The Role of Cofactors in Prion Propagation and Infectivity

article has not abstract

Published in the journal: . PLoS Pathog 8(4): e32767. doi:10.1371/journal.ppat.1002589
Category: Pearls


article has not abstract


The term “prion" was originally coined by Prusiner to explain the unusual infectious agent in transmissible spongiform encephalopathies (TSEs, also known as prion disease) [1]. Now the term has expanded to include a growing list of fungal proteins that stably maintain an atypical self-propagating conformation and epigenetically modify a variety of cellular processes [2]. Although fungal prions and the TSE agent share the capability of maintaining an atypical self-propagating conformation, fungal prions distinctly differ from the TSE agent in several aspects [3]. Thus far, the TSE agent is the only prion that behaves as a bona fide infectious agent, having an infectious cycle, capable of transmitting horizontally (among a community) and causing epidemic outbreaks [3]. The discussion in this article will be focused on mammalian prion, and the term “prion" specifically refers to the infectious TSE agent.

Prion Is a Protein Conformation-Based Infectious Agent

Prions were defined as small proteinaceous infectious particles that cause TSEs [1], a group of fatal neurodegenerative diseases including Creutzfeldt-Jakob disease (CJD) in human, scrapie in sheep, bovine spongiform encephalopathy (BSE) in cattle, and chronic wasting disease (CWD) in deer and elk. Prion protein (PrP) is an N-linked glycoprotein tethered to lipid membranes via a glycophosphatidylinositol (GPI)-anchor, widely expressed in various tissues and highly enriched in the central nervous system. The host-encoded normal prion protein (PrPC) is α-helical rich, soluble in mild detergents, sensitive to protease digestion, and releasable from lipid membrane by phosphatidylinositol-specific phospholipase C (PI-PLC) digestion. During prion disease, a portion of PrP converts to an aberrant conformational isoform called PrPSc, which is mostly β-sheeted, highly aggregated, protease-resistant, and unable to be released from lipid membrane by PI-PLC digestion.

The prion hypothesis postulates that, because of its self-propagating property, PrPSc isoform seeds the conversion of normal PrPC to the pathogenic PrPSc and causes the disease. The self-propagating characteristic of disease-associated PrPSc has been demonstrated by cell-free conversion assay and serial protein misfolding cyclic amplification (sPMCA) [4], [5]. The simultaneous propagation of protease-resistant PrPSc and prion infectivity in sPMCA provides strong evidence supporting the prion hypothesis [5]. Because crude brain homogenate is used in sPMCA for prion propagation, it is difficult to conclusively establish a causal relationship between PrPSc and prion disease.

Clearly establishing that PrPSc, an altered conformational PrP isoform, is the causative agent for prion disease has been a challenge for the prion field for decades, mainly because PrPSc is highly aggregated, preventing it from being purified to homogeneity using conventional biochemical methods. An alternative approach is to reconstitute prion infectivity in vitro with defined components. Amyloid fibers formed in vitro with bacterially expressed recombinant PrP (recPrP) have been shown to contain limited prion infectivity [6][8]. Infectious prions have been formed through unseeded sPMCA with native PrPC purified from brains [9] or recPrP [10] as substrate, or by converting recPrP into an infectious conformer in sPMCA seeded with partially purified PrPSc [11]. More importantly, the infectivity of prions generated via sPMCA has been demonstrated by causing bona fide prion disease in wild-type animals [9], [10], [11]. Because bacterially expressed recPrP does not contain any eukaryotic genetic informational molecules, generating prion infectivity with recPrP [10][12] is generally accepted as the most stringent proof of prion hypothesis.

Cofactors Promote Prion Propagation

A key concept of the prion hypothesis is that prion is a self-propagating PrP conformer, which elicits the conversion of host-encoded normal PrPC to pathogenic PrPSc. This self-propagating property of PrP conformers is best demonstrated by in vitro assays. Recombinant PrP in amyloid fibers represents a self-propagating conformational state that is distinct from normal PrPC, but recPrP in amyloid fibers is generally without the biochemical hallmark of PrPSc, C-terminal proteinase K (PK)-resistance. The in vitro assays that propagate the classic PK-resistant PrPSc conformation include cell-free conversion assay and sPMCA [4], [5]. The cell-free conversion uses partially purified PrPSc as seed and PrPC purified from cultured cells as substrate, which is far less efficient than sPMCA that employs crude brain homogenates. The difference between two assays suggests the presence of factors in the brain homogenates that promote efficient prion propagation. Polyanions, such as RNA molecules and proteoglycans, have been identified as one type of cofactors in the brain homogenate that enhance prion propagation [13], [14]. Lipids are another type of cofactors that promote prion propagation in cell-free conversion assay [15] and in propagating recombinant prions via sPMCA [10].

