can be used as a therapeutic biomarker. We will also develop a primary neuronal cell assay for prion neurotoxicity that will be applied to isolate and characterise the putative toxic PrP species.

Publications

10 25 50
 
Description CONACyT Mexican PhD Scholarship
Amount £75,000 (GBP)
Funding ID 329734 
Organisation Government of Mexico 
Department Prospera
Sector Public
Country Mexico
Start 02/2015 
End 01/2018
 
Description Commonwealth Scholarship
Amount £70,334 (GBP)
Funding ID CSC Reference Number: INCS-2014-212 
Organisation Government of the UK 
Department Commonwealth Scholarship Commission
Sector Public
Country United Kingdom
Start 10/2015 
End 09/2018
 
Description Policy Research Programme
Amount £493,145 (GBP)
Funding ID PR-R17-0916-23004 
Organisation Department of Health (DH) 
Sector Public
Country United Kingdom
Start 05/2018 
End 03/2022
 
Description Big Questions in Neuroscience, Cell Symposia, Society for Neuroscience Satellite meeting 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Presented poster on the development of a primary cell assay for prion induced neurotoxicity.
Year(s) Of Engagement Activity 2017
 
Description Conference: Glia in Health and Disease, Cold Spring Harbor Meeting July 16-19th 2020 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact International conference on Glia and Health and Disease
Year(s) Of Engagement Activity 2020
 
Description Jacques Monod Conference: Protein misfolding in disease - Toxic aggregation-prone proteins in aging and age-related diseases: from structure to pathology and spreading 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Presented poster on our work on the development of a cell based assay for prion induced neurotoxicity.
Year(s) Of Engagement Activity 2016
 
Description National Prion Clinic Open Day 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Patients, carers and/or patient groups
Results and Impact The National Prion Clinic (NPC) is the national referral centre for prion disease and is part of the University College London Hospitals NHS Foundation Trust. It is funded by the NHS to provide diagnosis and care for patients with, or suspected to have, any form of human prion disease (inherited, iatrogenic, sporadic and variant CJD). The NPC is integrally linked with the Prion Unit.

The Open Day is an annual public engagement event in which its stakeholders - patients, patients' relatives, health and allied health professionals are invited to attend. The Open Day seeks to inform and update stakeholders on the Unit's prion research; focus group discussions are also held to interact with the stakeholders and gauge their views about our research. Laboratory tours are also offered.
Year(s) Of Engagement Activity 2017
 
Description Poetry collaboration between scientists and artists with experimental words at Springer Nature 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact The event was a collaboration between scientists and artists to engage the public by explaining the science of prions by poetry.
Year(s) Of Engagement Activity 2017
 
Description Prion Conference 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Annual International Prion conference where researchers from all the world come to present their latest research.
Year(s) Of Engagement Activity 2014,2015,2016
 
Description Queen Square Symposium 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Postgraduate students
Results and Impact Poster presentation on the "Development of an in vitro assay of prion- induced toxicity".
Year(s) Of Engagement Activity 2017
 
Description UCL - Research Images as Art / Art Images as Research 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Undergraduate students
Results and Impact Involved in the 2018 UCL Research Images as Art / Art Images as Research, in which artists or scientists presented images that bridge the science with arts. I presented a digital image of neurons growing in a well of a culture plate. I explained how they network/communicate with one another and suggested analogies with the communication/network at a larger world-scale. Images were exhibited for around 1 month where any interested members of public or university students could view.
Year(s) Of Engagement Activity 2018
URL http://www.grad.ucl.ac.uk/comp/2018-2019/research-images-competition/