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MRC Prion Unit
From fundamental research to prevention and cure
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Cellular mechanisms of prion propagation

Prions, the infectious agents that cause the lethal brain disease CJD in humans infect a range of different cells in the body. In contrast to other infectious diseases, like viral or bacterial diseases, prions are not recognised as rogue proteins by the immune system and the hosts remain symptom-free for a long period of time. During this phase prions rapidly multiply in lymphoid organs, like the spleen, lymph nodes or tonsils and finally reach the brain where they cause a fatal and progressive loss of neurons. We seek to better understand how prions are passed from cell to cell and how they are replicated by cells. Such knowledge will help to provide fundamental insights into the mechanisms of prion propagation and is important to develop better tests and treatments for these diseases in the future.

In early phases of disease prions accumulate in lymphoid organs long before they reach the brain and lymphatic replication is considered to be an important prerequisite for prions to reach the brain. Seminal work in the past two decades helped to identifying critical cell types involved in prion accumulation, but the molecular mode of prion spread remains unknown. We therefore aim to understand how prions are transmitted from scrapie-infected to recipient cells. The route of infection from the primary entry site of prions to neuronal cells is considered to involve multiple cell types, but a comprehensive study to compare the prion load of candidate cells has not been performed to date due to prohibitively large numbers of animals required. We established a procedure to sequentially isolate specific splenic cell populations and utilise an in-vitro infectivity assay, the Scrapie cell assay that was established at the Prion Unit to assess which cells are most infectious. We investigate the mechanism of prion spread in primary culture and co-culture models and plan to investigate the transmission of prions in vivo.

Peter Klohn Figure 1

Figure 1. To determine which cell types of the spleen accumulate prions after infection we isolate cells of the spleen that play a distinct role in the immune system, like B cells, T cells, dendritic cells (DC), macrophages (MΦ) and follicular dendritic cells (FDC) and determine infectious titres with a cell-based assay, the Scrapie Cell Assay.

 

Identifying modifier genes associated with prion replication

Most studies of prions have required use of laboratory animals but more recently it has been possible to isolate types of cells that can be grown indefinitely in the laboratory and used to propagate prions. However, it turns out to be very difficult to find cells that will propagate prions efficiently and the ones that do typically will only propagate just one type, or strain, of prions. By studying different lines of cells that are highly susceptible or poorly susceptible to defined prion strains, we aim to identify the genes that are required to render cells susceptible or resistant.

Peter Klohn Figure 2

Figure 2. To explore which genes are required to replicate prions we investigate expression differences between cells that have a high propensity to replicate prions (susceptible) and those that are resistant to prion replication. Messenger RNA or mRNA, a copy of the information carried by a gene on the DNA is isolated, labelled with fluorescent dyes, combined and hybridised onto microarray chips. In the example above gene 1B is expressed primarily in susceptible cells, whereas gene 1C is expressed in resistant cells. Gene 1A not differentially expressed.


Reviews:

Exosome release from infected dendritic cells: A clue for a fast spread of prions in the periphery
Kloehn PC, Castro-Seoane R, Collinge J.  J Infect 2013; 67: 359-368.


Peer reviewed articles:
2014

Identification of a gene regulatory network associated with prion replication
Marbiah M, Harvey A, West BT, Louzolo A, Banerjee P, Alden J, Grigoriadis A, Hummerich H, Kan HM, Cai Y, Bloom GS, Jat P, Collinge J, Klöhn PC. EMBO J 2014

In-vitro screen of prion disease susceptibility genes using the scrapie cell assay
Brown CA, Schmidt C, Poulter M, Hummerich H, Klöhn PC, Jat P, Mead S, Collinge J, Lloyd SE.
Hum Mol Genet 2014

2012

Plasmacytoid dendritic cells sequester high prion titres at early stages of prion infection
Castro-Seoane R, Hummerich H, Sweeting T, Tattum HM, Linehan JM, Fernandez de Marco M, Brandner S, Collinge J, Klöhn PC.  PLoS Pathog 2012; 8: e1002538..

Prion protein antibodies do not trigger mouse hippocampal neuron apoptosis.
Klöhn PC, Farmer M, Linehan JM, O'Malley C, Fernandez de Marco M, Taylor W, Farrow M, Khalili-Shirazi A, Brandner S, Collinge J. Science 2012; 335: 52.

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