Self-Propagating PrP Conformation Does Not Always Correlate to Prion Infectivity

Prion infectivity obviously depends on the seeding or self-propagating property of PrP conformers. However, a PrP conformer with seeding or self-propagating capability does not necessarily contain prion infectivity in vivo, that is, cause spongiform encephalopathy in animals. This notion is exemplified by PrP amyloid fiber, which clearly contains a strong seeding or self-propagating capability, but does not have a strong association with in vivo infectivity. The PrP amyloid fibers in the brain tissues of a familial prion disease patient induce a prominent PrP-amyloid accumulation in “humanized" knock-in mice carrying the same mutation, demonstrating in vivo seeding or self-propagating property of PrP amyloid fibers [16]. However, a lack of spongiform encephalopathy or any neurological disorders during the life span of these PrP amyloid–bearing mice suggests a dissociation of prion infectivity from seeding capability [16]. Similarly, in vitro generated recPrP amyloid fibers contain a strong in vitro seeding capability, but very low prion infectivity [6][8]. Even with PK-resistant PrP conformers, their abilities to propagate the PK-resistant conformation in sPMCA do not always associate with in vivo prion infectivity [17].

Collectively, the seeding or self-propagating property of a prion is essential for its infectivity, but not all PrP conformers with seeding or self-propagating capability are competent to cause prion disease in animals.

Cofactors Appear to Play an Important Role in Prion Infectivity

The role of cofactors in prion infectivity remains to be clarified, but comparing several in vitro reconstituted infectious prions indicates that they play an important role in prion infectivity. When infectious prions were formed with recPrP or purified PrPC via sPMCA in the presence of lipid and RNA as cofactors, the newly formed prions contain high infectivity [9], [10]. When recombinant prions were formed via seeded sPMCA in the absence of any mammalian cofactors, the prion infectivity is lower but still sufficient to cause disease in wild-type animals [11]. Notably, the buffer used in cofactor-free sPMCA contains SDS and Triton X-100 detergents, which are similar to lipid in biophysical properties and may partially replace the function of a lipid cofactor. The in vitro recPrP amyloid fiber formation, however, is in a partially denaturing buffer and completely without any cofactors. Interestingly, recPrP amyloid fibers contain very low infectivity, fail to cause disease in wild-type animals, and only cause disease with prolonged incubation times in PrP overexpressing mice [6], [7]. The limited infectivity of recPrP amyloid fibers in animals could be explained by binding to cofactors in vivo, leading to further PrP adaptations and conformational changes [18]. Nevertheless, the comparison of in vitro reconstituted prions indicates that the presence of cofactor enhances in vivo prion infectivity.

Mechanism of Cofactor in Prion Propagation and Infectivity

To date, two types of cofactors, lipids and polyanions, are identified as influencing prion propagation and infectivity, but the precise mechanism remains unclear. It can be envisioned that cofactors may act on several steps of PrP conversion. Both polyanion and lipid bindings alter the normal PrPC conformation to increase β-sheet content and PrP aggregation, a conformational change similar to PrPC-to-PrPSc conversion [19][21]. Thus, cofactor binding may render the normal PrPC susceptible to conversion or simply facilitate PrPSc-steered PrPC conversion. Alternatively, cofactors may facilitate PrP conversion by concentrating both PrPSc and PrPC on the surface of a single lipid vesicle or a single polyanion molecule. Another possibility could be that the cofactors stabilize the infectious PrPSc conformation by forming a complex with PrPSc and being a part of the infectious particle.

Both lipids and polyanions enhance prion propagation, yet polyanion does not appear to be critical for prion infectivity [22]. Lipids, on the other hand, not only influence prion propagation, but also have a significant impact on prion infectivity. A ∼100-fold increase in prion infectivity has been observed when purified prion is re-incorporated into liposomes [23]. Therefore, these two types of cofactors may act on the same or different steps in prion propagation and infectivity.

Most cofactor-related studies use mouse or hamster prions. It is well known that prions from different species or different prion strains have very different properties, which could be due to the different combinations of various lipid or polyanion molecules. It is also possible that other types of molecules could serve as cofactors, contributing to the diverse biological properties of prions. Further investigation of prion cofactors will help us to gain the insights of this enigmatic infectious agent, which is essential for us to combat these devastating diseases.


1. PrusinerSB 1982 Novel proteinaceous infectious particles cause scrapie. Science 216 136 144

2. TuiteMFMarchanteRKushnirovV 2011 Fungal prions: structure, function and propagation. Top Curr Chem 305 257 298

3. AguzziA 2009 Cell biology: Beyond the prion principle. Nature 459 924 925

4. KociskoDAComeJHPriolaSAChesebroBRaymondGJ 1994 Cell-free formation of protease-resistant prion protein. Nature 370 471 474

5. CastillaJSaaPHetzCSotoC 2005 In vitro generation of infectious scrapie prions. Cell 121 195 206

6. LegnameGBaskakovIVNguyenHORiesnerDCohenFE 2004 Synthetic mammalian prions. Science 305 673 676

7. ColbyDWWainRBaskakovIVLegnameGPalmerCG 2010 Protease-sensitive synthetic prions. PLoS Pathog 6 e1000736 doi:10.1371/journal.ppat.1000736

8. MakaravaNKovacsGGBocharovaOSavtchenkoRAlexeevaI 2010 Recombinant prion protein induces a new transmissible prion disease in wild-type animals. Acta Neuropathol 119 177 187

9. DeleaultNRHarrisBTReesJRSupattaponeS 2007 Formation of native prions from minimal components in vitro. Proc Natl Acad Sci U S A 104 9741 9746

10. WangFWangXYuanCGMaJ 2010 Generating a prion with bacterially expressed recombinant prion protein. Science 327 1132 1135

11. KimJICaliISurewiczKKongQRaymondGJ 2010 Mammalian prions generated from bacterially expressed prion protein in the absence of any mammalian cofactors. J Biol Chem 285 14083 14087

12. WangFZhangZWangXLiJZhaL 2012 Genetic informational RNA is not required for recombinant prion infectivity. J Virol 86 1874 1876

13. DeleaultNRLucassenRWSupattaponeS 2003 RNA molecules stimulate prion protein conversion. Nature 425 717 720

14. DeleaultNRGeogheganJCNishinaKKascsakRWilliamsonRA 2005 Protease-resistant prion protein amplification reconstituted with partially purified substrates and synthetic polyanions. J Biol Chem 280 26873 26879

15. BaronGSCaugheyB 2003 Effect of glycosylphosphatidylinositol anchor-dependent and -independent prion protein association with model raft membranes on conversion to the protease-resistant isoform. J Biol Chem 278 14883 14892

16. PiccardoPMansonJCKingDGhettiBBarronRM 2007 Accumulation of prion protein in the brain that is not associated with transmissible disease. Proc Natl Acad Sci U S A 104 4712 4717

17. MillerMBGeogheganJCSupattaponeS 2011 Dissociation of infectivity from seeding ability in prions with alternate docking mechanism. PLoS Pathog 7 e1002128 doi:10.1371/journal.ppat.1002128

18. MakaravaNKovacsGGSavtchenkoRAlexeevaIBudkaH 2011 Genesis of mammalian prions: from non-infectious amyloid fibrils to a transmissible prion disease. PLoS Pathog 7 e1002419 doi:10.1371/journal.ppat.1002419

19. WongCXiongLWHoriuchiMRaymondLWehrlyK 2001 Sulfated glycans and elevated temperature stimulate PrP(Sc)-dependent cell-free formation of protease-resistant prion protein. Embo J 20 377 386

20. GomesMPMillenTAFerreiraPSe SilvaNLVieiraTC 2008 Prion protein complexed to N2a cellular RNAs through its N-terminal domain forms aggregates and is toxic to murine neuroblastoma cells. J Biol Chem 283 19616 19625

21. WangFYangFHuYWangXJinC 2007 Lipid interaction converts prion protein to a PrPSc-like proteinase K-resistant conformation under physiological conditions. Biochemistry 46 7045 7053

22. PiroJRHarrisBTSupattaponeS 2011 In situ photodegradation of incorporated polyanion does not alter prion infectivity. PLoS Pathog 7 e1002001 doi:10.1371/journal.ppat.1002001

23. GabizonRMcKinleyMPGrothDFKenagaLPrusinerSB 1988 Properties of scrapie prion protein liposomes. J Biol Chem 263 4950 4955

Hygiena a epidemiologie Infekční lékařství Laboratoř

Článek vyšel v časopise

PLOS Pathogens

2012 Číslo 4
Nejčtenější tento týden
Nejčtenější v tomto čísle

Zvyšte si kvalifikaci online z pohodlí domova

Důležitost adherence při depresivním onemocnění
nový kurz
Autoři: MUDr. Eliška Bartečková, Ph.D.

Koncepce osteologické péče pro gynekology a praktické lékaře
Autoři: MUDr. František Šenk

Sekvenční léčba schizofrenie
Autoři: MUDr. Jana Hořínková, Ph.D.

Hypertenze a hypercholesterolémie – synergický efekt léčby
Autoři: prof. MUDr. Hana Rosolová, DrSc.

Multidisciplinární zkušenosti u pacientů s diabetem
Autoři: Prof. MUDr. Martin Haluzík, DrSc., prof. MUDr. Vojtěch Melenovský, CSc., prof. MUDr. Vladimír Tesař, DrSc.

Všechny kurzy
Zapomenuté heslo

Zadejte e-mailovou adresu, se kterou jste vytvářel(a) účet, budou Vám na ni zaslány informace k nastavení nového hesla.


Nemáte účet?  Registrujte se